Microorganisms doi: 10.3390/microorganisms14091963

Authors: Fang Liang Tianyuan Nie Peixiu Lin Huawei Li Ruining Wang Xuyong Zhao Lin Wang Keshan Zhang Yaqiong Ye Mengmeng Zhao

Porcine reproductive and respiratory syndrome virus (PRRSV) poses a significant threat to the global swine industry. In this study, the genetic diversity, evolutionary dynamics, and key antigenic characteristics of the PRRSV-1 GP5 gene in China were systematically investigated. A total of 114 Chinese PRRSV-1 GP5 sequences, isolated between 1999 and 2024, were analyzed alongside global reference strains. The nucleotide and amino acid sequence similarities ranged from 60.03% to 100% and from 48.72% to 100%, respectively, with mutation and deletion hotspots concentrated in the N-terminal hypervariable region. Phylogenetic analysis further classified the Chinese strains into four major lineages: Amervac-like, BJEU06-1-like, HKEU16-like, and NMEU09-1-like. Notably, a preliminary phylogenetic observation indicated that several recent Chinese PRRSV-1 isolates clustered topologically close to PRRSV-2 strains. However, as this finding is based solely on a single-gene tree and has not been validated by formal tests for convergent evolution, homoplasy, or selection pressure, it should be interpreted with caution and warrants further investigation using whole-genome data. A Bayesian evolutionary analysis estimated the evolutionary rate at 1.94 × 10−3 substitutions per site per year and the time to the most recent common ancestor at approximately 1910. No statistically significant recombination events were identified within the GP5 gene. The transmembrane topology of GP5 was highly conserved, whereas B-cell linear epitopes were enriched in the N-terminal extracellular region. This study establishes a useful molecular foundation for understanding PRRSV-1 evolution in China and offers valuable insights for the development of broad-spectrum vaccines and effective control strategies.

Microorganisms doi: 10.3390/microorganisms14091961

Authors: Xueliang Zhan Yushan Pei Yuanyuan Huang Na Li Wenhui Huang Yitong Tang Haihua Liang

How individual diguanylate cyclases generate specific outputs within bacterial c-di-GMP networks remains unclear. Here, we characterized PA0290, a PAS-PAC-GGDEF protein of Pseudomonas aeruginosa, and examined its relationship with the c-di-GMP receptor PelD. Deletion or overexpression of PA0290 alone did not significantly affect biofilm formation. A bacterial adenylate cyclase two-hybrid screen identified PelD as a candidate PA0290-interacting protein, and co-expression of PA0290 and PelD markedly enhanced static and flow-cell biofilm formation. Purified PA0290 generated an HPLC product peak with a retention time closely matching that of the authentic c-di-GMP standard, whereas substitution of the conserved GGEEF motif with GGAAF reduced product formation and weakened the biofilm-enhancing phenotype observed upon PA0290–PelD co-expression. PA0290–PelD co-expression did not produce sustained activation of the bulk c-di-GMP-responsive cdrA-lux reporter. Clinical isolates also displayed heterogeneous biofilm-forming capacity and variable PA0290 and pelD transcript abundance. Together, these findings support a functional association between PA0290 and PelD in biofilm regulation. The absence of sustained bulk c-di-GMP-responsive reporter activation suggests that this phenotype is not accompanied by a generalized increase in c-di-GMP-responsive transcription.

Microorganisms doi: 10.3390/microorganisms14091960

Authors: Qiuyue Li Jiaming Wang Zhilong Wang Chenxu Cheng Shuting Bao Shatuo Chai Dongwen Dai Xun Wang Qizhu Song Yongwei Chen Jiaying Lv Yijuan Ma Tashi Sonam Junchao Qiu Shuxiang Wang

The yak is an iconic ruminant of the Qinghai-Tibet Plateau, yet segment-specific variation in its intestinal microbial functional potential and bile acid profiles under different feeding systems remains insufficiently characterized. Six healthy adult male yaks with similar body weights (320 ± 30 kg) were assigned to grazing (G) or stall-feeding (S) systems, with three animals per group, for a 90-day trial comprising a 10-day adaptation period and an 80-day formal experimental period. The individual yak was considered the experimental unit, and intestinal segments sampled from the same animal were treated as repeated observations. Liver tissue and digesta from the duodenum, ileum, cecum, and colon were analyzed using targeted bile acid metabolomics and shotgun metagenomics. Principal coordinate analysis based on Bray-Curtis dissimilarities showed segment-associated clustering of microbial communities, with PCo1 and PCo2 explaining 65.5% and 18.9% of the total variation, respectively. ANOSIM identified a significant intestinal-segment effect on microbial community composition (R = 0.2208, BH-FDR = 0.0144), whereas the overall feeding-system effect was not significant (R = 0.3747, BH-FDR = 0.1200). No statistically significant feeding-system differences were detected in Shannon, Simpson, Chao1, or ACE indices within individual intestinal segments (BH-FDR ≥ 0.800), and PERMDISP detected no significant differences in within-group dispersion (BH-FDR ≥ 0.1682). Bacillota and Bacteroidota were the dominant phyla. Descriptive functional profiling showed higher mean ileal abundances of GH2 (0.0035 vs. 0.0026), GH3 (0.0029 vs. 0.0024), and GH43 (0.0021 vs. 0.0013) in grazing yaks, whereas the starch-associated GH13 family showed its highest mean abundance in the colon of stall-fed yaks. These metagenomic patterns represent predicted genomic functional potential rather than gene expression, enzyme activity, or metabolic flux. Cecal total bile acid concentration showed a nominal between-group difference (unadjusted Welch’s p = 0.0109), but this difference did not remain significant after correction across the five anatomical sites (BH-FDR = 0.0545). In the colon, stall-fed yaks had a lower conjugated-to-unconjugated bile acid ratio and a higher secondary-to-primary bile acid ratio than grazing yaks (BH-FDR < 0.05). Feeding-system-associated descriptive patterns were observed in predicted microbial functional profiles, whereas statistically supported between-group differences were limited mainly to selected colonic bile acid ratios. Given the limited animal-level replication, these findings should be considered exploratory.

Microorganisms doi: 10.3390/microorganisms14091959

Authors: Matteo Fortunato Anne Tabard-Fougère Elio Paris Giacomo De Marco Oscar Vazquez Christina Steiger Ardian Ramadani Andreas Tsoupras Romain Dayer Dimitri Ceroni

Identifying the causative pathogen of paediatric osteoarticular infections (OAIs) is essential for guiding management. Kingella kingae infections are generally milder than pyogenic infections, which may have more severe and lasting consequences. Because standard inflammatory markers have limited discriminatory value, we assessed whether complete blood count-derived immune–inflammatory biomarkers (IIBs) could provide a simple, low-cost diagnostic aid. We retrospectively reviewed children younger than 5 years admitted to a tertiary hospital with confirmed OAIs. K. kingae cases were included from 2007 to 2025 and pyogenic cases from 1997 to 2025. Admission clinical data, conventional inflammatory markers, and six IIBs—NLR, MLR, PLR, SII, SIRI, and PIV—were analysed. The cohort included 118 K. kingae and 39 pyogenic OAIs. Pyogenic infections were associated with higher admission temperature and CRP levels and lower lymphocyte counts. All six IIBs were significantly increased in pyogenic infections and showed excellent univariable discrimination, with AUCs of 0.92–0.94. In a common complete-case cohort, the clinical model incorporating age, admission temperature, and CRP achieved an optimism-corrected AUC of 0.848. Adding lymphocyte count or an individual IIB increased corrected AUCs to 0.951–0.962; apparent AUCs were significantly higher than that of the clinical model after Holm adjustment. The abnormal-IIB-count model achieved the highest corrected AUC of 0.970; apparent sensitivity and specificity at the Youden-optimal threshold were 97.3% and 90.3%, respectively, but it did not significantly outperform the model with lymphocyte count alone. Routine CBC-derived IIBs may improve early differentiation between K. kingae and pyogenic OAIs, although external validation is required before clinical implementation.

Microorganisms doi: 10.3390/microorganisms14091958

Authors: Jia-You Xing Wen-Jie Xu Zi-Xuan Fu Ying-Na Xu Jing-Yang Li Jiang Wang Bei-Bei Chu Lei Zeng Sheng-Li Ming

Feline calicivirus (FCV) is a highly heterogeneous pathogen and a major cause of feline upper respiratory tract disease, highlighting the need for continuous surveillance of its genetic diversity and pathogenic characteristics. In this study, we isolated and comprehensively characterized a newly isolated FCV strain, HN/ZZ/2025, from cats at a feline trading market in Zhengzhou, China, and systematically evaluated its molecular features, in vitro replication characteristics, physicochemical properties, and pathogenicity in cats. Phylogenetic analysis classified HN/ZZ/2025 as genotype GI. The isolate replicated efficiently in CRFK, F81, and Fc3Tg cells, reaching peak titers of 107.18, 107.50, and 105.89 TCID50/0.1 mL, respectively, and exhibited typical calicivirus-like particles with diameters of 35–40 nm. Complete genome analysis showed that HN/ZZ/2025 shared the highest nucleotide and amino acid sequence identities of 85.7% and 62.8%, respectively, with the closely related Chinese FCV strain CH-JL4, and revealed distinct amino acid variations within the hypervariable E region of the VP1 capsid protein. Experimental infection of cats (n = 3/group) resulted in pyrexia, with rectal temperatures reaching 39.8–40.2 °C, weight loss, oral ulceration, and persistent viral RNA shedding from 1 to 21 days post-infection. Viral RNA and VP1 antigen were detected in the lungs, trachea, kidneys, and spleen, indicating systemic dissemination and broad tissue distribution. Although no mortality occurred during the observation period, the observed clinical signs, viral dissemination, and histopathological lesions indicate that HN/ZZ/2025 is capable of causing clinically evident disease in experimentally infected cats. These findings provide useful insights into the molecular epidemiology, tissue tropism, and host–virus interactions of circulating FCV strains and establish HN/ZZ/2025 as a useful isolate for future studies of FCV genetic diversity and pathogenesis.

Microorganisms doi: 10.3390/microorganisms14091957

Authors: Xinyi Chen Sijin Guo Xiaokai Zheng Yingchao Sun Rongzheng Huang Yongcheng Chen Chunhui Ma Fanfan Zhang

Paper mulberry (Broussonetia papyrifera) is a high-protein forage, but its high buffering capacity and low water-soluble carbohydrate (WSC) content hinder silage fermentation. This study evaluated the effect of tea tree (Melaleuca alternifolia) essential oil (TTO) as a natural additive for paper mulberry silage. TTO was added at 0 (CK), 500 (CSD), and 1000 mg/kg (CSG) fresh weight, with samples collected on days 7, 15, 30, and 90. After 90 days, CSD had the lowest pH and the highest lactic acid content. NH3-N/TN was significantly lower than in CK (p < 0.05), and the retention of dry matter, crude protein and water-soluble carbohydrate was improved (p < 0.05). Compared with CK, CSD exhibited the highest lactic acid bacteria (LAB) counts, along with the lowest mold and aerobic bacterial counts (p < 0.05). However, CSG did not further improve fermentation quality, and most of the indicators were not significantly different from CSD (p > 0.05). The in vitro gas production test showed that CSD significantly increased the theoretical maximum gas production and the maximum gas production rate (p < 0.05). Additionally, 16S rRNA sequencing revealed a distinct bacterial succession in CSD, which enriched beneficial lactic acid bacteria (LAB) such as Pediococcus while outcompeting less competitive taxa. This microbial reshaping redirected the community toward acidification and nutrient retention. Collectively, these results demonstrate that TTO at 500 mg/kg is a promising natural additive for improving paper mulberry silage quality through selective reshaping of the bacterial community.

Microorganisms doi: 10.3390/microorganisms14091956

Authors: Fahima Chowdhury Marjahan Akhtar Firdausi Qadri Taufiqur Rahman Bhuiyan

Enterotoxigenic Escherichia coli (ETEC) is a leading cause of bacterial diarrheal illness in low- and middle-income countries and among travelers to endemic regions, impacting morbidity and mortality and impairing childhood development. Despite its significant importance in public health, the propensity of ETEC diarrhea in endemic settings is influenced by complex interactions among epidemiological, immunological, environmental, and pathogen-related factors. This review aims to provide a comprehensive understanding of ETEC diarrhea in endemic regions by examining epidemiological trends, host immune responses, environmental determinants, and progress in current ETEC vaccine development. We discuss disease burden, seasonality, age-specific susceptibility, toxin and colonization factor diversity, and their relation to diarrheal disease severity at the community level and in hospitalized patients. In addition, we summarize current knowledge on natural immunity, including mucosal and systemic immune responses induced in ETEC-infected diarrheal patients. The review further highlights advances in ETEC vaccine development, focusing on candidate vaccines, safety, immunogenicity, protective efficacy, and challenges associated with implementation in resource-limited settings. By integrating clinical, microbiological, immunological, environmental, and public health perspectives, this review provides a multisectoral framework to better understand ETEC pathogenesis and prevention. It also highlights future research priorities, vaccine strategies, and policy interventions to reduce the burden of ETEC-associated diarrhea in endemic settings.

Microorganisms doi: 10.3390/microorganisms14091955

Authors: Esra Çiftci Cüneyt Özakın Sinem İrez Çetin Oktay Rodoplu Zeynep Gizem Ergün Özdel Nazmiye Ülkü Tüzemen Pelin Laleoğlu Deniz Camcı Erten Solmaz Çelebi Mustafa Kemal Hacımustafaoğlu

In the original publication [...]

Microorganisms doi: 10.3390/microorganisms14091954

Authors: Nilufar Akhmedova Gulomjon Kholov Ulugbek Ochilov Nodira Shonazarova Otabek Yuldashev Gulkhayo Olimova Sabina Istamova Sitora Mukhammadiyeva Nozima Kenjayeva Jahongir Sharipov

Pulmonary hypertension complicating chronic obstructive pulmonary disease (COPD-PH) has a worse prognosis than airflow limitation alone but its pathogenesis is thought to be primarily due to alveolar hypoxia affecting pulmonary circulation. Two observations are not fully explained by this model. First, the majority of exacerbations of COPD are infectious and there is a stable frequent-exacerbator phenotype that is not fully accounted for by spirometric severity. Second, the airways of COPD are not sterile. They are home to a resident microbial community that changes as the disease progresses and the gut microbial community also undergoes disease-associated changes. This review examines whether microbial factors actively contribute to pulmonary vascular remodelling or merely reflect advanced disease. Three candidate exposures are considered: chronic airway colonisation, dysbiosis of the airway and gut communities and recurrent infectious exacerbation. We then examine the virulence mechanisms and host-recognition pathways of non-typeable Haemophilus influenzae, Streptococcus pneumoniae, Moraxella catarrhalis, Pseudomonas aeruginosa, rhinovirus, influenza viruses, respiratory syncytial virus and SARS-CoV-2. We then trace the pathways linking microbial recognition to vascular injury: Toll-like receptor signalling and NF-κB activation; interleukin-6, tumour necrosis factor-α and interleukin-1β; endothelin-1 upregulation against nitric oxide depletion; reactive oxygen species and hypoxia-inducible factor signalling converging with hypoxic pulmonary vasoconstriction; and, along the gut–lung axis, lipopolysaccharide translocation, trimethylamine N-oxide and depletion of short-chain fatty acid-producing taxa. Therapeutically, there is evidence and a vascular rationale for interventions that decrease exacerbation frequency. Pulmonary vasodilators have a sound physiological rationale, although their clinical efficacy is frequently limited by worsening ventilation–perfusion mismatch. Current evidence supports biological plausibility rather than demonstrated causality: no direct longitudinal human study has established that microbial alterations or microbial exposure cause pulmonary vascular remodelling in COPD-PH. We identify study designs capable of directly testing this hypothesis.

Microorganisms doi: 10.3390/microorganisms14091953

Authors: Lakshmi T. Sunkara John Reddy Peasari Diane Davis Radhika Kakani Tanit Kasantikul Rob Harbert Maurice Byukusenge

The family Pasteurellaceae includes pathogenic and opportunistic bacteria affecting poultry, yet taxonomic resolution remains challenging due to phenotypic heterogeneity, limited genomic representation, and the poor performance of routine diagnostic systems for uncommon taxa. At the Clemson Veterinary Diagnostic Center, a Gram-negative bacterium was isolated from a backyard hen. The isolate produced two distinct colony morphotypes with identical biochemical profiles. They were misidentified as Sphingomonas paucimobilis by VITEK® 2, while MALDI-TOF MS failed to identify them. Oxford Nanopore long-read sequencing with PacBio HiFi polishing generated near-complete chromosome-level assemblies. Comparative genomic analyses demonstrated that morphotypes CVDC-smooth and CVDC-rough represent clonal variants of a distinct Spirabiliibacterium lineage sharing 93% average nucleotide identity (ANI) and 52% digital DNA–DNA hybridization (dDDH) with the closest relative, Spirabiliibacterium mucosae. Copy-number variation within a tandemly duplicated ~20 kb genomic region may contribute to differences in colony morphology. Family-wide analysis identified lineage-associated gene patterns and a limited number of VFDB-matched virulence genes in the CVDC isolates. This study identifies a distinct lineage within the genus Spirabiliibacterium, reports the first isolation and chromosome-level genome assemblies of a Spirabiliibacterium lineage from a chicken in the United States, and highlights the limitations of routine diagnostic methods for identifying uncommon bacterial taxa.

Microorganisms doi: 10.3390/microorganisms14091952

Authors: Pietro Crispino Antonello Viceconti Valentina Camardo

During the COVID-19 pandemic, healthcare-associated infections (HAIs) showed a significant increase, exacerbated by the prevalence of multidrug-resistant (MDR) organisms. Antibiotic overuse and infection control practices influenced HAI incidence rates, alongside the rising prevalence of resistant pathogens such as methicillin-resistant Staphylococcus aureus. Antibiotic-resistant pathogens causing healthcare-associated infections in COVID-19 patients include—in addition to methicillin-resistant Staphylococcus aureus—metallo-β-lactamase-producing carbapenem-resistant Enterobacteriaceae, carbapenem-resistant Acinetobacter baumannii, extended-spectrum β-lactamase-producing Klebsiella pneumoniae, and vancomycin-resistant enterococci. COVID-19 impacted bacterial healthcare-associated infections in various ways, with an increase in the incidence of metallo-β-lactamase-producing, carbapenem-resistant organisms—a trend already noted prior to the pandemic. Furthermore, the pandemic laid the groundwork for future challenges in HAI management, highlighting the need for rigorous infection prevention and control protocols. Poorer outcomes were observed in hospitalized COVID-19 patients with antibiotic-resistant infections. Although increased infection prevention and control (IPC) measures led to a reduction in certain site-specific hospital-acquired infections, in other contexts, COVID-19 was associated with an increased incidence of bacterial hospital-acquired infections. Further research is needed to determine the cost–benefit ratio of maintaining COVID-19-related infection prevention and control protocols beyond the pandemic in order to reduce the impact of hospital-acquired infections. Furthermore, it is necessary to assess the long-term impact of the high usage of certain broad-spectrum antibiotics during the COVID-19 pandemic.

Microorganisms doi: 10.3390/microorganisms14091951

Authors: Victoria Birlutiu Rares-Mircea Birlutiu

Antimicrobial resistance is now documented in a substantial proportion of laboratory-confirmed bacterial infections worldwide, and although the most extreme extensively drug-resistant (XDR) and pandrug-resistant (PDR) phenotypes are concentrated among Gram-negative bacilli, resistance in Gram-positive cocci continues to rise and remains a major determinant of outcomes in severe infection. This structured comprehensive narrative review, based on a documented search of PubMed/MEDLINE, Scopus and Web of Science to June 2026, is confined to the three Gram-positive cocci of greatest clinical importance—Staphylococcus aureus, Streptococcus pneumoniae and Enterococcus faecium—and summarises current global and European surveillance data for this organisms, describes the principal molecular mechanisms responsible for resistance to β-lactams, glycopeptides, fluoroquinolones, oxazolidinones, lipopeptides, tetracyclines, folate-pathway inhibitors, macrolides–lincosamides–streptogramins and aminoglycosides, and reviews the therapeutic options and investigational agents that may extend the treatment repertoire against multidrug-resistant (MDR) Gram-positive cocci. Surveillance indicators for the three species are compared side by side, and the agents discussed are separated according to whether they are licensed, clinically evaluated but not widely licensed, or still preclinical. Its scope is deliberately narrower than that of pan-bacterial accounts of antimicrobial resistance: it concentrates on the determinants that routine molecular confirmation does not detect, on the divergent pathogen-specific trajectories visible in recent surveillance, and on the distinction between agents supported by randomised evidence and those still at the discovery stage. Our review is structured rather than systematic; the PRISMA 2020 flow diagram is used as a reporting template only, and no claim of PRISMA compliance is made.

Microorganisms doi: 10.3390/microorganisms14091950

Authors: Yiping Guo Meng Bai Yan Liu Leicheng Li Shihang Ni Camilo Ayra-Pardo Wei Yuan

The widespread occurrence of antibiotics in aquatic environments has raised concern over the dissemination of antibiotic resistance genes (ARGs). Although vertical flow constructed wetlands (VFCWs) efficiently remove antibiotics from wastewater, the relationships between substrate configuration, microbial communities, and ARG abundance remain poorly understood. Here, five laboratory-scale VFCWs with different substrate configurations, including cobalt ferrite (CoFe2O4)-modified biochar and its composite with zeolite, were evaluated using synthetic domestic wastewater amended with oxytetracycline (OTC). CoFe2O4 modification increased the specific surface area and pore volume of biochar and altered its adsorption behavior. All VFCWs achieved high OTC removal (>97%), despite differences in OTC accumulation within the substrate. In contrast, ARG and mobile genetic element abundances and bacterial community profiles varied among reactors. Combined tetA and tetX abundance ranged from 2.55 × 102 to 1.90 × 105 copies g−1, with the CoFe2O4-modified biochar–zeolite reactor showing the lowest mean ARG abundance. Exploratory co-occurrence analysis identified associations of Acinetobacter with tetA and Flavobacterium with tetA and tetX (nominal Spearman p < 0.05), although these correlations do not establish ARG host identity or horizontal gene transfer. Overall, the findings indicate that microbial ecological patterns associated with ARG abundance may differ among VFCWs even when antibiotic removal efficiencies are comparable.

Microorganisms doi: 10.3390/microorganisms14091949

Authors: Yu-Xiang Zhang Li-Xian Wang Jian-Gen Zhang Chen Li Luo-Fa Wu Xin-Feng Jiang Ye Chun

Tea quality is largely determined by the accumulation of specialized metabolites in fresh leaves, yet the effects of the propagation method on tea quality and its belowground ecological basis remain insufficiently understood. In this study, sexually propagated (SR) and asexually propagated (AR) tea plants were compared by integrating soil physicochemical analysis, leaf quality and physiological measurements, widely targeted metabolomics, and rhizosphere metagenomic profiling. Compared with AR, SR plants exhibited more favorable rhizosphere nutrient conditions, with soil organic matter, total nitrogen, alkali-hydrolyzable nitrogen, and available phosphorus increasing by 23.1%, 18.2%, 27.8%, and 161.5%, respectively, although available potassium decreased by 24.4%. SR leaves also contained higher dry matter, tea polyphenol, and soluble sugar contents, which increased by 11.5%, 58.8%, and 8.6%, respectively. In addition, superoxide dismutase, peroxidase, and indole-3-acetic acid oxidase activities were 30.4%, 92.0%, and 21.7% higher under SR, whereas hydrogen peroxide content remained unchanged. Metabolomic profiling revealed marked differences in leaf metabolic composition between the two propagation types, with differentially accumulated metabolites mainly enriched in flavonoid biosynthesis, phenolic acid metabolism, caffeine metabolism, carotenoid biosynthesis, plant hormone signaling, and α-linolenic acid metabolism. Rhizosphere metagenomic analysis further showed that SR was characterized by higher relative abundances of Actinomycetota, Pseudomonadota, Planctomycetota, Alphaproteobacteria, and Streptomycetales, together with distinct microbial functional profiles related to glycolysis, the tricarboxylic acid cycle, and pyruvate metabolism. Significant correlations were identified between several SR-enriched microbial taxa and quality-related metabolites, particularly flavonoids and phenolic acids. Overall, under the present field conditions, sexual propagation was more favorable than asexual propagation for tea quality formation, as reflected by improved nitrogen and phosphorus availability, greater accumulation of quality-related metabolites, higher antioxidant enzyme activities, and distinct rhizosphere microbial carbon-metabolic potential. These findings provide an integrated soil–microbiome–metabolome perspective for understanding propagation-related differences in tea quality.

Microorganisms doi: 10.3390/microorganisms14091948

Authors: Daniela Sateriale Giuseppina Forgione Paola Salvatore Caterina Pagliarulo

Pomegranate is increasingly being recognized as a versatile source of bioactive compounds with antimicrobial, antioxidant, anti-inflammatory, and microbiota-modulating potential. Beyond the edible arils and juice, peel, seeds, leaves, flowers, and other agro-industrial by-products are rich in ellagitannins, particularly punicalagins, as well as ellagic acid, gallic acid, flavonoids, anthocyanins, fatty acids, and related phytochemicals. Their recovery depends strongly on plant fraction, cultivar, solvent, and extraction technology, including conventional hydroalcoholic extraction, ultrasound- and microwave-assisted processes, high-pressure treatments, and enzyme-assisted methods. Pomegranate-derived preparations exhibit activity against Gram-positive and Gram-negative bacteria, fungi, multidrug-resistant pathogens, and microbial biofilms, while selected compounds may enhance the efficacy of conventional antimicrobials. Emerging evidence also indicates bidirectional interactions with microbial communities, including microbial biotransformation of ellagitannins into urolithins and modulation of beneficial taxa and microbial metabolites. This narrative review integrates agronomic and phytochemical diversity, extraction and standardization strategies, mechanisms of antimicrobial action, synergistic interactions, microbiota-related effects, and applications in food preservation, biomedicine, animal nutrition, agriculture, and environmental sustainability. Key translational limitations include compositional variability, methodological heterogeneity, insufficient standardization, limited in vivo validation, and scarce evidence from realistic application models and clinical studies. Addressing these gaps is essential for developing safe, reproducible, and scalable pomegranate-derived preparations.

Microorganisms doi: 10.3390/microorganisms14091947

Authors: Cristoper Ramírez-Sandoval María-Elena Campos-Aldrete Benjamín Nogueda-Torres Rogelio Gómez-Escobedo

Chagas disease remains a major neglected tropical disease with limited therapeutic options and substantial drug-associated toxicity. A previously synthesized series of 1-(2-methyl-5-nitro-1H-imidazol-1-yl)-2-phenyl-N-(4-arylthiazol-2-yl)ethan-1-imines (5a–e) was evaluated for trypanocidal activity. Molecular docking against Trypanosoma cruzi trypanothione reductase (PDB ID: 1BZL) suggested favourable molecular recognition for several derivatives. Compound 5e, bearing a p-nitro-substituted arylthiazole moiety, showed the most favourable predicted binding profile, with a binding energy of −8.41 kcal/mol and an estimated inhibition constant of 0.689 µM. Biological activity was assessed against bloodstream trypomastigotes of two T. cruzi isolates representing acute and chronic Chagas disease models, NINOA and INC-5, respectively; however, both isolates were evaluated during acute experimental infection in mice. Compound 5e exhibited the strongest trypanocidal activity, with IC50 values of 18.08 and 22.41 µM against NINOA and INC-5, respectively, outperforming nifurtimox and benznidazole under the same experimental conditions. In addition, 5e exhibited low cytotoxicity in BHK-21 cells (CC50 = 2230 µM), yielding selectivity indices of 123 for NINOA and 99.6 for INC-5. Substitution at the para position of the arylthiazole ring markedly influenced trypanocidal activity and predicted molecular recognition. Overall, these findings identify the thiazolyl–nitroimidazole scaffold, particularly compound 5e, as a promising platform for further trypanocidal and mechanistic evaluation.

Microorganisms doi: 10.3390/microorganisms14091946

Authors: Vladimir Paredes-Cervantes Cecilia Rosel-Pech Sandra Angélica Rojas-Osornio Edith Reyes-Serrato Laura López-Pelcastre Leticia Manuel-Apolinar Laura Arcelia Montiel-Cervantes José Molina-López María Pilar Cruz-Domínguez José Guadalupe Rendón-Maldonado Fernando Minauro-Sanmiguel Martha Eugenia Ruiz-Tachiquín Salvador Vázquez-Vega Emiliano Tesoro-Cruz

Antimicrobial resistance poses a critical challenge to global public health. Urinary tract infections are one of the leading causes of morbidity in Mexico. This study aimed to identify the most prevalent bacterial pathogens in urinary bacterial isolates and to determine their antimicrobial and selected multidrug-resistance pattern profiles in outpatient and inpatient isolates at a tertiary care hospital in Mexico City. In this retrospective laboratory-based surveillance study, a census of 3434 urine samples collected between January and December 2023 was analyzed. Bacterial identification and antimicrobial susceptibility testing were performed using a VITEK 2 XL automated system. Escherichia coli was the predominant uropathogen in both groups, followed by Enterococcus spp. and Klebsiella spp. A significant disparity was observed in the resistance profiles: E. coli resistance to third-generation cephalosporins and fluoroquinolones was higher in inpatient isolates than in outpatient isolates. The selected multidrug-resistance patterns were generally higher in the inpatient isolates than in the outpatient isolates. Although common enteric pathogens with lower resistance levels predominated in the outpatient isolates, inpatient isolates required coverage targeting higher resistance. These findings reflect the need to tailor the treatment to the patients’ clinical context, launch awareness campaigns for the strategic use of antibiotics, and strengthen epidemiological surveillance.

Microorganisms doi: 10.3390/microorganisms14091945

Authors: Tewodros Fentahun Jember Mark Edward Westman Sameer Dinkar Pant Seyed Ali Ghorashi

Bovine bacterial mastitis remains a major challenge for the dairy industry, and timely identification of causative bacteria is essential for effective disease management. This study compared six regions of the 16S rRNA gene for PCR high-resolution melt (HRM) curve analysis to identify the most suitable target for species-level differentiation of mastitis-associated bacteria. A total of 547 clinical bovine milk samples from cows with clinical mastitis were cultured, and representative isolates of 18 bacterial species were selected for comparative analysis. Species identity was confirmed using matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) prior to molecular analysis. All six primer pair combinations were tested across the 18 species panel, and discriminatory performance was assessed using a genotype confidence percentage (GCP)-based classification model that allowed for objective interpretation without visual inspection. Among the six target regions, the 203 bp amplicon generated by primer pair F1R1, spanning the V1-to-V2 region and part of V3, showed the strongest overall discriminatory performance with no cross-classification observed. Sequencing of F1R1 amplicons confirmed species identity and showed concordance with HRM-based classification. Phylogenetic analysis of sequences from this region showed clustering broadly consistent with established taxonomic relationships. Proof-of-concept testing showed that the workflow could also be applied to DNA extracted directly from selected milk samples without prior bacterial isolation. Overall, this approach provides a rapid, cost-effective, and high-throughput method for preliminary discrimination of mastitis-associated bacteria.

Microorganisms doi: 10.3390/microorganisms14091944

Authors: Lorreta Aboagyewa Opoku Stephanie V. Trefry Maame Konadu Jonathan Ontivero Sanchez Alison Gomeiz Shannon D. Walls Michael D. Barrera Dylan Valerio Scarton Rémi Veneziano Mariaelena Pierobon Elsa Ronzier Aarthi Narayanan

Arthropod-borne viruses such as Alphaviruses and Flaviviruses are the causative agents of severe human disease, highlighted by fatal encephalitis and neurological sequelae in survivors. The lack of FDA-approved vaccines and therapeutics that can prevent or treat these infections results in a significant global disease burden. An important unmet need to address this capability gap is the need for affordable, scalable, clinically relevant human-based neuronal models to study neuroinvasive viruses and evaluate therapeutic options. Here, we described the application of a human neuronal precursor cell model, LUHMES (Lund human mesencephalic) cells, that can be differentiated into dopaminergic midbrain neurons and used to study virus infections. In this study, we demonstrated the susceptibility of LUHMES cells to infection by three arthropod-borne neurotropic viruses: Venezuelan equine encephalitis virus, dengue virus serotype 2, and West Nile virus. We also demonstrated how the model may be applied to evaluate potential therapeutic options using an FDA-approved small molecule, Omaveloxolone. Finally, we analyzed host cell responses to infection and treatment using gene expression and phospho-signaling analyses. These findings highlight the value of this model to interpret the pathogenic mechanisms of neurotropic viral infections and evaluate potential therapeutic intervention strategies in a clinically relevant in vitro human neuronal model.

Microorganisms doi: 10.3390/microorganisms14091943

Authors: Ying Li Lei Shen Miaomiao An Xuming Wang Guozhu Zhao Tianlei Qiu

A novel bacterial strain, CR191T, was isolated from maize (Zea mays) rhizosphere soil. The strain is Gram-stain-negative, oxidase-positive, rod-shaped, non-motile, and aerobic. Phylogenetic analysis based on 16S rRNA gene sequences placed CR191T within the genus Dyella, with highest similarity to Dyella flava DHOC52T (98.23%) and Dyella dinghuensis DHOA06T (98.16%). The genomic DNA G+C content was 66.19%. Average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between strain CR191T and the type strains of the other 36 validly published Dyella species ranged from 79.0 to 86.6% and 20.2–28.7%, respectively, both below the recognized thresholds for species delineation. Genome mining identified several putative biosynthetic gene clusters showing similarity to known clusters associated with diverse secondary metabolites, including saframycin, cosmomycin, and malleobactin biosynthesis. The major cellular fatty acids were iso-C17:0, iso-C15:0, and summed feature 9 (comprising iso-C17:1 ω9c and/or C16:0 10-methyl). The polar lipid profile comprised phosphatidylglycerol, phosphatidylethanolamine, phosphatidylmonomethylethanolamine and two unidentified polar lipids; the sole respiratory quinone was ubiquinone Q-8. Based on a combination of phenotypic, chemotaxonomic, genomic, and phylogenetic characteristics, strain CR191T is considered to represent a novel species of the genus Dyella, for which the name Dyella cornirhiza sp. nov. is proposed. The type strain is CR191T (=CCAM 2032T = JCM 36807T).

Microorganisms doi: 10.3390/microorganisms14091942

Authors: Liwen Yin Yiming Li Xuetao Gong Peishan Chen Weihui Wu Un-Hwan Ha Shouguang Jin Yongxin Jin

Pseudomonas aeruginosa is a nosocomial pathogenic bacterium that causes a wide range of human infectious diseases. The type III secretion system (T3SS) serves as a key virulence determinant underlying the pathogenesis of this bacterium in acute infections. As the master transcriptional activator of T3SS, ExsA binds to target promoter regions and modulates the expression of all currently identified T3SS genes. In this study, we identified tyrosine phosphoprotein A (TypA) as a repressor that restricts expression of the T3SS in P. aeruginosa. TypA interacts with ExsA to block its binding to target promoters, thereby inhibiting T3SS expression. We show that the typA expression is induced in response to low calcium, low temperature, a biofilm lifestyle, and direct contact with host cells. Additionally, the absence of TypA caused a growth defect in P. aeruginosa at low temperatures. Collectively, these data confirm the significant role of TypA and reveal a novel molecular mechanism by which P. aeruginosa regulates T3SS.

Microorganisms doi: 10.3390/microorganisms14091941

Authors: Cristiana Georgeta Bujor Marilena Dinuti Felicia Sfrijan

Blood cultures are frequently obtained in hospitalized patients with COVID-19, although the diagnostic yield and clinical meaning of culture positivity vary markedly according to case mix, disease severity, and timing of sampling. Omicron-era data remain comparatively limited. We aimed to characterize blood-culture positivity, pathogen distribution, inflammatory correlates, antimicrobial exposure, and short-term outcomes in a homogeneous cohort hospitalized during the SARS-CoV-2 Omicron BA.5 wave. This retrospective observational study included 395 patients hospitalized with SARS-CoV-2 infection between 12 July and 25 September 2022. Patients were categorized as blood-culture positive when a named bacterial species was recorded and as culture negative when “No growth” was documented. Continuous variables were compared using the Mann–Whitney U test and categorical variables using Fisher’s exact or chi-square tests, as appropriate. A parsimonious multivariable logistic regression model was used to explore factors independently associated with blood-culture positivity. Blood cultures were positive in 89/395 patients (22.5%). Gram-positive organisms accounted for 52.8% of positive cultures and Gram-negative organisms for 47.2%. The most frequent isolates were Streptococcus pneumoniae (20.2% of positive cultures), Enterococcus faecalis (18.0%), Klebsiella pneumoniae (18.0%), Staphylococcus aureus (14.6%), Escherichia coli (14.6%), and Pseudomonas aeruginosa (14.6%). Compared with patients with no growth, those with positive cultures had higher admission leukocyte counts (median 13.42 vs. 11.22 × 109/L; p < 0.001), C-reactive protein (116.15 vs. 105.50; p = 0.007), and interleukin-6 (17.24 vs. 9.39; p = 0.027). A documented sepsis diagnosis was more frequent in the culture-positive group (48.3% vs. 28.1%; OR 2.39, 95% CI 1.47–3.88; p < 0.001). Mortality was numerically higher with culture positivity (20.2% vs. 14.4%) but did not reach statistical significance (p = 0.188). In the adjusted model, higher leukocyte count remained independently associated with culture positivity (aOR 1.10 per 1 × 109/L, 95% CI 1.05–1.15; p < 0.001), while vaccination status showed an inverse exploratory association (aOR 0.58, 95% CI 0.35–0.96; p = 0.033). The exploratory model AUC was 0.672 (bootstrap 95% CI 0.608–0.731); calibration was not assessed. Blood-culture positivity was common in this hospitalized BA.5 cohort and showed a nearly balanced Gram-positive/Gram-negative distribution. Leukocytosis and clinically documented sepsis were the clearest correlates of positivity, whereas mortality did not differ significantly. These findings support selective blood-culture use driven by bacterial infection signals and emphasize the need to connect antimicrobial decisions to microbiological evidence.

Microorganisms doi: 10.3390/microorganisms14091940

Authors: Laura Sutarlie Sian Yang Ow Karrie Kwan Ki Ko Chayaporn Suphavilai Kar Mun Lim Patipan Boonsimma Darren Wei Tan Niranjan Nagarajan Xiao Di Su

Surveillance of methicillin-resistant Staphylococcus aureus (MRSA) in the environment is crucial for reducing its transmission, particularly in hospitals, nursing homes, and high-contact public areas. Rapid Point-of-Need (PON) tools at healthcare facilities, without involving prolonged bacterial culture in sophisticated laboratories, are needed for environmental surveillance of MRSA. We have developed a gold nanoparticle (AuNP)-based “Swab-and-Stain” assay for rapid and on-site detection of PBP2a-expressing MRSA from environmental surfaces. The assay utilizes anti-PBP2a antibody immobilized on cotton swabs to specifically capture PBP2a-expressing MRSA and anti-MRSA antibody conjugated on AuNPs to detect the captured MRSA on the swabs via sandwiched complex formation. In the presence of PBP2a-expressing MRSA, the swabs were stained in red, as seen with the naked eye. The color intensity of the stain can be quantified by smartphone image analysis for MRSA quantification. Using PBP2a-expressing MRSA samples spiked in real hospital sink and drain matrices, this pilot study demonstrated a “Swab-and-Stain” assay as a proof-of-concept method for detecting MRSA at a concentration as low as 12 CFU/mL. The assay was validated to specifically detect MRSA spiked in real hospital sink and drain matrices. This “Swab-and-Stain” assay provides a rapid, portable, and easy-to-use platform with potential application for PON environmental surveillance of MRSA.

Microorganisms doi: 10.3390/microorganisms14091939

Authors: Nize Otaru Erik Eckhardt Matthew R. Hayward Tomoya Isaji Yuki Kamioka Britt Blokker Gabriel Vinderola

Unusually frequent and soft stools may negatively impact the quality of life. A postbiotic containing heat inactivated Limosilactobacillus fermentum CNCM I-2998 and Lactobacillus delbrueckii subsp. lactis CNCM I-4831 has previously been shown to alleviate gastrointestinal symptoms and diarrhea in patients with clinically diagnosed gastrointestinal disorders. This study evaluated whether this postbiotic can support bowel movement frequency in Japanese healthy adults with self-reported frequent and soft stools. In a randomized, double-blind, placebo-controlled clinical trial, participants (n = 100) received the postbiotic (20 billion heat-inactivated cells per day) or placebo for four weeks. The primary endpoint was weekly bowel movement frequency. Secondary endpoints included days with bowel movements per week, sense of relief, stool characteristics (shape, color, odor, water content), gastrointestinal symptoms, and fecal microbiome composition. After four weeks, the postbiotic group showed significantly lower bowel movement frequency and fewer days with bowel movements when compared with the placebo group. Importantly, these reductions were not accompanied by decreased stool volume or increased hard stools or constipation symptoms. Mild changes in microbiome composition were detected across groups, and no adverse events were reported. These findings suggest that the postbiotic is safe and can help reduce bowel movement frequency in healthy individuals experiencing unusually frequent and soft stools.

Microorganisms doi: 10.3390/microorganisms14091938

Authors: Siyang Dai Weihao Lan Weijia Geng Pan Liu Bujian Wang Xun Li Yongqiang Fan Fuhui Wang Dake Xu Ying Zheng

Microbiologically influenced corrosion (MIC) at oral biomaterial interfaces is viewed as an ecological phenomenon, yet how microbial composition and interspecies interactions shape corrosion remains unclear. Here, we investigated whether oral microbial community composition and interspecies interactions contribute to medical 316L stainless steel corrosion. Consortia-enhanced Cr and Fe release and localized surface pitting, with marked inter-subject variability. Interface-associated biofilms exhibited trends toward compositional shifts and showed enrichment of predicted pathways for fermentation and riboflavin metabolism, along with higher genome-based metabolic interaction potential than planktonic communities. Extracellular riboflavin accumulated in MIC systems and correlated positively with dissolved Cr and Fe concentrations. In perturbation assays, riboflavin supplementation increased corrosion current density (icorr) and metal dissolution, whereas roseoflavin reduced extracellular riboflavin availability and corrosion-related parameters without marked changes in the measured biofilm biomass or surface-associated ATP levels. A defined three-strain consortium (C. tsuruhatensis, R. erythropolis, and T. aromatica) reconstituted the S3 high-corrosion phenotype, including elevated icorr, extracellular riboflavin accumulation, and induced pitting, consistent with a proposed riboflavin-linked model involving species-dependent metabolic interactions. These findings suggest that extracellular riboflavin may represent a candidate redox-active factor associated with microbial community interactions and corrosion activity, providing an ecological framework for understanding microbiota-associated corrosion resistance at oral biomaterial interfaces.

Microorganisms doi: 10.3390/microorganisms14091936

Authors: Miguel Anchundia Stalin Santacruz

Fresh cheeses are highly perishable dairy products due to their high moisture content, pH, and other intrinsic characteristics that support microbial growth. This scoping review synthesized the available evidence on spoilage-associated bacteria in fresh cow cheeses, the physicochemical and sensory changes associated with spoilage, and the methods for their identification and quantification. The review was conducted in accordance with the PRISMA-ScR guidelines and registered in the Open Science Framework (OSF). Literature searches were performed in PubMed/MEDLINE, Scopus, SciELO, and Google Scholar through 2 April 2026. Of 7379 records identified, 30 studies met the inclusion criteria. Across the reviewed studies, Pseudomonas spp. emerged as the predominant spoilage-associated bacteria, followed by members of the Enterobacteriaceae, lactic acid bacteria (LAB), and spore-forming bacteria. Total viable count was the microbiological indicator most frequently used to monitor spoilage progression. Microbial spoilage was consistently associated with acidification, proteolysis, lipolysis, discoloration, gas production, texture deterioration, and reduced sensory acceptability. These findings highlight the central role of psychrotrophic bacteria in the spoilage of fresh cow cheeses and emphasize the importance of microbiological monitoring and spoilage-control strategies throughout production and storage.

Microorganisms doi: 10.3390/microorganisms14091937

Authors: Jing Yang Zhuo Wang Chuanxu Wang Yunjie Li Yajie Niu Jia Feng Shulian Xie Xin Li

Salinity is a key driver of microbial community structure and function in salt lake ecosystems, yet how it shapes functional genes involved in carbon (C), nitrogen (N), and phosphorus (P) cycling remains poorly understood. We collected metagenomic samples along a natural salinity gradient in Yuncheng Salt Lake and examined how salinity was associated with microbial taxonomic and functional diversity and with C, N, and P cycling genes. Both diversity metrics decreased significantly with increasing salinity and were positively correlated with each other. The composition and abundance of C, N, and P cycling genes differed significantly among the low-, medium-, and high-salinity groups. In carbon cycling, most carbon fixation genes were more abundant at higher salinity, whereas most carbon degradation genes were less abundant; within carbon fixation, reductive tricarboxylic acid (rTCA) cycle and Calvin cycle gene abundances were higher. In nitrogen cycling, nitrogen mineralization and assimilation genes were significantly more abundant. In phosphorus cycling, transporter and pyrimidine metabolism genes were more abundant, whereas the relative contribution of purine metabolism genes declined. Co-occurrence network analysis revealed dense positive co-occurrence associations among C, N, and P cycling genes, with mer, GLU, and ppk1 as highly connected genes. Mantel tests identified salinity and pH as the primary environmental factors associated with functional gene variation. These results suggest that salinity may regulate C, N, and P cycling genes partly by reshaping microbial community structure in salt lake ecosystems.

Microorganisms doi: 10.3390/microorganisms14091935

Authors: Chao Cui Yujie Zheng Zhiguo Guo Zhichang Wang Xiaoyan Zhu Yalei Cui Defeng Li Chen Wen Yinghua Shi Boshuai Liu

The specific differences, unique characteristics, and functional linkages of rumen microbiota and their metabolic pathways across different ruminant species remain poorly understood. To investigate these cross-species relationships, 16 Holstein cows, 28 Simmental crossbred cattle, and 15 Boer goats were fed under standardized conditions for 97 days. Rumen microbial composition and function were evaluated using 16S rRNA gene sequencing and PICRUSt. Results revealed distinct, host-specific microbial architectures. Holstein cows exhibited an overall enrichment of the phylum Proteobacteria and ABC transporter pathways. Notably, their micro-ecosystem was dominated by the Succinivibrionaceae_UCG-001 flora type, which is computationally associated with propionate synthesis for milk production and is predicted to potentially contribute to altered methane metabolism. In contrast, Simmental cattle were characterized by Succinivibrionaceae_UCG-002 and computationally upregulated glycolysis/gluconeogenesis pathways, which are associated with volatile fatty acid conversion for energy deposition. Furthermore, Boer goats harbored a unique fiber-adapted ecosystem exclusively enriched with the norank_f_Bacteroidales_BS11_gut_group and Lachnospiraceae_ND3007_group, correlating with butyrate production and maintaining a classic acetate-type fermentation profile, while exhibiting an elevated predicted genomic potential for methane production based on functional profiling. This study demonstrates that rumen microbiota, metabolic pathways, and fermentation parameters form a highly coordinated network shaped by host phylogeny, energy allocation, and specific diets, providing a theoretical basis for targeted microbiome modifications to improve animal health and feed utilization.

Microorganisms doi: 10.3390/microorganisms14091934

Authors: Muqadas Altaf Waseem Khalid Zhijia Fang Haroon Munir Afifa Aziz Muhammad Bilal Hussain Qingping Wu Ravi Gooneratne

The widespread exposure of multidrug-resistant bacterial strains due to excessive use of antibiotics has accelerated the bacteriophage applications as highly specific antibacterial agents competent to target resistant pathogenic Vibrio through receptor-mediated adsorption and lytic infection. Vibrio pathogens are responsible for substantial economic losses, environmental instability, and foodborne infections in humans through contaminated seafood and water. However, the molecular determinants remain incompletely understood, commanding the Vibrio-phage host specificity, host range, and resistance evolution. This review highlights the comprehensive overview of structural, and functional framework of Vibrio surface receptors responsible for phage recognition, including outer membrane vesicles (OMVs), porins, lipopolysaccharides (LPS), capsular polysaccharides (CPS), flagella, and pili along with specialized bacteriophage tail spike proteins known as receptor-binding proteins (RBPs). We also explored the mechanisms against phage-mediated lysis, including receptor modification, intracellular defense systems, physiological remodeling, and structural adaptations that regulate burst size, lysis timing, and resistance evolution with emphasis on significant progress in phage biology and therapeutic development. It is critical to grasp the molecular-level interaction mechanisms due to limited marine phage ecological data, lack of standardized resistance databases, and regulatory constraints. Future perspectives emphasize the engineered phages, multi-omics integration, and artificial intelligence-driven phage design for the sustainable management of Vibriosis and improved aquaculture biosecurity.

Microorganisms doi: 10.3390/microorganisms14091933

Authors: Folliero Veronica Dell’Annunziata Federica Radocchia Giulia Leone Raimondo Gualano Maria Rosaria Fusco Alessandra Putignani Lorenza

Tuberculosis (TB) remains one of the most significant global public health challenges, as it represents the world’s leading infectious cause of death. The clinical efficacy of currently available anti-TB drugs is increasingly compromised due to the growing prevalence of antibiotic-resistant strains, side effects, and prolonged treatment times. This scenario highlights the urgent need to identify new anti-TB drugs with alternative mechanisms of action and improved anti-TB activity and bioavailability. In this context, natural products derived from microorganisms represent a key source of chemical diversity for drug discovery. Actinomycetes, fungi, and other environmental microbes produce a wide range of secondary metabolites with broad antimicrobial activity. These compounds can interfere with essential bacterial structures and processes, including membrane integrity, redox homeostasis, protein synthesis, and DNA replication. This review aims to evaluate new microbial natural products for their antibacterial activity, focusing on their efficacy, mechanisms of action, chemical nature, and potential clinical applications, with the aim of informing the development of new anti-TB agents.

Microorganisms doi: 10.3390/microorganisms14091932

Authors: Seyedmohammad Hosseinpourlamardi Shaiqa Labiba Li Li Ali Dadvar Manita Guragain Ute Römling

The phosphoethanolamine-modified exopolysaccharide cellulose synthesized by the type IIa cellulose biosynthesis nanomachine is a major extracellular matrix component of Escherichia coli. Here, we aim to investigate whether homologs of bcsABC genes of the core genome EcType 1 bcs cellulose biosynthesis gene cluster, which are diminished or abolished in high-virulence isolates, and entire clusters are mobilized on plasmids and occasionally manifested on E. coli chromosomes at alternative locations. While EcType BcsA2 and BcsA3 cellulose synthases are restricted to the genus Escherichia, the EcType BcsA4 cellulose synthase and associated bcs gene products are highly similar to Klebsiella pneumoniae counterparts. Cyclic di-GMP turnover proteins not previously recognized to post-translationally regulate cellulose biosynthesis on the chromosomal level are frequently co-localized with novel bcs genes. Thermotolerant meat-derived E. coli 730V1 uniquely harbors an EcType 4 bcs gene cluster of type Ib with a type IIb bcsG gene on a plasmid. Chemical, phenotypic and genetic evidence showed cellulose biosynthesis, cellulose-dependent cell aggregation and activation of biosynthesis by the second messenger cyclic di-GMP, an allosteric activator of the cellulose synthase. With gene duplication, horizontal gene transfer and recombination to contribute to the multiplication and diversification of bcs gene clusters in a number of different bacterial species, the extent and ecological role(s) of the multiplication, transfer and replacement of cellulose biosynthesis gene clusters, as well as in species other than E. coli, still need to be unraveled. Our data indicate, however, that although diversification and multiplication of bcs clusters is ongoing in the species E. coli, these events become rarely manifested in the population.

Microorganisms doi: 10.3390/microorganisms14091930

Authors: Antonio Braga Gustavo Ribeiro Lima Fernanda da Costa Negraes Maria Vitória Moura Fajardo Karine Mello Duvivier Luis Fernando Lima Bueno Susana Cristina Aidé Viviani Fialho Edward Araujo Júnior Jorge Rezende-Filho

The vaginal ecosystem undergoes profound physiological adaptations during pregnancy and the puerperium through dynamic interactions among the vaginal microbiota, epithelial barrier, local immune system, and hormonal milieu. Disruption of this homeostasis may lead to vaginal dysbiosis and infection, with potential consequences for maternal, fetal, and neonatal health. This narrative review aimed to provide a comprehensive and updated overview of the vaginal ecosystem throughout pregnancy and the puerperium, integrating current evidence on physiological changes in the vaginal microbiota, mechanisms of dysbiosis, major vaginal infections, diagnostic approaches, therapeutic management, and maternal–fetal implications. A structured literature search was conducted in PubMed/MEDLINE and Scopus through July 2026. Priority was given to recent systematic reviews, meta-analyses, international clinical guidelines, randomized clinical trials, and observational studies addressing the vaginal microbiota, dysbiosis, bacterial vaginosis, vulvovaginal candidiasis, trichomoniasis, aerobic vaginitis, pregnancy, and the puerperium. Pregnancy is generally characterized by a stable, low-diversity, Lactobacillus-dominated vaginal microbiota, whereas the postpartum period is associated with reduced Lactobacillus abundance, increased microbial diversity, and gradual restoration of eubiosis. Disruption of this ecosystem promotes biofilm formation, microbial persistence, inflammation, and ascending infection. Bacterial vaginosis, vulvovaginal candidiasis, trichomoniasis, and aerobic vaginitis represent the major vaginal infections during pregnancy and the puerperium and have varying associations with adverse outcomes, including preterm birth, preterm premature rupture of membranes, chorioamnionitis, postpartum infection, and neonatal morbidity. Molecular diagnostics and microbiome profiling have improved etiological characterization, while emerging microbiome-directed interventions offer potential strategies for restoring vaginal homeostasis. Current evidence supports an increasingly ecosystem-centered approach to vaginal health during pregnancy and the puerperium. Accurate etiological diagnosis and evidence-based treatment remain essential, while preservation and restoration of vaginal homeostasis may represent important complementary objectives for improving maternal, fetal, and neonatal outcomes.

Microorganisms doi: 10.3390/microorganisms14091931

Authors: Sizhu Pan Wei Zheng Zhimei Feng Zhanhong Ouyang Mingchen Ma Luying Huang Lulu Wang Xitao Wang Chunshan Quan

Intensive aquaculture and antibiotic overuse have rendered largemouth bass (Micropterus salmoides) prone to frequent mass mortalities and economic losses. Probiotics are promising antibiotic alternatives for improving intestinal health and immunity. This study evaluated a novel butyrate-producing strain Clostridium butyricum DMZ-SG06 to elucidate its growth-promoting and immunomodulatory mechanisms. Three dietary treatments were used: control (Con, basal diet), T1 (basal diet supplemented with 1 g C. butyricum powder containing 5 × 108 CFU per 10 g diet), and T2 (basal diet supplemented with 1 mL C. butyricum suspension containing 5 × 108 CFU per 10 g diet). A total of 270 healthy juvenile largemouth bass (5.63 ± 0.03 g) were randomly allocated to three groups (90 fish per group), with each group divided into three replicate tanks (60 × 40 × 34 cm) (30 fish per tank). Fish were fed the corresponding diets to apparent satiation for a 60-day rearing period. Results showed that T2 increased weight gain rate (WGR) by 11.35% compared with control (p < 0.05, 213.90 ± 12.30% vs. 192.10 ± 10.20%) and significantly improved the specific growth rate (SGR, p < 0.05), with T2 performing best. Probiotic treatments markedly improved intestinal morphology, nonspecific immune indices, and antioxidant status (p < 0.05), with T2 exerting the most prominent effects. C. butyricum DMZ-SG06 significantly reduced the levels of pro-inflammatory cytokines. For instance, TNF-α levels in T2 decreased 1.39-fold compared with the control group (128.57 ± 4.97 pg/g vs. 178.67 ± 7.80 pg/g, p < 0.05). Meanwhile, the levels of anti-inflammatory cytokines were upregulated. For example, IL-10 levels in T2 increased 1.32-fold relative to the control group (321.89 ± 15.03 pg/g vs. 244.31 ± 7.57 pg/g, p < 0.05). Metagenomic analysis revealed reduced Acinetobacter abundance, enriched beneficial genera (Lactobacillus, Parabacteroides, p < 0.05), and enhanced microbial functions related to the phosphotransferase system and galactose metabolism (p < 0.05). In conclusion, C. butyricum DMZ-SG06 promotes largemouth bass growth and intestinal health via butyrate metabolism, immune modulation, and microbiota remodeling. Notably, the liquid bacterial suspension formulation exerts a more significant effect on enhancing the fish’s growth performance and intestinal health than the powder counterpart, supporting its application as a safe probiotic in antibiotic-free aquaculture.

Microorganisms doi: 10.3390/microorganisms14091929

Authors: Guiying Cui Peng Zhang Qian Chen Jiuming Zhang Qingyi Wang Shanshan Zhang Yiming Shi Zhidan Zhang Yang Wang

The process of straw decomposition is highly complex and is regulated by a multitude of interacting factors. However, how the straw return rate influences the metabolic byproducts, extracellular enzymes, and microbial communities during straw decomposition remains an unresolved question. To address this, we integrated straw and soil chemistry, amplicon sequencing, untargeted metabolomics, and enzyme assays across four straw return rates (no return, 1/3, 1/2, and the Full treatment) at 30 and 90 d. Straw mass loss was greatest under the 1/2 return treatment at 90 d, reaching 53.7%, whereas soil organic carbon (SOC) and total nitrogen (N) were highest under the 1/3 return, exceeding the control by 17.6% and 17.2%, respectively. This indicates that decomposition and short-term soil C and N accumulation were decoupled. Microbial communities underwent clear temporal turnover, and hydrolytic enzymes increased from 30 to 90 d and were positively associated with mass loss. In contrast, oxidative enzymes showed no positive association with decomposition. Metabolomic profiles shifted from early-stage labile compounds to later-stage aromatic and phenolic compounds, and straw and soil metabolomes were closely coupled in a stage-specific manner (Procrustes M2 = 0.15, p = 0.001; Mantel r = 0.69, p = 0.001). Mantel tests further indicated that the metabolome was significantly associated with return rate and SOC. Partial least squares path modeling revealed that the direct and indirect pathways linking return rate, metabolome, enzymes, and decomposition were reorganized between 30 and 90 d. Overall, straw return rate influenced decomposition through stage-dependent biochemical and microbial changes, and straw mass loss and short-term soil C and N accumulation were decoupled, responding nonlinearly and peaking under different return rates.

Microorganisms doi: 10.3390/microorganisms14091928

Authors: Carolina Magri Ferraz Valéria Modolo Peterle Matheus Zorzal Bernardes Rangel Gabrielly de Moura Paris Enzo Bernardes Rocha Fávaro Sarah Bernardes Simões Lucas Possa Oliveira Heloisa Cristina Brugnera Júlia Silva Santana Gustavo Guimarães Fernandes Viana Alessandra Figueiredo de Castro Nassar Vanessa Castro Ricardo Pinto Schuenck Juliano Gonçalves Pereira João Pedro Rueda Furlan Marita Vedovelli Cardozo Gabriel Augusto Marques Rossi

Foods can be contaminated by a wide range of microorganisms, originating from multiple sources within domestic kitchens. However, the role of flies and ants as carriers of bacteria in these environments remains poorly understood, and data on the antimicrobial susceptibility profiles of insect-associated bacteria remain scarce. Therefore, this study aimed to identify Enterobacterales and Staphylococcus spp. recovered from flies and ants collected in domestic kitchens and to characterize the antimicrobial susceptibility profiles of the isolates. Additionally, selected virulence genes were investigated in Staphylococcus aureus and Escherichia coli, two recognized foodborne pathogens. A total of 240 insects (113 flies and 127 ants) were collected from 58 domestic kitchens. Twenty-six Enterobacterales isolates belonging to nine species were recovered from 17 kitchens, including a single multidrug-resistant (MDR) isolate identified as Enterobacter hormaechei. In addition, 45 staphylococcal isolates representing nine species were recovered from 25 kitchens, originating from 36 flies and 9 ants. Among these, 13 isolates exhibited an MDR phenotype, including S. aureus and several non-aureus Staphylococcus species. None of the E. coli isolates carried the eae or stx1/stx2 genes, whereas none of the S. aureus isolates harbored the sea, seb, sec, see, or mecA genes. To the best of our knowledge, this is the first study to report the simultaneous recovery of MDR bacteria from flies and ants collected in domestic kitchens. These findings provide new evidence that household insects may act as carriers of antimicrobial-resistant bacteria, highlighting their potential role in the dissemination of clinically relevant bacterial species, including MDR strains, within domestic environments. Effective insect control and good household food hygiene practices may help reduce the spread of antimicrobial-resistant bacteria and mitigate the risk of difficult-to-treat foodborne infections.

Microorganisms doi: 10.3390/microorganisms14091925

Authors: Yongsheng Ge Zhi Li Caiyun Yu Weitong Guo Guangying Fan Guiyu Lin Han Yu Ying Wang

Autism spectrum disorder (ASD) is a heterogeneous neurodevelopmental condition characterized by impaired social interaction and repetitive stereotyped behaviors, with pathogenic mechanisms that remain incompletely understood. The gut microbiota has emerged as a key regulator of ASD; however, its impact on hippocampal proteomic and phosphoproteomic signatures has not been fully characterized. In this study, we performed fecal microbiota transplantation (FMT) by transferring fecal samples from children with ASD and typically developing controls into antibiotic-treated mice. Gut microbiota from children with ASD induced several ASD-like behaviors in recipient mice, accompanied by aberrant activation of microglia, astrocytes, and neurons, as well as impaired neurogenesis. Phosphoproteomic profiling combined with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that differentially phosphorylated proteins were predominantly enriched in synapse-related pathways. ASD-derived microbiota markedly reduced synaptic density, downregulated the synaptic proteins SYP and PSD-95, and inhibited the expression of blood–brain barrier (BBB) tight junction proteins. In silico structural simulations using AlphaFold3 (AF3) and HADDOCK further supported that ASD-FMT may promote abnormal phosphorylation, potentially remodeling SHANK3 and SRRM2 conformations and weakening the binding affinity of SHANK3. Integrative proteomic and phosphoproteomic screening identified FNDC3A as a potential susceptibility-associated protein upregulated by gut microbiota from children with ASD, which was verified in mouse hippocampal tissues and plasma samples from children with ASD using Western blotting and ELISA, respectively. Mechanistically, ASD pathogenesis may be attributable not only to the dysregulation of classical ASD susceptibility genes but also to gut microbiota-driven post-translational phosphorylation remodeling of multiple protein structures. Importantly, this study established an innovative research framework that integrates in silico analyses with wet-lab experiments, yielding novel insights into ASD pathogenesis from the perspective of gut microbiota-induced alterations in the hippocampal phosphoproteome and revealing a plausible molecular mechanism underlying ASD.

Microorganisms doi: 10.3390/microorganisms14091927

Authors: Mesut Yilmaz Naim Mahroum

Meningitis and encephalitis necessitate rapid pathogen identification to guide therapy, as conventional methods are time-consuming. This study evaluated both the wet-lab analytical performance and the clinical performance evaluation of the Bioeksen Meningitis/Encephalitis Panel (BS-MEP) integrated onto the fully automated, high-throughput Sigmoida Lab platform. Analytical limits of detection (LoD) were defined via Probit analysis (95% threshold) by spiking negative cerebrospinal fluid (CSF) matrices. Target detection was verified using characterized reference materials for all 14 analytes, and in silico primer/probe coverage was assessed against taxon-specific sequence databases. Cross-reactivity was evaluated using high-prevalence non-target organisms. Clinical performance was evaluated retrospectively using 500 archived, anonymized CSF specimens from a single-center repository, with analyte-specific classifications compared with prespecified routine comparator methods. The automated platform provided an 80-min sample-to-result turnaround for up to 23 samples. Verified LoDs ranged from 472 to 2118 genome copies/mL. All inclusivity strains were successfully detected in 5/5 replicates. In silico coverage exceeded 98% combined, and zero wet-lab cross-reactivity was observed. Precision coefficients of variation (CVs) were consistently ≤1.38%. In the clinical evaluation, the pooled clinical agreement was high, with PPA ranging from 90.9% to 100% and NPA of 100% across the evaluated analytes. Analyte-level clinical sensitivity ranged from 90.9% to 100%, with zero false-positive results across all targets. The fully automated molecular system demonstrates excellent analytical robustness and strong clinical agreement for syndromic detection of major CNS pathogens. The automated workflow combines broad pathogen coverage with an approximately 80-min sample-to-result turnaround and warrants further prospective evaluation in routine clinical settings.

Microorganisms doi: 10.3390/microorganisms14091926

Authors: Klaudia Grudzińska Kacper Bułdyś Cezary Dubaj Ernest Kuchar Anna Piwowarczyk

Early identification of children at risk of severe acute respiratory infection remains a clinical challenge. Although C-reactive protein (CRP), procalcitonin (PCT), and white blood cell (WBC) count are commonly assessed at hospital admission, their predictive value for severity assessment remains uncertain. We aimed to evaluate the utility of clinical features, a respiratory multiplex PCR test detecting 23 pathogens, imaging findings, and early inflammatory biomarkers in predicting subsequent supplemental oxygen requirement, used as a marker of severe acute lower respiratory infection (ALRI), among hospitalized children. The primary outcome was the need for supplemental oxygen during hospitalization. We conducted a single-center retrospective cohort study of children admitted to the pediatric department between May 2022 and June 2023 with ALRI or other conditions for which respiratory infection was considered but subsequently excluded. All included patients underwent multiplex respiratory PCR testing. We included 439 children in the analysis [median age, 20 mo (IQR: 5–54); 52.7% males]. Out of them, 178 (40.5%) required supplemental oxygen. The strongest positive association with subsequent supplemental oxygen requirement was observed for low oxygen saturation on admission [OR 8.83 (95% CI, 3.00–26.02; p < 0.001)], although this variable was documented in only about half of the cohort (52.4%). The strongest predictors of subsequent supplemental oxygen requirement in multivariable analysis were age, abnormal auscultatory findings, and the cumulative number of danger signs, including hypoxemia, tachypnea, dyspnea, and increased work of breathing. Among the 23 pathogens tested, respiratory syncytial virus (RSV) was the only pathogen consistently associated with a more severe course: 60.3% of RSV-positive children required supplemental oxygen, compared with 36.9% of RSV-negative children (OR 2.59, 95% CI 1.53–4.40; p < 0.001). For predicting subsequent supplemental oxygen requirement, no single marker had strong utility. These findings support the importance of structured bedside clinical assessment for identifying hospitalized children who may subsequently require supplemental oxygen. Given incomplete documentation of some vital signs, associations involving these variables should be interpreted with caution.

Microorganisms doi: 10.3390/microorganisms14091924

Authors: Yunhan Qin Yidan Tian Tingfu Li Changliang Nie Xueyun Geng Xiaomin Sun Lingyu Li Yoshizumi Kajii

Airborne bacteria exhibit high taxonomic diversity and a well-documented capacity to influence human health through pathogenic infection, allergenic sensitization, and immunomodulatory or commensal exposure. However, characterization of the airborne bacterial community structure, and particularly the seasonal dynamics, size-resolved distribution, and functional potential of predicted potential pathogens in Qingdao, a coastal megacity on the Yellow Sea, remains limited. Results revealed pronounced seasonal variation in community structure: Staphylococcus, Lactobacillus, Escherichia–Shigella, and Klebsiella were consistently dominant across all samples. LEfSe analysis identified Staphylococcus as significantly enriched in autumn, supporting its utility as a robust seasonal biomarker (LDA > 3.5, p < 0.05). This genus is predominantly associated with human sources and comprises opportunistic pathogens, thereby posing a potential risk to human health. Significant compositional divergence between summer and autumn communities was observed as a higher beta deviation index, whereas there were no statistically significant differences between particle size groups. The Chao and Shannon indices were consistently higher in PM10 than in PM2.5 across all seasons; however, this size-associated difference reached statistical significance only for autumn PM10 (p < 0.05), indicating a seasonally amplified effect of coarse particulate matter on microbial richness and evenness. These analyses provide new insight into the total airborne bacterial and potential pathogens in an urban coastal environment.

Microorganisms doi: 10.3390/microorganisms14091923

Authors: Shuaibo Han Xiaolong He Jianglong Guo Jinping Deng Xueye Li Jiayin Wang Xinxing Wu Hui Wang Yan Zhang Fangli Sun

A lignin-degrading bacterium, strain BE17T, was isolated from decaying bamboo and identified as a novel species within the genus Brucella, proposed as Brucella ligniniphila, sp. nov. (type strain = MCCC 1K08845T = JCM 36600T). BE17T exhibited a lignin degradation rate of 7.9% with lignin as the sole carbon source, which increased to 17.2% upon glucose supplementation. Structural analysis indicated that BE17T degraded lignin by cleaving β-O-4 bonds, oxidizing aliphatic side chains and disrupting aromatic rings. Low-molecular-weight lignin fractions were more readily degraded. Genome analysis revealed genes encoding DyPs, peroxidases, monooxygenases, and dioxygenases. GC-MS analysis identified aromatic monomers and organic acids among the degradation products, and together with genome annotation, these data suggested the involvement of CoA-dependent non-β-oxidative, β-ketoadipate, and phenylacetic acid pathways. To enhance lignin degradation efficiency, a co-culture system (BT) of BE17T and Trametes versicolor was established. This system achieved a degradation rate of 36.7% under optimized conditions (25 °C, pH 6.0, BE17T inoculated on day 9). Microstructural and chemical analyses confirmed that fungal pretreatment facilitated lignin degradation. This study reveals the lignin-degrading potential of a novel Brucella species and demonstrates the synergistic effect of a bacterial–fungal system, offering insights into lignin biodegradation and bioconversion strategies.

Microorganisms doi: 10.3390/microorganisms14091921

Authors: Jielan Mi Xiao Zhang Jing Liu Kun Yue Zhihan Yang Yujia He Yuanming Huang Liqiong Song Zhihong Ren Jianguo Xu

Klebsiella pneumoniae (KP) is an important opportunistic pathogen that can persist in the gastrointestinal tract and serve as a reservoir for subsequent infection. The increasing prevalence of multidrug-resistant KP highlights the need for non-antibiotic strategies to limit intestinal KP burden and associated host injury. Lactiplantibacillus plantarum GUANKE (L. plantarum GUANKE) is a candidate probiotic strain with reported effects on mucosal barrier protection and inflammatory regulation, but its role in intestinal KP challenge remains unclear. In this study, the effects of L. plantarum GUANKE and its metabolite phenyllactic acid (PLA) on KP growth, intestinal KP burden, and associated inflammatory injury were investigated using antibacterial assays in vitro, metabolomic analyses, an antibiotic-pretreated mouse model of intestinal KP challenge, and an influenza A virus (IAV)/KP intestinal co-exposure model. L. plantarum GUANKE and its culture-derived products inhibited KP growth in vitro. Metabolomic analysis identified PLA as a metabolite enriched in L. plantarum GUANKE fermentation supernatants, and exogenous PLA directly inhibited KP growth in a dose-dependent manner. In an antibiotic-pretreated mouse model of intestinal KP challenge, oral administration of L. plantarum GUANKE reduced intestinal KP burden and was associated with changes in gut microbiota composition, increased cecal PLA abundance, improved intestinal barrier-related parameters, and reduced inflammatory responses, while exogenous PLA partially reproduced these effects. IAV infection increased susceptibility to intestinal KP expansion, and oral L. plantarum GUANKE reduced KP burden in the IAV/KP intestinal co-exposure model. In mice challenged with multidrug-resistant KP strain NK04152, L. plantarum GUANKE and PLA reduce intestinal KP burden and associated tissue injury in mice, supporting their potential as candidate microbiota and metabolite strategies against intestinal KP challenge.

Microorganisms doi: 10.3390/microorganisms14091922

Authors: Ibrahim A. Alqarni Zeina S. Alkudmani Naelah A. Alghamdi Hamad Almatrudi Muath Alsaidan Yazeed Alshuweishi

Early identification of ICU patients with sepsis who are at risk of early death remains clinically important. The aspartate aminotransferase-to-platelet ratio index (APRI) is a simple laboratory-derived index, but its clinical value for very early mortality in adult ICU patients with sepsis remains insufficiently characterized. This study evaluated the association between admission APRI and short-term mortality in ICU patients with sepsis. This retrospective cohort study included 266 adults admitted to the ICU with sepsis. APRI was calculated from AST and platelet measurements obtained within 24 h of admission. Patients were categorized according to mortality timing. Associations with 7-day mortality were evaluated using logistic regression adjusted for age, sex, and comorbidity status. ROC and survival analyses were also performed. Of 266 patients, 32 died within 7 days, 88 between 8 and 28 days, and 72 after 28 days, and 74 survived. APRI was highest among patients who died within 7 days (median 1.919, IQR 0.631–5.009) compared with survivors (0.472, IQR 0.192–1.531; p = 0.0056). Higher log-transformed APRI remained associated with 7-day mortality after adjustment for age, sex, and comorbidity status (OR = 2.77, 95% CI 1.57–5.01; p = 0.0004). APRI showed modest discrimination for 7-day mortality (AUC = 0.687, 95% CI 0.580–0.793) and lower discrimination for 28-day mortality (AUC = 0.623, 95% CI 0.555–0.690). Elevated APRI was also associated with reduced short-term survival. Elevated admission APRI is independently associated with early mortality and reduced short-term survival in ICU patients with sepsis. However, its modest discrimination supports its use as an adjunct to established clinical risk assessment tools.

Microorganisms doi: 10.3390/microorganisms14091919

Authors: Ruiwen Zhang Xiazhen Yao Zhen Xing

Habitat transition can reorganize plant-associated microbial communities, yet whether rhizosphere bacteria and fungi respond similarly to the transition from native to introduced habitats remains unclear. Here, we investigated this question in the endangered plant Paeonia ludlowii by comparing rhizosphere bacterial and fungal communities across three native and two introduced sites in Xizang. Both bacterial and fungal communities showed significant site-associated differentiation and strongly concordant spatial patterns. However, their responses differed markedly: bacterial richness remained relatively stable despite compositional turnover, whereas fungal communities showed reduced richness at the Lhasa introduction site, lower OTU sharing among habitats, and significantly greater divergence from native assemblages than bacterial communities. Soil pH and available phosphorus were the environmental variables most consistently associated with differentiation in both microbial groups. These findings demonstrate that rhizosphere bacteria and fungi exhibit concordant spatial differentiation but contrasting responses to habitat transition, with fungal communities showing greater departure from native states. This contrast highlights the importance of considering bacterial and fungal communities jointly when evaluating rhizosphere reorganization during the introduction and ex situ conservation of endangered plants.

Microorganisms doi: 10.3390/microorganisms14091920

Authors: Leticia Jaimes Betancourt Alfonso Vallejos Parás Porfirio Felipe Hernández Bautista David Alejandro Cabrera Gaytán Bernardo Cacho Díaz Adriana Josefina Toríz Saldaña Lumumba Arriaga Nieto Óscar Cruz Orozco Gabriel Valle Alvarado Mónica Grisel Rivera Mahey Mónica Alethia Cureño Díaz Víctor Gómez Bocanegra Héctor Alonso Téllez Medina

Dengue is a major public health problem in Mexico, where large epidemics have occurred during recent years. Although demographic and clinical predictors of severe dengue have been widely studied, the impact of epidemic transmission periods on hospitalization and mortality remains poorly understood. This study evaluated the association between epidemic years and hospitalization and mortality among laboratory-confirmed dengue cases in Mexico. A nationwide retrospective analytical study was conducted using laboratory-confirmed dengue cases reported through the Mexican National Epidemiological Surveillance System between 2020 and 2025. Years were classified as epidemic (2023–2024) or non-epidemic (2020–2022 and 2025) according to official endemic channel analyses. Hospitalization and mortality were evaluated as primary outcomes. Multivariable logistic regression models were developed to identify factors independently associated with hospitalization and mortality. A total of 235,226 laboratory-confirmed dengue cases were included, of whom 90,661 (38.5%) were hospitalized and 1604 (0.68%) died. Overall, 170,819 cases (72.6%) occurred during epidemic years. Hospitalization was more frequent during epidemic years than during non-epidemic years (40.2% vs. 34.1%; crude OR 1.30, 95% CI 1.27–1.32; p < 0.001), whereas mortality did not differ significantly (0.68% vs. 0.69%; crude OR 0.99, 95% CI 0.89–1.11; p = 0.919). In multivariable analysis, epidemic-year status remained independently associated with hospitalization (aOR 1.26, 95% CI 1.24–1.29; p < 0.001) but not with mortality (aOR 1.00, 95% CI 0.90–1.12; p = 0.946). Chronic kidney disease showed the strongest independent association with both hospitalization (aOR 5.36, 95% CI 4.33–6.65) and mortality (aOR 6.98, 95% CI 5.23–9.32). Advanced age and other underlying comorbidities were also independently associated with adverse outcomes. Epidemic years were associated with increased hospitalization among laboratory-confirmed dengue cases in Mexico but were not associated with increased mortality. Mortality was more strongly associated with age and underlying comorbidities, particularly chronic kidney disease. These findings highlight the importance of strengthening hospital preparedness during epidemic periods while prioritizing clinical monitoring of high-risk patients.

Microorganisms doi: 10.3390/microorganisms14091918

Authors: Salar Ali Salvatore Dimonte Stefano Aquaro Muhammed Babakir-Mina Giovanni Di Bonaventura Arianna Pompilio

Carbon monoxide-releasing molecules (CORMs) display promising antibacterial activity, yet their global cellular targets and underlying mechanisms remain incompletely understood. This study aimed to characterize the proteomic response of Escherichia coli MG1655 exposed to a sub-inhibitory concentration (10 µM, 40 min) of the ruthenium-based CORM-3 compared to its inactive counterpart, iCORM-3, in defined minimal medium. Utilizing label-free quantitative LC-MS/MS proteomics combined with differential abundance, functional annotation, and STRING-based network analyses, we explored treatment-associated proteomic signatures and subcellular localization patterns. While iCORM-3 exhibited no antibacterial activity, CORM-3 inhibited growth in a concentration-dependent manner. Although no individual protein reached FDR significance after multiple-testing correction, candidate abundance changes converged on functionally related modules, including envelope/periplasmic stress, sulfur metabolism and transport, redox/metal homeostasis and selected energy-associated functions. These findings support a hypothesis-generating model in which sub-inhibitory CORM-3 exposure is associated with multifactorial bacterial stress adaptation rather than a single dominant protein-level target. Envelope homeostasis, sulfur-containing pathways, redox/metal adaptation, and attenuation of selected energy-associated functions therefore emerge as candidate components of the E. coli response to CORM-3 and require future targeted validation.

Microorganisms doi: 10.3390/microorganisms14091917

Authors: Camila Tassano Matías Javier Garavaglia Marcos Leopoldo Esteso Catalina Basile Dazzi Juan Orlowski Walter Patricio Mac Cormack Jaco Vangronsveld Sofie Thijs Lucas Adolfo Mauro Ruberto Francisco Massot

More than six decades of human activity in Antarctica resulted in chronic diesel contamination of soils surrounding research stations. Phytoremediation assisted by the native vascular plant Deschampsia antarctica is one of the few remediation strategies compatible with the Antarctic Treaty System guidelines. Securing suitable root-associated microbial resources is central to this approach. This work characterized the bacterial communities of the rhizosphere and root endosphere of D. antarctica growing in a chronically diesel-contaminated soil at Carlini Station and in three pristine sites on 25 de Mayo (King George) Island, South Shetland Islands, combining culture-independent 16S rRNA gene amplicon sequencing, PICRUSt2 functional prediction and culture-dependent bacterial strain isolation. The rhizospheric microbial community was consistently more diverse than those inhabiting the endosphere and was more strongly structured by site and associated soil physicochemical variables, with the contaminated site showing the most distinct composition, whereas the endosphere remained comparatively stable and host-selected. The community associated with the chronically contaminated site showed pronounced compositional differences and higher predicted abundances of hydrocarbon-degradation pathways in the rhizosphere. Polaromonas, Rhodococcus, Mycobacterium and Devosia were repeatedly associated with the community from the contaminated site across taxonomic, biomarker and predicted functional analyses. Members of the genera Polaromonas, Rhodococcus and Devosia were also recovered in culture. These taxa represent suitable candidates for the future development of microbe-assisted phytoremediation strategies in Antarctica.

Microorganisms doi: 10.3390/microorganisms14091916

Authors: Liang Cui Shasha Wang Xuewen Gao Rongfeng Hong Zongze Shao Lijing Jiang

In deep-sea hydrothermal ecosystems, inorganic sulfur compounds serve as key energy sources for microbes through oxidation, reduction, or disproportionation reactions. However, to date, the bacteria that disproportionate sulfur remain poorly understood. Here, we characterized the physiological and metabolic characteristics of Sulfurovum sp. MH2-6, which was isolated from hydrothermal sediments of the South Mid-Atlantic Ridge. Based on the results of 16S rRNA gene sequence, average nucleotide identity, and DNA–DNA hybridization value, strain MH2-6 belonged to the same species as Sulfurovum mangrovi ST1-3T. The isolate was able to grow chemolithoautotrophically using thiosulfate, sulfite, or sulfide as the sole energy source, and molecular oxygen as the sole electron acceptor. When using hydrogen as the sole energy source, this bacterium could utilize a wide range of electron acceptors, including oxygen, elemental sulfur, thiosulfate, nitrate, and sulfate. Various organic compounds also supported growth as carbon sources during hydrogen oxidation, suggesting a potential for chemolithomixotrophy. Notably, the isolate could grow via the disproportionation of thiosulfate and elemental sulfur in the presence of ferrihydrite. Further, genome analyses revealed that this bacterium contains a complete reductive citric acid cycle (rTCA) for carbon fixation, multiple hydrogenases, sulfur oxidation, reduction, and transfer enzymes, nitrogenase, and oxygen reductases. Transcriptomic comparisons between sulfur reduction and disproportionation conditions revealed that thiosulfate reductase, type IV sulfide: quinone oxidoreductase, and sulfite dehydrogenase were highly abundant in thiosulfate-disproportionating cultures, while rhodanese-like sulfurtransferases and sulfide dehydrogenase showed increased abundances when grown via elemental sulfur disproportionation. Together, these flexible energy- and carbon-utilizing strategies may enhance the persistence of MH2-6 in hydrothermal vent environments.

Microorganisms doi: 10.3390/microorganisms14091915

Authors: Felipe da Silva Valente Karla Renata Kaminski Andrioli Marcus Adonai Castro da Silva André Silva Barreto

This decade-long surveillance study (2016–2025) investigates the acquisition of multidrug-resistant (MDR) bacteria in South Atlantic cetaceans to evaluate how ecological niches modulate exposure to biological pollution. Analyzing clinical isolates from stranded cetacean carcasses (n = 346), we used Generalized Linear Mixed-Effects Models (GLMMs) to mitigate multi-center analytical biases and compare resistance profiles across coastal and oceanic species. We observed a significant, progressive upward trend in the overall MDR probability over the decade. Although raw MDR was higher in the demersal-feeding Pontoporia blainvillei (58.2%) than in the sympatric, water-column-foraging Sotalia guianensis (22.0%), multivariate modeling revealed that this difference was primarily driven by the geographic stranding location rather than by intrinsic foraging ecology. High gastrointestinal MDR (73.9%) suggests dietary intake as a primary biological gateway. Demographic modeling revealed a significant sex-based association in P. blainvillei, with females facing a higher risk (p = 0.009), although the specific ecological or physiological mechanisms underlying this difference remain unknown. High MDR rates in deep-diving Lagenodelphis hosei (70.8%) and Kogia breviceps (67.9%) suggest that resistant pathogens may reach bathypelagic food webs, potentially via vertical trophic pathways. These findings suggest that spatial environmental contamination, alongside foraging and reproductive ecologies, is a key driver of exposure to the anthropogenic resistome. Because carcass-based sampling inherently targets a diseased or senescent fraction, these high prevalences may overestimate the resistome burden of healthy free-ranging populations. The detection of human pathogens across coastal and offshore habitats indicates persistent deficiencies in terrestrial effluent management, reinforcing cetaceans as One Health sentinels.

Microorganisms doi: 10.3390/microorganisms14091914

Authors: Xionglian Jin Huichun Xie Xiaoping Kong Jiawei Yan Yonggui Ma Zhe Chen Feng Qiao

Biological soil crusts (BSCs) are critical components of alpine sandy land ecosystems, yet how their diazotrophic communities respond to plantation type and successional stage remains largely unknown. This study examined diazotrophic community differentiation in BSCs under four sand-fixing plantations (Salix psammophila SL, Caragana korshinskii NT, Salix cheilophila WL, Populus simonii XYY) across three successional stages in an alpine sandy land. Soil properties, microbial biomass, and enzyme activities were measured, along with nifH gene sequencing (Illumina), to characterize community composition and its relationships with environmental factors along BSCs succession. The results indicate that Skermanella, Mastigocladus, and Nostoc were the dominant diazotrophic groups in the study area. The Nostoc was the dominant genus across all plantation types, with the highest average relative abundance (37.92%) recorded in moss crusts under the XYY plantations. Diazotrophic α-diversity increased with BSCs succession, and β-diversity analysis (PCoA with ANOSIM) demonstrated significant community differentiation among plantation types (variance explanation = 50.91%, R = 0.59458, p = 0.001). Spearman’s and redundancy analysis (RDA) showed that sand-fixing plantations drive the differentiation of diazotrophic communities by regulating soil physicochemical properties and microbial metabolism. Furthermore, the diazotrophic communities in moss crusts exhibited stronger environmental responsiveness than those in algae crusts. In the COG functional categories, J (translation and ribosomal biogenesis, 7.92%), E (amino acid metabolism, 10.14%), and C (energy production, 7.11%) were identified as the core foundational functions of BSCs in the study area. The relative abundances of these functional categories showed convergence. In summary, different plantations were associated with distinct diazotrophic community compositions, and these associations were largely mediated by soil environmental factors. Concurrently, BSC succession covaried with enhanced differentiation in community structure and functional traits. These patterns are consistent with a stepwise response cascade linking vegetation and soil properties to diazotrophic communities and nitrogen metabolism functions. This study provides a theoretical basis for enhancing the stability and long-term restoration of artificial ecosystems by regulating diazotrophic communities in BSCs in alpine sandy land.

Microorganisms doi: 10.3390/microorganisms14091913

Authors: Mark L. Cannon John Peldyak Paul R. Reynolds Gustavo Ferrer

The oral cavity is a mucosal and mineralized interface shared by the digestive tract and the upper airway. This narrative review proposes the oral–nasal gateway microbiome as a clinically useful model for understanding oral, nasal, and systemic health. The model includes bacteria, fungi, archaea, protozoa, viruses, bacteriophages, microbial metabolites, and host-derived salivary components. Its gateway role is supported by anatomy, continuous salivation, periodontal vascular exposure, oral–gut microbial overlap, nitrate–nitrite–nitric oxide biology, oral and nasal airway interactions, maternal–child microbial transmission, and enrichment of oral organisms in selected distal diseases and tumors. Oral communities respond rapidly to diet, salivary flow, airway physiology, smoking and vaping, xerostomic medications, antibiotics, and antiseptic rinses, and these changes may influence the nasal microbiome. Published evidence summarizes bacterial pathobionts and protective commensals; Candida and other oral fungi; herpesviruses; papillomaviruses; bacteriophages; salivaomics; pregnancy and early-life prevention; probiotics; polyols; remineralization chemistry; environmental exposures; and tumor microbiology. As of manuscript preparation, SalivaDB catalogs 15,821 salivary biomarker entries across 201 diseases and 48 disease categories. The practical endpoint is not sterilization of the oral cavity but restoration of microbial homeostasis, salivary competence, airway stability, dietary balance, and biologically informed, timely prevention.

Microorganisms doi: 10.3390/microorganisms14091912

Authors: Shufeng Weng Qingchun Li Yamin Zhao Taiyue Lin Zihan Wang Mingrui Zhu Miaochengyue Xin Ying Xu

Mycobacterium tuberculosis (M. tb) possesses a unique, lipid-rich cell envelope that is critical for virulence, drug resistance, and persistence. Here, we identify the PPE family protein PPE57 as a key regulator of mycolic acid transport and host lipid exploitation. PPE57 physically interacts with the essential lipid transporter MmpL3, promoting trehalose monomycolate (TMM) translocation, enhancing trehalose dimycolate (TDM) synthesis, and increasing cell wall lipid content. Site-directed mutagenesis identified G175 as a critical residue for PPE57-MmpL3 binding. Deletion of PPE57 reduces cell wall thickness, alters lipid composition, and impairs biofilm formation. Mechanistically, PPE57 facilitates bacterial cholesterol acquisition, and during infection, activates the host PPAR-γ pathway in macrophages, leading to enhanced cholesterol uptake, lipid droplet accumulation, and increased intracellular triglyceride and cholesteryl ester levels. These changes provide a nutrient-rich niche that promotes bacterial survival and persistence. In a mouse model of infection, PPE57 deficiency results in reduced bacterial burden, milder lung pathology, and diminished lipid droplet-positive cell accumulation in lung tissues. These findings establish PPE57 as an essential accessory component of the MmpL3 lipid transport system, bridging mycobacterial cell wall assembly with host nutrient exploitation during persistent infection.

Microorganisms doi: 10.3390/microorganisms14091911

Authors: Meng-Ling Deng Yan Zhang Man-Li Jiang Tong Zhang Jun Ma Jian-Fa Yang Feng-Cai Zou Jun-Jun He Lu-Yang Wang

Ticks are hematophagous arthropods that parasitize livestock, humans, and wild animals, posing serious threats to public health worldwide. Diverse pathogens have been discovered in ticks, including viruses, bacteria, protozoa, and filarial nematodes; however, ticks also harbor a variety of fungal microbes, which remain largely unexplored. In this study, we isolated and identified culturable fungi associated with ticks collected on cattle and goats in Yunnan Province, China. Of the 1730 ticks obtained, 1690 were identified as Rhipicephalus microplus, and only 40 as Haemaphysalis longicornis via morphology and 16S rRNA gene analysis. A total of 90 fungal strains were isolated from the ticks and identified by morphological examination and ITS sequencing, representing 25 species across 20 genera, with Fusarium being the most prevalent. Among these isolates, Fusarium verticillioides (19 isolates) poses a threat to grains, animals, and humans; Cladosporium cladosporioides (six isolates) and Sarocladium zeae (one isolate) are primarily pathogenic to plants, while Schizophyllum commune (eight isolates), Rhizopus arrhizus (two isolates), Diaporthe phaseolorum (two isolates), and Purpureocillium lilacinum (two isolates) are recognized animal-associated pathogens. The diversity of isolated fungi with documented pathogenicity suggests that ticks may serve as potential vectors for fungal transmission, raising concerns about their role in cross-kingdom disease spread across agriculture and livestock. Furthermore, fungi with entomopathogenic potential, such as P. lilacinum, offer valuable resources for developing biocontrol agents against ticks. Our findings highlight the dual role of ticks as both potential vectors of phytopathogenic fungi and reservoirs of entomopathogenic biocontrol agents. This necessitates a paradigm shift in tick surveillance, integrating mycological monitoring to mitigate agricultural losses and exploring tick-borne fungi for sustainable vector control strategies.

Microorganisms doi: 10.3390/microorganisms14091910

Authors: Peng Zhang Lei Ji Wei Yan Liping Chen Fenfen Dong

Vibrio parahaemolyticus is a leading cause of seafood-associated gastroenteritis worldwide. Traditional serotyping based on 13 O and 71 K antigens fails to classify many isolates, designated as K-untypable (KUT), whose genetic basis and evolutionary dynamics remain unclear. In this study, we conducted whole-genome sequencing of 47 O4:KUT strains from 40 clinical and 7 retail aquatic products (snail, river shrimp, Macrobrachium rosenbergii, etc.) to decipher the genomic diversity and structural variation in their O/K antigen loci. Phylogenomic analysis revealed a polyphyletic population structure spanning multiple sequence types, with O/K antigen loci classified into 13 distinct structural types comprising 149 biosynthetic genes. These clusters exhibited mosaic architectures and varied functional profiles. Notably, pervasive phylogenetic incongruence and robust recombination signals identified horizontal gene transfer as the primary mechanism driving O/K antigen loci diversification. Our findings reveal the evolutionary mechanisms of the foodborne O4:KUT serotype prevalent in local seafood and clinical samples from Huzhou, China, and provide a preliminary gene-signature framework that may inform the future development of molecular serotyping assays.

Microorganisms doi: 10.3390/microorganisms14091906

Authors: Wahauwouélé Hermann Coulibaly Tano Marie-Ange Sakia Mian Yabo Majoie Géroxie Tohoyessou Muiz O. Akinyemi Bassey Ebenso Ange Olivier Parfait Yao Cécile Meex Paul-Alexandru Popescu Thierry Fievez Phillipe Maesen Hary Razafindralambo

Aquaponics combines aquaculture and hydroponics, offering an integrated and sustainable food production system. This study investigated the probiotic properties, plant growth-promoting (PGP) activity, and nitrifying capacity of twelve lactic acid bacteria (LAB) strains isolated from an aquaculture farm environment on the Ivory Coast for their potential application in aquaponic systems. All isolates demonstrated antagonistic activity against key pathogenic indicator strains, except Vibrio cholerae, and displayed varying levels of surface hydrophobicity (5.50 ± 0.70% to 22.83 ± 1.17%) and auto-aggregation (32.50 ± 0.08% to 52.89 ± 0.39%) after 24 h. Antioxidant activity was significantly higher in cell-free supernatants (~71–79%) than in intact cells (~30–33%). Bile salt tolerance (0.3%, 4 h) ranged from 2.13 ± 0.76% to 40.87 ± 2.12%, and survival under pH 1.5 with pepsin for 3 h varied from 4.84 ± 0.26% to 53.98 ± 13.28%. All isolates produced lactic, acetic, citric, malic, and propionic acid and exhibited amylase and cellulase activity; none showed hemolytic activity. Only LAB 11 produced indole-3-acetic acid (17.72 ± 0.06 μg/mL), siderophores, and phosphate-solubilization activity for PGP traits, and this group significantly enhanced maize seed germination (86.66 ± 5.77%) and radicle length (7.00 ± 0.52 mm) compared to the control (63.33 ± 32.14% and 4.94 ± 1.52 mm), respectively. LAB 1 and LAB 10 demonstrated the highest ammonia-oxidizing capacity in vitro and in trout pond water. LAB 1, LAB 10, and LAB 11 were confirmed by whole genome sequencing analysis to be Enterococcus faecalis strains with a favorable-safety genomic profile and probiotic characteristics. These three strains therefore represent promising candidates for consortium-based applications in aquaponics systems.

Microorganisms doi: 10.3390/microorganisms14091908

Authors: Eugen Mihail Arnautu Elena Leocadia Plesea Ovidiu Mircea Zlatian Georgiana Cristiana Camen Radu Razvan Mititelu Andrei Osman Maria-Loredana Tieranu Andreea Loredana Golli Alice Elena Ghenea

Colistin resistance is an increasing concern in critically ill adults, but reported rates, clinical consequences, laboratory detection, and resistance mechanisms vary widely between studies. This systematic review evaluated epidemiological, clinical, laboratory, and molecular evidence on colistin resistance in adult intensive care populations. MEDLINE/PubMed, the Cochrane Library, Scopus, and Web of Science Core Collection were searched, and the findings were synthesised descriptively because of substantial differences between studies. A total of 120 studies were included. In studies using BMD in broader ICU populations, colistin non-susceptibility ranged from approximately 0.3–57.4% at the isolate level, while patient-level carriage/acquisition estimates ranged from approximately 1.2% to 9.4%. Clinical studies often reported poorer outcomes in patients with colistin-resistant infections. However, the strength and direction of these associations were inconsistent, and most comparative analyses had substantial risk of bias. Laboratory methods influenced resistance detection, while heteroresistance required specific testing approaches. Molecular studies identified several resistance pathways, including mcr genes, mgrB disruption, regulatory alterations, clonal spread, and within-host evolution. Overall, colistin resistance in critical care is highly heterogeneous and should be interpreted according to the clinical, microbiological, and epidemiological context.

Microorganisms doi: 10.3390/microorganisms14091909

Authors: Damir I. Khasanov Natalia L. Rudakova Iuliia V. Danilova Iuliia A. Vasileva Aigul I. Gilmutdinova Margarita R. Sharipova

The Bacillus pumilus 3-19 strain exhibits increased secretion of various hydrolases, including proteases, and is a promising plant growth-promoting (PGP) agent. A unique secreted minor metalloproteinase, MprBp, which has no homologs among prokaryotic enzymes, has been identified in its genome. The role of the enzyme in B. pumilus cells is poorly understood. Using CRISPR/Cas9 genome editing, a deletion mutant of B. pumilus 3-19 with an inactivated mprBp gene was obtained. Comparative analysis of the mutant and native strains by quantitative reverse transcription PCR (RT-qPCR) revealed a significant decrease in the expression of other minor proteases. The mutant strain also demonstrated a two-fold increase in biofilm formation, suggesting that MprBp acts as a negative regulator of this process. Additionally, seed treatment with B. pumilus ΔmprBp led to an increase in the morphometric parameters of barley seedlings (Hordeum vulgare L.).

Microorganisms doi: 10.3390/microorganisms14091907

Authors: Rui Li Chi Zhang Yu Miao Fangze Li Ge Zhang Qiwei Sun Tianci Hua Zhikun Pang Xingjie Lin

Salt-affected soil constrains maize establishment, and rhizosphere and non-rhizosphere responses to soil biochar remain difficult to establish. We evaluated soil physical and nitrogen properties, qPCR marker genes, 16S and ITS communities, and molecular profiles of dissolved organic matter (DOM) in a 90-day randomized complete block pot experiment (four biochar rates; five blocks; 20 pots). Control maize did not survive; control pots therefore yielded only non-rhizosphere soil, whereas rhizosphere and non-rhizosphere samples from biochar pots were paired. Block-adjusted comparisons showed higher water content and porosity and lower bulk density in all biochar non-rhizosphere groups than in the control (Holm-adjusted p < 0.05). Biochar-associated shifts in mineral-N partitioning and selected DNA-level marker-gene abundances accompanied design-aware community differences for NR treatment, R dose and pooled compartment (16S R2 = 0.360, 0.435 and 0.218; ITS R2 = 0.720, 0.712 and 0.304; all p < 0.001); fungal differences also included heterogeneous dispersion. D90 FT-ICR MS profiles described relative DOM molecular variation, but cross-layer residual associations did not survive false-discovery rate correction. Thus, biochar-associated physical and nitrogen changes coincided with microbial and relative DOM restructuring under the tested pot conditions. Because control survival was confounded with treatment, these are observed-group associations rather than pure causal biochar effects.

Microorganisms doi: 10.3390/microorganisms14091905

Authors: Mingxuan Che Jingbo Zhang Kunduziayi Kudelaiti Jiajun Zhang Yunhao Liusui Zhengwu Dong

Long-term excessive nitrogen application in arid cotton fields increases nitrate leaching risk during fallow and disrupts rhizosphere microecology. To clarify the cross-growth-stage regulatory effects of the biostimulant γ-polyglutamic acid (γ-PGA) combined with chemical fertilizer on rhizosphere microbial communities, we compared chemical fertilizer alone (NK) and γ-PGA plus chemical fertilizer (GT) using rhizosphere soils collected at boll-setting (August) and fallow (October), with physicochemical measurements and metagenomic sequencing technology. At boll-setting, GT lowered pH by 0.74 units compared with NK and increased NH4+-N, NO3−-N, and TN by 339.3%, 491.4%, and 23.0%, respectively. By fallow, GT increased TOC by 70.6% and maintained NH4+-N at 18.38 mg/kg, while NK accumulated 66.85 mg/kg NO3−-N. GT buffered post-harvest fungal community disturbance (Shannon: GT 4.06 vs. NK 2.80) and shifted bacterial communities toward oligotrophic taxa and archaea toward ammonium-preferring taxa. A metagenomic LEfSe analysis showed that GT was enriched in functional genes related to [Q]: Secondary metabolite biosynthesis, transport and catabolism, [T]: Signal transduction mechanisms, and [V]: Defense mechanisms, indicating a shift from resource acquisition to conservative maintenance. Mantel tests revealed that microbial functional profiles showed the strongest association with NH4+-N (r = 0.828 in August, r = 0.883 in October, p < 0.001). Thus, γ-PGA with chemical fertilizer stabilizes fallow rhizosphere microbial communities, reduces nutrient leaching, and promotes carbon–nitrogen co-retention.

Microorganisms doi: 10.3390/microorganisms14091904

Authors: Ziqiao Zhao Yixin Li Yunhan Li Chengshuo Shen Youyu Liu Peng Zhang

Prophages are widespread in bacterial genomes and can influence pathogen evolution by modulating virulence, antimicrobial resistance, environmental adaptation, and competitive fitness. However, prophage research in aquaculture pathogens remains limited, with abundant genomic predictions but scarce functional validation, numerous cross-sectional surveys but little longitudinal tracking, and extensive phenomenological observations but insufficient mechanistic investigation. This review synthesizes current knowledge of prophages in aquatic bacterial pathogens, focusing on their roles in virulence, antimicrobial resistance, and competitive advantage, while drawing on mechanistic paradigms established in human and animal pathogens. We further propose a pathogen-centered “molecular bridge” framework in which environmental perturbations alter bacterial physiological states, are interpreted through prophage regulatory mechanisms, and ultimately translate into changes in pathogen phenotypes, fitness, and pathogenic potential. Particular attention is given to multi-signal interactions, induction thresholds, and the context-dependent fitness consequences of prophage induction in dynamic aquaculture environments. Finally, we propose future priorities encompassing systematic prophage resource construction, functional validation, multi-stressor experiments, and longitudinal monitoring. By adopting a paradigm-borrowing review approach, this review aims to provide a transferable roadmap for prophage research in aquaculture, to facilitate translational applications, and to offer theoretical support for precision disease control in aquaculture.

Microorganisms doi: 10.3390/microorganisms14091903

Authors: Patricia Vargas-Gutiérrez José Augusto Ramírez-Trujillo Ana Busto-Ulloa Luis Lozano-Aguirre Juan Téllez-Sosa Paola Bocanegra-Ibarias Gabriel Lizama-Uc Ismael Hernández-Lucas Ramón Suárez-Rodríguez Josefina Duran-Bedolla Humberto Barrios-Camacho

Antimicrobial resistance (AMR) is one of the main public health problems that also affects environmental ecosystems. Sinkholes are understudied groundwater ecosystems that may act as reservoirs and dissemination routes for AMR. This study evaluated the composition, distribution, and antimicrobial resistance profiles of culturable ampicillin-resistant Gram-negative bacteria recovered from sinkholes of the Yucatán Peninsula, Mexico. Water samples were collected from 27 sinkholes during dry and rainy seasons. The isolates were identified by MALDI-TOF MS and evaluated by broth microdilution. Representative E. coli isolates were analyzed by whole-genome sequencing. A total of 193 ampicillin-resistant Gram-negative isolates were recovered. The most frequently recovered genera were Enterobacter, Klebsiella, and Escherichia, which were detected across all hydrogeological zones. Resistance to cefotaxime, levofloxacin, gentamicin, and tetracycline was detected among several bacterial genera. Opportunistic bacterial species associated with healthcare-associated infections, including K. pneumoniae, A. baumannii, and P. aeruginosa, were also identified. Genomic analysis of 19 E. coli isolates revealed resistance determinants associated with multiple antimicrobial classes and a diverse population structure dominated by phylogroup B1. These findings show that the Ring of Cenotes aquifer harbors diverse populations of culturable ampicillin-resistant Gram-negative bacteria and emphasize the importance of groundwater ecosystems as environmental reservoirs of AMR.

Microorganisms doi: 10.3390/microorganisms14091902

Authors: Yunqi Huang Mi Hu Huiting Zhu Qianwen Sha Qiaomei Wang Qiongfen He Xiujuan Deng Wenxia Yuan Yihu Guan Niuniu Shi Yapeng Li Wendi Zhang Baijuan Wang Xinghua Wang

Organic fertilizer and season are established sources of variation in tea plantation soil microbiomes. A repeated-measure field experiment was conducted in Yunnan, China, comprising an unfertilized control (CK), sheep manure (SM), pig manure (PM), rapeseed cake manure (RM), and commercial organic fertilizer (OF). The same 15 mature tea trees were sampled in spring, summer, and autumn, yielding 45 composite soil samples. Soil properties, as well as 16S rRNA gene and ITS amplicon profiles, were evaluated using mixed-effect models, restricted-permutation PERMANOVA, and design-adjusted partial redundancy analysis (pRDA). Fertilization regime, season, and their interaction showed property-specific associations, most clearly for alkali-hydrolyzable nitrogen, available phosphorus, and available potassium. Bacterial alpha diversity was more responsive than fungal alpha diversity. Both factors were significantly associated with bacterial and fungal community structures. Season accounted for most bacterial variation (68.70%), whereas fungal variation was associated more evenly with season (27.33%) and fertilization regime (18.61%); no significant interaction was detected for either group. Nine fungal genera retained significant treatment-associated differences after FDR correction, whereas the bacterial genus-level patterns remained exploratory. After conditioning for fertilization regime and season, measured soil variables collectively explained 6.51% and 13.56% of bacterial and fungal variation, respectively, but no individual variable remained significant after FDR correction. Because fertilizer inputs were not nutrient-standardized, treatment contrasts represent integrated material-and-dose regimes rather than isolated fertilizer-type effects. These context-specific results illustrate the value of repeated seasonal sampling with design-adjusted analyses for evaluating practical fertilization regimes.

Microorganisms doi: 10.3390/microorganisms14091901

Authors: Huilin Yan Panda Ye Huigang Pei Jiajia He Yujiao Liu

This study characterized regional variation in faba bean rhizosphere bacterial communities across a broad survey of the Qinghai Plateau and examined their associations with soil and spatial factors. Rhizosphere soil samples were collected from 15 faba bean-growing field sites across Haidong City, Xining City, and Hainan Tibetan Autonomous Prefecture, Qinghai Province, China, spanning elevations from 1842.9 to 3206.3 m. Nine within-site subsamples were collected at each field site, with the field site treated as the independent unit of replication. Soil physicochemical properties were measured, and bacterial communities were characterized by high-throughput sequencing of the 16S rRNA gene. At the site level, bacterial alpha-diversity indices showed numerical variation among the three regional elevation groups, but did not differ significantly. Bacterial community composition differed among groups in the site-level PERMANOVA based on Bray–Curtis dissimilarities (pseudo-F2.12 = 1.789, R2 = 0.230, p = 0.031), although substantial overlap among sites remained. However, a conservative locality-level sensitivity analysis was not significant (p = 0.060). Geographic distance was significantly associated with bacterial community dissimilarity, whereas elevation difference showed no detectable independent association after controlling for geographic distance, indicating a substantial contribution of spatial structure to the observed community differentiation. Dominant phylum-level composition was broadly conserved across sites, while genus-level relative abundances showed descriptive spatial variation. Neither RAVD-based compositional consistency nor community-level niche breadth differed significantly among groups. Among the measured soil properties, only pH differed significantly, whereas most nutrient variables did not. Overall, the study revealed spatially structured regional differentiation in faba bean rhizosphere bacterial communities across the Qinghai Plateau. The observed patterns likely reflect the combined influence of geographic structure, elevation, background soil conditions, agricultural management, plant phenology, and other unmeasured environmental factors, which could not be independently disentangled in the present survey.

Microorganisms doi: 10.3390/microorganisms14091900

Authors: Xiaoyu Chen Xinye Chen Jinrong Shou Jiaojiao Zhang Jiening Zou Xinyi Bao Zonghe Yu Xiaoyong Zhang

While the gut bacterial communities of sea cucumbers have been extensively studied, the extent to which environmental filtering may shape bacterial assembly across different host compartments remains poorly understood. This study employed Illumina MiSeq high-throughput sequencing technology to investigate the bacterial communities in environmental sediments (E), body epidermis (B), foregut (F) and hindgut (H) samples of Holothuria leucospilota (n = 4 per group) from the South China Sea. A total of 749,222 high-quality 16S rRNA sequences were dereplicated into 911 amplicon sequence variants (ASVs). Alpha diversity revealed that environmental sediments exhibited the highest diversity among the four samples (Shannon index: 7.33 ± 0.11). Beta-diversity analysis revealed pronounced differentiation in bacterial community structure among sample types. Proteobacteria, Bacteroidetes, and Firmicutes were the dominant bacterial phyla. The bacterial taxonomic composition across compartments of H. leucospilota varied considerably. Woeseia was not detected from host niches but was present only in environmental sediments. The genus Vibrio dominated in environmental sediments and foregut samples. The epidermal bacterial communities were characterized by the prevalence of the NS11-12 marine group, Rhodobacteraceae, and Rhizobiaceae. The distinct successional gradient observed in Woeseia, Vibrio, and Pseudomonas suggested that bacterial colonization of the gut is consistent with selective environmental filtering. Functional annotation (KEGG) indicated dominance in core metabolic pathways, particularly carbohydrate and amino acid degradation, revealing complementary metabolic functions among dominant taxa with the potential for metabolic niche partitioning to support host energy harvest. Our findings provide a baseline framework that may assist in future screening of probiotic candidates and disease management strategies, pending experimental validation of the functional roles of the identified taxa.

Microorganisms doi: 10.3390/microorganisms14091899

Authors: Luis Adrián De Jesús-González Ignacio Martínez Jorge Procopio-Velázquez Muslim Schabib-Hany Bertha Espinoza

Chagas disease is difficult to diagnose in the chronic phase, especially when serological assays disagree. We examined whether Trypanosoma cruzi metacyclic trypomastigotes’ antigenic extracts may increase ELISA/Western blot concordance in initially discordant serum compared to epimastigotes. A total of 178 human blood samples were studied, including seropositive persons, Chagasic cardiomyopathy patients, discordant sera, seronegative controls, and sera from various parasitic illnesses. Antigenic extracts from epimastigotes and metacyclic trypomastigotes were extracted using sonication or urea, and Western blot and ELISA were used to compare their behavior. Sonicated metacyclic trypomastigote extracts produced concordant ELISA/Western blot results in 31/35 originally discordant sera, compared to 19/35 for sonicated epimastigotes, 20/35 for urea-extracted trypomastigotes, and 17/35 for urea-extracted epimastigotes. All extracts in the serum panel previously classified using the standardized in-house serological platform showed 100% positive percent agreement, but negative percent agreement varied between 93.3% for sonicated metacyclic trypomastigotes and 66.7% for urea-extracted epimastigotes. These estimates reflect agreement with the previous serological classification, not diagnostic accuracy versus an independent reference standard. Cross-reactivity was mainly observed in sera from patients with leishmaniasis, while sera from the cardiopathic group showed an exploratory preferential recognition pattern with bands of 18, 22, 24, and 26 kDa in the sonicated metacyclic trypomastigote preparation. The comparison did not account for potential clinical variables or molecularly identify these bands. These data demonstrate that stage-informed antigen selection, particularly using sonicated metacyclic trypo-mastigote extracts, can affect serum panel assay concordance and nonspecific reactivity.

Microorganisms doi: 10.3390/microorganisms14091898

Authors: Yidan Zhang Yu Zhang Shengnan Song Jing Zhang Zhihua Sun Xia Zhou Chuangfu Chen Jia Guo Hui Zhang

Brucella spp. is a bacterium that can survive under conditions of nutrient starvation and is responsible for foodborne illnesses. The OmpR-type transcriptional regulator is characterized by an N-terminal receiver domain and a C-terminal domain, and it plays a regulatory role in diverse physiological processes, notably nitrogen and carbon metabolism. The genome of Brucella melitensis 16M contains genes for multiple OmpR-family regulators. However, the genetic program associated with nitrogen metabolism remains elusive. Herein, it was demonstrated that the B. melitensis 16M ΔftcR mutant, which lacks the OmpR-type regulator FtcR, displayed compromised viability upon recovery in a nitrogen- and carbon-free medium. Chromatin immunoprecipitation and next-generation sequencing were used to characterize the DNA-binding sites of FtcR. Our genome-wide analysis revealed extensive FtcR-binding sites throughout the B. melitensis 16M genome, including the identification of glnB as a novel target. As glnB encodes a P-II nitrogen regulator, this suggests FtcR plays a direct role in modulating nitrogen, carbon, and energy metabolism under prolonged nutrient starvation. In summary, our findings not only advance our understanding of the transcriptional regulation of nitrogen metabolism but also highlight its critical role in brucellosis pathogenesis, thereby paving the way for the development of novel therapeutics.

Microorganisms doi: 10.3390/microorganisms14091896

Authors: Zhilian Gong Lingshan Li Yajie Li Xiao Zhang Sidan Gong Yong Li

Microbial degradation of phthalate esters (PAEs) is regarded as a highly promising green remediation strategy for PAE-contamination, but it still remains constrained by the scarcity of efficient microbial resources capable of degrading PAEs completely under complex conditions. This study systematically explored microbial resources resistant to dimethyl phthalate (DMP)/dibutyl phthalate (DBP) stress along with the putative degradation mechanisms in recycled plastic sewage treatment plants through high-throughput sequencing, DMP/DBP-degrading bacteria screening, and whole-genome sequencing of representative efficient DBP-degrading strains. High-throughput sequencing result revealed substantial enrichment of PAEs-degrading bacteria in activated sludge exposed to 500 mg L−1 DMP/DBP. Further analysis demonstrated that the recycled plastic sewage treatment plant harbored rich culturable DMP/DBP-degrading bacteria, comprising 19 genera and 27 species of DMP-degrading bacteria, as well as 11 genera and 20 species of DBP-degrading bacteria. Notably, strain SWLYDMP 33, a potential novel species, along with 11 genera such as Paenirhodobacter, Ciceribacter, and Neorhizobium has been scarcely reported in association with PAEs degradation. Given its affiliation with a dominant genus and its superior broad-spectrum degradation performance, the representative efficient DBP-degrading strain SWLYDBP 51 was selected for whole-genome sequencing. Genome annotation of strain SWLYDBP 51 uncovered a wealth of functional genes implicated in DBP metabolism, and the putative complicated DBP degradation pathways were reconstructed, suggesting that SWLYDBP 51 might possess the genetic capacity for effectively degrading DBP under diverse environmental conditions. Overall, this study provided abundant efficient microbial resources and a theoretical reference for the bioremediation of PAEs contamination.

Microorganisms doi: 10.3390/microorganisms14091897

Authors: Laly Abigail Rojas Rodríguez Carlos Michell Gálvez Ramírez Marco Aurelio Salvatierra Celis Justo Nilo Balcazar Conde Margarita Fé Requena Mendizábal Rocio Del Pilar Bocanegra Arista Roger Dámaso Calla Poma Tania Valentina Rosales Cifuentes

Dental caries is the most prevalent chronic noncommunicable disease in Peruvian schoolchildren (>70%). Full-length 16S Oxford Nanopore Technology (ONT) sequencing enables species-level resolution unachievable with conventional Illumina V3–V4 platforms, critical for resolving intrageneric diversity in the dominant oral genus Streptococcus This is a cross-sectional study (STROBE/STORMS) of 30 children aged 8–10 years (15 with caries, ceod/CPOD ≥ 1; 15 caries-free, ceod/CPOD = 0) from IE N°3036 José Andrés Rázuri, San Martín de Porres, Lima, with caries status assessed by clinical examination using ICDAS-II criteria and summarized using the ceod/CPOD indices. Full-length 16S sequencing (~1500 bp; 27F/1492R; SQK-16S024) was performed on MinION MK1B (MN40465), with Flongle AUB828 in two runs (Run 1: 21,651 PASS reads, Q = 12.50; Run 2: 3750 PASS reads, Q = 11.89). The pipeline used was wf-16s v1.2.0 (EPI2ME, NCBI 16S rRNA), while for statistics, Mann–Whitney U test, Fisher’s exact test (α = 0.05), and PERMANOVA (999 permutations) were used. In total, 126 species were identified in 63 genera and 5 phyla. Streptococcus salivarius was dominant (22.5%; 30/30). Twenty-six species were exclusive to the caries group, led by S. mutans (7/15, 46.7%; OR = ∞; raw p = 0.006) and S. anginosus (4/15, 26.7%; OR = ∞; raw p = 0.038), plus Lancefieldella parvula, Veillonella infantium, and Actinomyces naeslundii. Fifteen species were exclusive to the healthy group, including Rothia aeria, Gemella sp., Aggregatibacter kilianii, and Bulleidia extructa. None of these taxon-level differences survived Benjamini–Hochberg correction for the 126 species tested (all FDR-adjusted p > 0.45); these findings are therefore presented as exploratory candidates rather than confirmed differences. Alpha diversity (Shannon p = 0.507) and global beta diversity (PERMANOVA p = 0.215) did not differ; dysbiotic convergence was significant (intragroup Bray–Curtis: 0.322 vs. 0.414; p < 0.001). This is the first ONT full-length 16S characterization of the salivary microbiome in Peruvian children, simultaneously identifying cariogenic and health-associated species profiles. These findings suggest dietary nitrate supplementation and xylitol as candidate prebiotic strategies warranting future intervention studies to restore health-associated taxa in caries-free children.

Microorganisms doi: 10.3390/microorganisms14091895

Authors: Liang Guo Yinghan Liu Yijun Weng Liangliang Hu Tan Ke Yuqin Mao Yin Lu

The invasive golden apple snail (Pomacea canaliculata) threatens rice agroecosystems, yet its direct density-dependent effects on flooded paddy soil biogeochemistry and bacterial communities remain unclear. We established flooded soil microcosms with four snail densities (0, 2, 4, and 6 snails/box) for 20 days, without external food inputs. Soil dissolved organic carbon (DOC), ammonium nitrogen (NH4+-N), nitrate nitrogen (NO3−-N), and the activities of β-glucosidase, N-acetyl-β-D-glucosaminidase, urease, and dehydrogenase were measured, and bacterial communities were characterized by full-length 16S rRNA gene amplicon sequencing. Snail density was significantly and positively related to all three nutrient variables and all four enzyme activities. The dominant bacterial phyla and genera remained stable, and bacterial α-diversity changed little among treatments, despite a small but significant increase in Simpson diversity in the high-density treatment. PERMANOVA detected significant differences in overall bacterial community structure among density treatments, while environmental fitting identified DOC, urease, and dehydrogenase as variables significantly associated with community variation. These findings indicate that living golden apple snails can alter nutrient availability, soil biochemical activity, and bacterial community organization in flooded paddy soil, revealing a belowground pathway through which this invader may influence paddy ecosystem functioning.

Microorganisms doi: 10.3390/microorganisms14091894

Authors: Luís Fernandes Diana S. Paiva Emília Pereira Célia Cabral Nuno Mesquita António Portugal

Fungal organisms play a major role in the contamination and biodeterioration of natural history museum collections and cultural heritage as a whole, leading to the potential loss of key knowledge preserved by these. As part of the ongoing efforts to study the fungal contaminations observed in the Science Museum of the University of Coimbra (Portugal) collections, eight taxidermized birds were sampled and 27 fungal isolates were retrieved. A comprehensive analysis of these isolates led to the identification of 14 different species belonging to 13 different genera, with Aspergillus accounting for 37% (1 species), Talaromyces for 11.11% (1 species); Cladosporium, Paramicrodochium, and Periconia accounting for 7.4% each (2, 1, and 1 species, respectively); and Arcopilus, Beauveria, Coprinellus, Hyphodermella, Mycoaciella, Neokalmusia, Paraeutypella, and Penicillium collectively accounting for 3.7% each, all represented by a single species. Among these were two isolates of an unknown Periconia species. These isolates were thoroughly studied through an integrative analysis based on a multi-locus phylogeny of a combined dataset of ITS rDNA, LSU, SSU, TEF1α, and RPB2, along with morphological characteristics. Based on the data obtained from this study, we propose a new addition to this genus, Periconia callaina sp. nov. This discovery provides further insight into the fungal diversity present in natural history museums and the communities that colonize and threaten our cultural heritage.

Microorganisms doi: 10.3390/microorganisms14091893

Authors: Jia-You Xing Zi-Xuan Fu Wen-Jie Xu Jing-Yang Li Zi-Ji Wang Yu-Xin Xiang Jiang Wang Sheng-Li Ming Yue-Ting Zheng Jian-Li Li Lei Zeng

Feline bocavirus (FBoV) is an emerging enteric virus associated with gastrointestinal diseases in cats and has attracted increasing attention in feline health. However, no commercial vaccine is currently available for the prevention and control of FBoV infection. VP2 is the major capsid protein of FBoV and represents a promising target for vaccine development. In this study, the FBoV VP2 protein was expressed using the insect baculovirus expression system. The purified VP2 protein self-assembled into virus-like particles (VLPs), which were formulated with Alum, ISA 206, or GEL 02 adjuvants to prepare VP2 VLP vaccines. The immunogenicity, cellular immune responses, antigen uptake, biodistribution, germinal center responses, and safety of the vaccines were evaluated in BALB/c mice. The results showed that all VP2 VLP vaccine formulations induced VP2-specific IgG antibodies and neutralizing antibodies, promoted B- and T-lymphocyte activation, enhanced dendritic cell maturation, and stimulated germinal center-related immune responses. Among the tested formulations, VP2+GEL 02 induced the strongest immune responses and showed favorable safety in mice. These findings demonstrate that FBoV VP2 exhibits favorable immunogenicity and represents a promising vaccine antigen for further development against feline bocavirus.

Microorganisms doi: 10.3390/microorganisms14091892

Authors: Jia Su Xugang Lin Yina Xia Jing Zhao Yue Chen Depei Wang Xianli Xue

Filamentous fungi are ubiquitous eukaryotic microorganisms characterized by multicellular architectures and branched hyphal structures. They are used in industrial production for the manufacture of enzyme preparations, various organic acids, and secondary metabolites. Transcription factors serve as core regulatory molecules governing vital biological processes in filamentous fungi. This review systematically elaborates on the classification, structural features, and functional diversity of transcription factors in filamentous fungi, with a focus on their regulatory roles in hyphal growth, morphological differentiation, metabolite synthesis, and environmental stress responses. We further summarize the key transcription factors in three representative fungal genera, namely, Aspergillus, Trichoderma, and Penicillium. Additionally, the potential applications of genetically engineering transcription factors to increase the yield of metabolites are discussed. This review provides a comprehensive theoretical framework for the genetic modification and high-value utilization of filamentous fungi.

Microorganisms doi: 10.3390/microorganisms14091891

Authors: Lei Wang Junzhe Shi Liwen Li Kaipeng Jiang Jingwei Lian Dezong Sui Sian Liu Yingdan Yuan Yingzhou Tang

Common reed (Phragmites australis) is a native dominant plant in many coastal wetlands. To determine how rhizosphere effects and soil depth shape microbial communities, we sampled the rhizosphere and three bulk-soil layers (0–15, 15–30, and 30–45 cm) in a monodominant common-reed stand in a coastal salt marsh. Soil physicochemical properties and bacterial and fungal α-diversity, community composition, and assembly processes were evaluated using one-way ANOVA, principal coordinates analysis (PCoA), permutational multivariate analysis of variance (PERMANOVA), neutral community models, and phylogenetic null models. Rhizosphere pH was lower than that of 0–15 cm bulk soil (mean 8.434 vs. 8.712) but remained alkaline; soil organic matter, total nitrogen, hydrolyzable nitrogen, and total phosphorus were greatest in the rhizosphere. Neither bacterial nor fungal richness or Shannon diversity differed significantly among compartments (p > 0.05); fungal Shannon means ranged from 2.083 to 3.077, with relatively higher Bacteroidota and lower Acidobacteriota abundance in the rhizosphere. Fungal composition did not differ significantly (pseudo-F = 0.609, R2 = 0.102, p = 0.9112), although Mucoromycota and Rozellomycota were relatively more abundant in the rhizosphere. Phylogenetic null models indicated predominantly deterministic bacterial assembly, with 10% dispersal limitation in the 15–30 cm layer. Fungal assembly was predominantly stochastic in bulk soils, whereas the rhizosphere was an exception: heterogeneous selection accounted for 60% of pairwise comparisons and median βNTI exceeded +2. The Mantel test identified only the association between total phosphorus and bacterial diversity as significant (0.01 < p < 0.05). These results show that rhizosphere filtering strongly structured bacterial composition and imposed deterministic selection on rhizosphere fungi.

Microorganisms doi: 10.3390/microorganisms14091890

Authors: Zhihua Feng Cunyi Qiu Zhiding Zhou Meilin Yang Huaxin Wang Yefei Zhou

Autophagy is a highly conserved degradation and recycling process in eukaryotic cells that plays a critical role in maintaining cellular homeostasis and responding to external stress. During viral infection, autophagy exhibits a classic “double-edged sword” effect—it can act as a host defense mechanism by directly degrading viral components, but it can also be hijacked by viruses to promote their own replication. Porcine epidemic diarrhea virus (PEDV), an important enteric coronavirus that severely affects the global swine industry, engages in a complex and sophisticated interplay with the host autophagy system. This review systematically dissects the dual regulatory mechanisms of autophagy during PEDV infection and reveals two intertwined functional axes. On one hand, PEDV utilizes multiple viral proteins to cooperatively manipulate the autophagic pathway—inducing mitophagy to suppress innate immune responses, utilizing autophagic membranes to construct replication platforms, and blocking autophagic flux to evade degradation—thereby establishing a multi-level pro-viral network. On the other hand, host cells deploy a unified molecular axis of “ubiquitination–autophagy receptor–lysosome” by mobilizing a broad array of restriction factors to target and degrade viral proteins, forming a coordinated defense system. These two axes converge at the oxidative stress–endoplasmic reticulum stress–autophagy hub, where PEDV NSP1 and NSP2 synergistically inhibit the NRF2 antioxidant system to trigger this cascade, while host factors such as DDX6 and ACE2 finely regulate the process. Based on this mechanistic framework, we discuss the therapeutic implications of targeting autophagy for PEDV intervention, with particular emphasis on the development of selective autophagy modulators as potential antiviral agents. We also identify key knowledge gaps and propose future research directions to translate these mechanistic insights into clinical or field applications. This review synthesizes the peer-reviewed literature published between 2013 and 2026, identified through systematic searches of PubMed, Web of Science, and Scopus databases. Notably, the majority of mechanistic findings discussed are derived from in vitro cell culture models, and their translation to in vivo settings remains a significant challenge. Bridging this gap will require validation in physiologically relevant models, such as porcine intestinal organoids and controlled piglet challenge studies, to assess the efficacy and safety of autophagy-targeting interventions in the context of intestinal homeostasis and mucosal immunity.

Microorganisms doi: 10.3390/microorganisms14091889

Authors: Mengyao Wang Yuhong Zhou Mengxiao Li Shanhua Qin Ziyue Qi Meifang Li Nan Yang Yi Zhou Xiaoya Wei Yujie Zhang Zhonghui Yang Zhihui Cheng

Ehrlichia chaffeensis is an obligate intracellular bacterium that proliferates within monocytes or macrophages and causes human monocytic ehrlichiosis (HME), an emerging life-threatening zoonosis. Doxycycline is the choice of treatment for HME, yet it has prominent side effects. Host cells lack the lysine biosynthetic pathway; thus, the enzymes in this pathway are essential for bacterial growth and recognized as potential targets for the development of novel antibiotics. Here, we demonstrated that inhibitors targeting DapE, which is a key enzyme in the lysine biosynthetic pathway, especially disulfiram, effectively inhibit E. chaffeensis infection and intracellular growth. Through complementation experiments and peptide nucleic acid-mediated dapE knockdown, we showed that DapE in E. chaffeensis is functional and essential for bacterial intracellular growth. Using purified recombinant protein, we found that DapE induces IL-8 expression in host cells. Finally, we identified that NtrX, the response regulator of the two-component system NtrY/NtrX, regulates dapE expression using an electrophoretic mobility shift assay and a reporter assay. Our findings deepen the understanding of E. chaffeensis pathogenesis as well as illustrate that DapE in E. chaffeensis is a potential therapeutic target for the development of novel HME treatments.

Microorganisms doi: 10.3390/microorganisms14091888

Authors: Po-Hsiu Huang Chien-Hao Tseng Chia-Wei Liu Wei-Hsuan Huang Ting-Kuang Yeh Po-Yu Liu Hsien-Po Huang

Catheter-related bloodstream infections (CRBSIs) caused by Brevundimonas diminuta are rare, and polymicrobial infections involving this organism are exceptionally uncommon. To the best of our knowledge, we describe the first reported case of polymicrobial CRBSI caused by B. diminuta and Streptococcus anginosus in a woman with a pituitary gland tumor and intracranial hypertension who was receiving intravenous treatment through an indwelling peripherally inserted central catheter (PICC). The patient made a full clinical recovery after prompt PICC removal and intravenous piperacillin–tazobactam therapy. This case emphasizes the emerging clinical significance of B. diminuta and S. anginosus as potential causes of polymicrobial bloodstream infection in patients with indwelling central venous catheters and underlines the need to consider uncommon environmental Gram-negative organisms in catheter-related infection.

Microorganisms doi: 10.3390/microorganisms14091887

Authors: Ling Liu Kang Zhao Chang-Run Zhao Tao-Ni Zhang Yi Li Qin Wu Qi Wang Chuan-Rui Yang Wen-Qing Zhao Qiu-Ying Chen Tianchao Wei Teng Huang Jianni Huang Meilan Mo

Avian infectious bronchitis virus (IBV) is widely distributed worldwide and causes substantial economic losses to the poultry industry. Because IBV undergoes frequent mutation, prevention and control of infection remain challenging. Immunization is an important measure for the prevention and control of IB. Therefore, there is an urgent need for a rapid, sensitive, specific, and convenient method for the detection of antibodies against IBV. In this study, we firstly developed an indirect colloidal gold immunochromatographic strip for the rapid detection of antibodies against IBV based on a conserved epitope peptide. The recombinant epitope peptide recognized by N2D5 monoclonal antibody (mAb) against the N protein of IBV was expressed as a GST fusion protein (GST-N2D5) based on the conserved antigenic epitope previously identified in our laboratory. Colloidal gold-labeled GST-N2D5 was used as the detection reagent to generate visual signals. Rabbit anti-chicken IgY and mouse anti-GST mAb were immobilized on the nitrocellulose membrane as the test line (T line) and control line (C line), respectively. The optimal pH and optimal protein concentration for conjugation of gold nanoparticles (AuNPs) with GST-N2D5 were pH 8.5 and 72 µg/mL, respectively. Specificity was evaluated using common avian pathogens, and no cross-reactivity was observed. The detection limit of the strip for IBV-positive serum was 1:180. In addition, the assay showed good reproducibility and stability, and results could be observed within 5 min without any specialized equipment. Clinical chicken serum samples were tested using both the developed strip and an enzyme-linked immunosorbent assay (ELISA), and the strip showed high agreement with the ELISA. In conclusion, the established immunochromatographic strip is rapid, sensitive, specific, and easy to operate, and therefore has potential as an on-site tool for the rapid detection of antibodies against IBV, particularly in resource-limited settings.

Microorganisms doi: 10.3390/microorganisms14091886

Authors: Teresa Maria Assunta Fasciana

Legionella pneumophila epitomizes the concept of a microorganism with “a thousand faces”, reflecting its extraordinary ecological adaptability, genetic diversity, and multifaceted clinical relevance [...]

Microorganisms doi: 10.3390/microorganisms14091885

Authors: Zhihao Chen Yiqiong Zhang Zhouchang Zhang Tianjiao Song Yufei Ji Xinrui Li Liangxuan Xu Hua Yi Yao Wang Yanbing Lin

Forests store carbon in downed logs, whose decomposition is driven by bacterial and fungal communities. Yet most studies treat individual logs as homogeneous units, obscuring internal–external gradients in wood chemistry as fine-scale habitat filters. To address this gap, we combined ALDEx2, SpiecEasi networks, Mantel tests and PLS-SEM on 24 samples from Larix principis-rupprechtii logs spanning three decay stages (I, III, and V) and two radial positions. Chemical heterogeneity between positions peaked at Stage III (5.9-fold difference in total nitrogen), with severe internal nitrogen and phosphorus depletion. Bacterial alpha diversity converged between positions by Stage V, whereas fungal communities maintained persistent divergence, and differentially abundant taxa showed stage- and position-associated enrichment patterns. Network topology did not differ detectably between internal and external wood. Path modelling showed contrasting associations: fungal composition tracked primarily the decay-stage axis, whereas bacterial composition was associated more weakly and mainly through an indirect nutrient channel. These results provide a fine-scale, descriptive account of divergent bacterial and fungal responses to within-log heterogeneity during downed-log decomposition in a temperate coniferous forest.

Microorganisms doi: 10.3390/microorganisms14091884

Authors: Takashi Onodera Sungwook Seo Akikazu Sakudo Antonio Toniolo

Cats represent a susceptible host and a possible translational model for investigating coronavirus pathogenesis and therapeutics. Recent immunohistochemical (IHC) and histopathological studies in both human and feline tissues have demonstrated SARS-CoV-2 nucleocapsid protein (NP) and spike protein expression within pancreatic islet cells, following a classic temporal infection course. Notably, IHC analysis also reveals NP expression within exocrine ductal epithelial cells. Given that ductal epithelium functions as an islet progenitor pool during tissue injury or metabolic stress, pancreotropic coronaviruses may gain access to the endocrine compartment by exploiting this intrinsic cellular differentiation pathway. Although the precise mechanisms governing intra-islet viral entry remain to be elucidated, this review highlights the capacity of SARS-CoV-2 to compromise both the exocrine (digestive) and endocrine functions of the pancreas. Finally, we evaluate the therapeutic potential of direct-acting antivirals—specifically RNA-dependent RNA polymerase (RdRp) and protease inhibitors—as monotherapies and in synergistic combination to limit pancreatic injury and mitigate the diabetogenic effects of coronaviruses across both acute infection and post-acute sequelae, such as human long-COVID syndrome.

Microorganisms doi: 10.3390/microorganisms14091883

Authors: Faik Sertel Secer

Lactococcosis, caused by Lactococcus garvieae, is a persistent problem of farmed rainbow trout, and the reduced efficacy of conventional antibiotics has prompted interest in plant-derived alternatives. The in vitro antibacterial activity of five commercial essential oils from Melaleuca alternifolia, Mentha piperita, Citrus limon, Rosmarinus officinalis and Lavandula hybrida was evaluated against six L. garvieae isolates from rainbow trout. Minimum inhibitory and minimum bactericidal concentrations (MIC and MBC) were determined by broth microdilution, and the mode of action was defined by the MBC/MIC ratio; susceptibility to oxytetracycline (OTC) and trimethoprim–sulfamethoxazole (SXT) was also assessed. The major constituents were terpinen-4-ol (tea tree), menthol (peppermint), limonene (lemon), 1,8-cineole (rosemary) and linalool (lavandin). Activities were strain-dependent, with MIC values mostly from 6.25 to 100% (v/v) (single lowest, 1.5625% for rosemary), and lemon was among the least active, but the Kruskal–Wallis test showed no significant differences among oils (p > 0.05), so these differences are descriptive only. Where effective, the oils acted mainly through a bactericidal mechanism (MBC/MIC ≤ 4), but only at high concentrations, so their practical significance appears limited. All six isolates were resistant to OTC (MIC 8–16 µg/mL) and SXT (MIC 8–32 µg/mL). In vivo studies are required before practical use.

Microorganisms doi: 10.3390/microorganisms14091882

Authors: Karolina Kraus Paweł Mikziński Bindu Subhadra Emil Paluch

Periprosthetic joint infection (PJI) remains one of the most serious complications of arthroplasty, largely due to the formation of microbial biofilms on implant surfaces. Biofilm-associated infections exhibit increased tolerance to antimicrobial therapy and host immune responses, making eradication difficult and often requiring repeated surgical interventions. Consequently, there is a growing need for effective local therapeutic strategies capable of delivering high concentrations of antimicrobial agents directly to the site of infection while minimizing systemic toxicity. Hydrogels have emerged as promising drug delivery platforms for the management of biofilm-associated PJI. Their biocompatibility, injectability, high water content, and tunable physicochemical properties enable controlled and localized release of therapeutic agents within the infected peri-implant environment. This narrative review summarizes recent advances in hydrogel-based approaches, including antibiotic-loaded hydrogels, systems incorporating anti-biofilm enzymes, bacteriophage-loaded formulations, and nanoparticle-enhanced platforms. It also highlights future research directions, with particular emphasis on the need for expanded clinical studies to facilitate the translation of emerging hydrogel-based therapies into clinical practice. Further development of these systems should focus on the incorporation of novel therapeutic agents into hydrogel platforms, aiming to enhance biofilm eradication and improve treatment outcomes in patients with PJI. Particular attention is given to stimuli-responsive (“smart”) hydrogels that release therapeutic payloads in response to infection-related triggers such as pH changes, with emphasis on the need for expanded clinical studies to facilitate the translation of emerging hydrogel-based therapies into clinical practice. Further development of these systems should focus on the incorporation of novel therapeutic agents into hydrogel platforms, aiming to enhance biofilm eradication and improve treatment outcomes in patients with PJI.

Microorganisms doi: 10.3390/microorganisms14091881

Authors: Ping Fu Zizhen Tang Wenjie Chai Ling Ke Bingting Wu Zhan Gao Yang Huang Dan Yuan Qiulei Zhong Yan Yu Zhenxin Fan Miao He

The accurate characterization of human immunodeficiency virus (HIV) genetic diversity and drug resistance is critical for effective surveillance and treatment, yet current sequencing technologies face limitations in sensitivity and scalability for community-level implementation. We present G-HIV, an integrated platform combining long-read sequencing (G-seq500) with an automated bioinformatics pipeline. G-HIV processes raw FastQ data to generate automated reports on point mutations, drug resistance predictions, viral quasispecies diversity, and haplotype networks via a two-step analytical approach. Applied to 44 HIV-1 plasma samples (42 used in the final comparison after excluding 2 samples with low-quality Sanger chromatograms), G-HIV detected 3–48 candidate minority variants per sample that were not observed by Sanger sequencing, identifying drug-resistant quasispecies in two samples with undetectable Sanger signals, and revealed mixed infection cases (e.g., inter-subtype CRF07_BC/CRF08_BC) through phylogenetic analysis. G-HIV addresses an integration of long-read sequencing with a fully automated, one-stop bioinformatics pipeline designed for frontline laboratories without specialized bioinformatics expertise—providing a scalable solution for community-based resistance surveillance and personalized therapy optimization in resource-limited settings. This research addresses an integrated long-read sequencing and automated bioinformatics platform for rapid and precise HIV-1 surveillance. G-HIV surpasses conventional approaches like Sanger sequencing in resolution, efficiency, and accessibility for community-level surveillance. By integrating long-read sequencing, streamlining workflows and eliminating the need for specialized bioinformatics expertise, G-HIV is positioned to become a new solution, providing more effective one-stop services for HIV-1 prevention and control.

Microorganisms doi: 10.3390/microorganisms14091880

Authors: Károly Szili Csilla Dézsi Viktor Gulyás-Oldal Dániel Sallai Gábor Patay Ekaterine Paschali Sándor Nagy

Chronic non-communicable conditions—type 1 and type 2 diabetes mellitus (T1DM, T2DM), metabolic obesity syndrome (MOS), polycystic ovary syndrome (PCOS), colorectal and extra-intestinal cancers, and systemic autoimmune disease—share a prolonged asymptomatic phase during which conventional screening is invasive, insensitive, or resource-intensive. This review synthesizes the 2021–2026 literature on fecal microbiome-based artificial intelligence (AI) diagnostics across these conditions, extracting reported discrimination, validation strategy, microbial and short-chain fatty acid (SCFA) biomarkers, and cross-cohort reproducibility. Across the primary classifier studies tabulated here, reported areas under the curve (AUCs) span 0.76–0.99 under internal validation but 0.69–0.91 under external or cross-population validation; in the four studies reporting both, the median AUC falls from 0.875 to 0.810. Verified external-validation values include 0.82 for colorectal cancer, 0.79 for T2DM and 0.792 for discrimination of systemic lupus erythematosus from rheumatoid arthritis and controls. Clinical readiness turns on this internal-to-external gap more than on the headline AUC. We propose a multimodal deep learning architecture coupled with explainable AI; no component has been implemented or evaluated on data, and it is presented as a design proposal. Fecal-microbiome-based multimodal AI is technically feasible but clinically unvalidated, pending prospective, harmonized cross-cohort trials.

Microorganisms doi: 10.3390/microorganisms14091879

Authors: Xin Jiang Xianfei Huang Xianliang Wu

Microplastics (MPs) pollution caused by agricultural film residues, organic fertilizer application, sewage irrigation, and atmospheric deposition has gradually become an unignorable interference factor to the sustainable development of the rhizosphere soil and crop in farmland. However, their specific impacts on the rhizosphere and crops remain unclear. Therefore, this review focuses on the current knowledge on the response mechanisms of rhizosphere soil and crops to MP contamination. The density of MPs is generally lower than that of soil mineral particles. Their substantial accumulation in soil can significantly reduce both the bulk density (by increasing total porosity) and the particle density (by diluting the heavy solid phase with light plastic components). The introduction of MPs disrupts the normal metabolism of soil bacterial communities; a disruption directly reflected in functional genes associated with carbon cycling. MPs can interfere with the activity of key metabolic enzymes involved in fungal nutrient cycling, thereby disrupting normal energy allocation and material metabolism. Viruses can regulate the turnover and metabolism of microbial communities through lytic and lysogenic cycles, consequently influencing the carbon fate of MPs. The toxicity and underlying mechanisms of MPs on soil fauna primarily manifest in aspects such as feeding behavior, growth and development, oxidative stress, intestinal toxicity, and reproductive toxicity. The direct effects of MPs on plants include physical barriers and mechanical damage, induction of oxidative stress, interference with nutrient uptake, disruption of photosynthesis and carbon metabolism, and disruption of plant hormone networks. This review identifies critical knowledge gaps, particularly regarding crop quality, field-based soil faunal studies, virus-microbe interactions, and degradation products, and proposes future research directions to better understand the risks MPs pose to agricultural sustainability and food safety.

Microorganisms doi: 10.3390/microorganisms14091878

Authors: Meng Li Yinyu Gu Chuanjie Chen Zongshuai Wang Xiaohong Guo Xiaoyan Liang Kuihua Yi Junlin Li Dongyang Li Haiyang Zhang

The application of biochar and bioorganic fertilizer (BOF) in agricultural systems has garnered increasing attention in recent years. Nevertheless, research remains scarce on the impacts of biochar and BOF on the rhizosphere microecological characteristics of saline-alkali soils. This research involved the execution and analysis of 16S rRNA sequencing using Illumina technology to explore how biochar, whether used alone or in conjunction with BOF, along with varying application rates, impacts the microbial community in the saline-alkali rhizosphere soil during quinoa cultivation. In the conducted field trial, sole BOF application, sole biochar application, and their combined application (referred to as BOFB) led to a substantial enhancement of 23.88%, 74.08–97.00%, and 188.88–220.59% in quinoa aerial biomass, respectively. Meanwhile, sole biochar application or biochar combined with BOF reduced soil electrical conductivity (EC) by 26.42–39.81%. Biochar and BOF significantly improved most soil parameters, with the exception of total phosphorus (TP). In comparison to the control (CK), the relative abundances of Pseudomonas, Arthrobacter, Skermanella, and Bacillus were elevated in the biochar and BOFB treatments, while Sphingomonas was more abundant in the BOF treatment. In addition, Skermanella exhibited a significant positive correlation with EC and available potassium (AK). Biochar exerted a stronger effect on soil bacterial community structure than BOF. Furthermore, the complexity of the bacterial community in biochar and BOFB treatments far exceeded that in the BOF and CK treatments. Overall, the application of biochar effectively reduced soil EC and improved soil fertility, enhanced bacterial community stability, and optimized bacterial community structure, thereby increasing quinoa aerial biomass. Under the conditions of this study, the optimal application rate for biochar was 15 t/ha, and the combined application of biochar and BOF produced superior effects relative to either amendment alone.

Microorganisms doi: 10.3390/microorganisms14091877

Authors: Luna Gao Lin Wei Siqi Wang Jun Li Jingli Liu Zhao Guo Zhi-Min Li Zhimin Li

β-Nicotinamide mononucleotide (NMN) is an important intermediate in nicotinamide adenine dinucleotide (NAD+) metabolism and has attracted increasing interest as a bioactive compound and biomanufacturing product. In this study, Pseudomonas putida KT2440 was engineered for NMN production using a “block–enhance–transport” strategy. Deletion of nicB impaired nicotinic acid degradation and resulted in the accumulation of 66.2 μM nicotinic acid in cell extracts, whereas additional deletion of the putative NMN-consuming genes pncC and ushA did not lead to detectable NMN accumulation. Coexpression of endogenous pncB and engineered Francisella tularensis nadE* enabled low-level NMN formation through a Preiss–Handler pathway-based route. By contrast, overexpression of endogenous nadA, nadB, and nadC strengthened precursor supply through the NAD+ de novo biosynthetic pathway and resulted in approximately 0.17 mM NMN in cell extracts. Chromosomal integration of an engineered Salmonella enterica pnuC* transporter cassette was associated with pronounced extracellular NMN accumulation. Additional overexpression of genes involved in downstream NAD+ metabolism or phosphoribosyl pyrophosphate supply did not considerably improve production, possibly because of metabolic competition or expression burden. The best-performing strain, LW10, produced 1.28 mM extracellular NMN, corresponding to approximately 0.43 g/L, after 96 h of shake-flask cultivation in basal salt medium containing glucose and L-aspartate. These results suggest the feasibility of NMN biosynthesis in engineered P. putida KT2440 and highlight the importance of balancing precursor supply, competing reactions, and product transport. Thus, P. putida KT2440 represents an alternative chassis for further pathway balancing and process optimization toward fermentative NMN production.

Microorganisms doi: 10.3390/microorganisms14091876

Authors: Pedro Antonio Pérez-Ferrer Michele Kiyoko Nishiguchi

Bacterial communication or quorum sensing (QS) is a common yet complex system where multiple factors influence the extent to how this chemical dialog is transferred from a single clone to the larger community of microbes in the population. More often, when bacteria are in large concentrations, their genetic and subsequent biochemical response to different chemical cues is influenced by not only which microorganisms are present but also the environmental variables that surround those individuals. This is especially relevant when symbiotic bacteria are dependent upon host functions yet are in high enough concentrations that can manifest their own behaviors in response to the present host prior, during, and after colonization. This review will examine the various abiotic and biotic factors that regulate QS when bacteria are in the process of detecting, colonizing, and persisting in a host that uptakes its microbial partner from the environment, as well as the consequences of multiple stressors on this dynamic communication process.

Microorganisms doi: 10.3390/microorganisms14091875

Authors: Francesco Giuseppe De Rosa Tommaso Lupia Valentina Fornari Davide Vita Marco Casarotto Silvia Corcione Alessandra Oliva

Antimicrobial resistance (AMR) in Gram-negative pathogens represents a critical global health challenge. Combining cefepime with novel β-lactamase inhibitors (enmetazobactam, taniborbactam, zidebactam, and nacubactam) revitalizes the role of this fourth-generation cephalosporin in contemporary therapy. We present a narrative review according to the Scale for the Assessment of Narrative Review Articles (SANRA) criteria, summarizing the existing literature regarding new cefepime combinations with β-lactamase inhibitors. Based on 68 identified studies, data were structured by molecule. Each section explores in vitro activity, preclinical in vivo data, PK/PD parameters, clinical evidence, and dosage. Findings demonstrate that these inhibitors successfully restore cefepime’s efficacy against diverse resistance mechanisms, notably ESBLs, AmpC, KPC, OXA-48, and metallo-β-lactamases. “Cefepime 2.0” therapies expand the therapeutic armamentarium against severe multidrug-resistant infections, offering vital carbapenem-sparing strategies. Because they possess distinct microbiological spectra, these agents serve complementary rather than interchangeable roles. Their clinical success demands targeted integration into antimicrobial stewardship programs to optimize efficacy and safely reduce carbapenem dependence.

Microorganisms doi: 10.3390/microorganisms14091874

Authors: Zenan Zhang Xing Guo Kui Guo Wei Guo Cheng Du Yan Yang Wenjing Dong Xiaoyu Chu Yuezhi Lin Xiaojun Wang

Rotavirus A (RVA) causes severe diarrhea in donkey foals, but rapid quantitative antigen assays for vaccine development and manufacturing control are lacking. We developed a VP6-based antigen-capture ELISA (acELISA) for donkey-origin RVA using rabbit anti-RVA polyclonal IgG as the capture antibody and horseradish peroxidase-conjugated mouse anti-VP6 monoclonal antibody 3F6 as the detector antibody. Assay conditions were optimized, and performance was evaluated through standard-curve analysis, specificity and repeatability testing, Western blot (WB) comparison, median tissue culture infectious dose (TCID50), reverse transcription quantitative polymerase chain reaction (RT-qPCR), ultrafiltration membrane screening, and pilot-scale tangential flow filtration (TFF) monitoring. The optimized acELISA detected VP6 at 1 ng/mL and showed a linear range of 1–25 ng/mL, with no cross-reactivity with eight common equine pathogens, and intra- and inter-assay coefficients of variation below 10%. The acELISA estimates were consistent with Western blot densitometry. In process applications, the assay supported the selection of a 100 kDa membrane and monitored pilot-scale TFF, yielding 75.69% VP6 recovery and an 18-fold increase in antigen purity. This acELISA provides a specific and practical in-process tool for quantitative antigen monitoring in donkey rotavirus vaccine production and is intended to complement, rather than replace, infectivity- and genome-based assays.

Microorganisms doi: 10.3390/microorganisms14091873

Authors: Sok Song Kyu-Sik Shin So-Hee Park Yong Yi Joo Cho-Yeon Lee Hyun-Ok Ku Wooseog Jeong

Chemical disinfectants used in livestock production commonly combine citric acid (CA) and benzalkonium chloride (BZK), yet the respective contributions of these active ingredients to virucidal efficacy remain poorly understood. This study quantified the relative contributions of CA and BZK against the non-enveloped foot-and-mouth disease virus (FMDV) and the enveloped avian influenza virus (AIV). Virucidal efficacy was evaluated using a full-factorial design comprising six CA concentrations and six BZK concentrations at contact times of 3 and 30 min. Log reduction values (LRVs) were analyzed by two-way ANOVA and multiple linear regression, and the relative importance of each predictor was estimated using the Lindeman–Merenda–Gold (LMG) method. FMDV inactivation was primarily determined by CA, which accounted for 99.99% and 99.87% of the explained variance at 3 and 30 min, respectively, whereas BZK and the interaction term contributed minimally. In contrast, BZK was the dominant determinant of AIV inactivation, explaining 69.79% and 78.02% of the variance at 3 and 30 min, respectively, while CA made a smaller contribution. These findings demonstrate that the dominant active ingredient in CA–BZK combination disinfectants differed markedly between the FMDV and AIV models examined. Quantifying the relative contribution of individual components provides a basis for the rational formulation and optimization of veterinary disinfectants for different target viruses.

Microorganisms doi: 10.3390/microorganisms14091872

Authors: Chengwei Shao Jianguang Fu Fei Deng Huiyan Yu Huan Fan Yanjun Chen Ke Xu Mingwei Wei Siyue Jia Xiaoyan Jia Liguo Zhu Jingxin Li

Population immune pressure from vaccination and prior infection continues to drive the evolution of SARS-CoV-2. Systematic characterization of RBD mutations under complex immune backgrounds is essential for understanding viral adaptation and evolutionary trajectories. Here, we applied a deep mutational scanning (DMS) to comprehensively map the neutralization escape landscape of the Omicron variant JN.1 and its descendant lineage XEC, under immune pressure from individuals who experienced Omicron breakthrough infections following three doses of inactivated vaccines. A neutralization escape map for the single amino acid substitutions in the RBD of JN.1 or XEC was generated, and the escape efficiency of each mutation was determined. The results show that RBD escape mutations are hierarchically organized: low-intensity signals are widespread, whereas high-intensity escape is confined to a few key sites. These escape mutations are not confined solely to the receptor-binding motif (RBM) but are broadly distributed across the entire RBD. Many escape sites could accommodate multiple amino acid substitutions. Integration of DMS data with genomic surveillance of circulating variants from 2024 to 2025 revealed significant overlap between experimentally identified escape sites and mutations observed in natural isolates. This overlap increased substantially in 2025, with site concordance rising from 27.17% and 26.81% to 45.09% and 47.10% for JN.1 and XEC, respectively. The natural prevalence of these escape mutations is further shaped by factors such as receptor-binding affinity, protein stability, and epistatic interactions. Overall, our findings suggest that SARS-CoV-2 antigenic evolution follows the pattern of multiple pathways within a constrained space, providing new insights into the adaptive mechanisms of Omicron-derived variants under hybrid immune pressure.

Microorganisms doi: 10.3390/microorganisms14091871

Authors: Qiuyu Wang Weihao Chen Yuchi Zhao Jiajing Liu Jingyan Xu Chunshi Wang Qi Sun Weiwei Dong Wenxiu Ji

Panax ginseng, a high-value medicinal plant, faces substantial yield losses due to black spot disease, while conventional chemical controls cause pesticide residues and soil ecological damage, necessitating green biocontrol strategies. Here, two antagonistic strains, Bacillus sp. WY17 and WY26, were isolated from the surface-sterilized internal root tissues of 10-year-old ginseng. Through systematic optimization of carbon/nitrogen sources, inorganic salts, and fermentation parameters (temperature, pH, agitation, inoculum size, and duration), the optimal culture conditions were established. The optimal consortium consisted of WY17 and WY26 in a 2:1 ratio (WY17:WY26 = 2:1), which achieved an antifungal inhibition rate of 84.94% against the pathogen compared to the untreated control group (pathogen only). Mechanistic investigations revealed that the crude protein extract exerted its antifungal effect by compromising the integrity of the pathogen’s cell membrane, leading to increased permeability and leakage of intra-cellular contents, and produced cell wall-degrading enzymes (chitinase and β-1,3-glucanase), thereby inhibiting mycelial growth and spore germination. In vitro efficacy tests demonstrated that this crude protein extract performed comparably to the chemical fungicide 70% mancozeb, with no statistically significant difference observed between them (p > 0.05). These findings identify a promising compound biocontrol agent derived from indigenous Bacillus strains, offering an effective and environmentally friendly alternative for managing ginseng black spot disease.

Microorganisms doi: 10.3390/microorganisms14091870

Authors: Oana-Elena Ioniţă Roxana-Carmen Cernat Nicola-Maria Militaru Maria-Elena Vodarici Maria Fulina Daniela Pițigoi Elena Mocanu Beatrice Severin Claudia-Simona Cambrea Irina-Magdalena Dumitru

Healthcare-associated infections (HAIs) caused by multidrug-resistant Klebsiella spp. represent a critical and escalating global public health threat. Carbapenem-resistant Klebsiella pneumoniae (CRKP) has been designated a critical-priority pathogen by the World Health Organization, and in the 2024 WHO Bacterial Priority Pathogens List, it was the top-ranked pathogen overall. The convergence of carbapenem resistance with hypervirulence in emerging strains has further complicated therapeutic decision-making. This review provides a narrative synthesis of the evidence published between 2020 and 2025 on the prevalence, resistance mechanisms, molecular epidemiology, clinical outcomes, and therapeutic strategies for Klebsiella pneumoniae infections acquired in healthcare settings, with particular attention to the Eastern European and Romanian context. PubMed/MEDLINE, Embase, Web of Science, and the Cochrane Library were searched for relevant publications from January 2020 to June 2025, supplemented by WHO and ECDC surveillance reports. Studies were selected narratively for their relevance to the themes addressed. No new quantitative pooling was undertaken; all summary estimates reported below are cited from the published meta-analyses and surveillance reports that generated them. In the most recent global meta-analysis of hospital-acquired CRKP infection, which pooled 61 studies and 513,307 patients from 14 countries, the global prevalence of CRKP among nosocomial K. pneumoniae infections was 28.69% (95% CI: 26.53–30.86%), with pronounced regional variation from 14.29% in high-income North America to 66.04% in South Asia, and 42.05% in Western Europe. Pooled mortality among patients infected with CRKP has been estimated in a separate meta-analysis at 42.14%, compared with 21.16% among patients infected with carbapenem-susceptible strains, rising to 54.30% in bloodstream infections. Surveillance data place Romania third in Europe for carbapenem resistance among invasive K. pneumoniae isolates, at 50.30%, with a distinctive predominance of NDM plus OXA-48-like co-producers. Ceftazidime-avibactam is recommended for KPC- and OXA-48-producing strains, whereas metallo-beta-lactamase producers require aztreonam-containing combinations. CRKP in HAIs constitutes a global epidemiological emergency characterised by marked regional heterogeneity in carbapenemase distribution, high attributable mortality and rapidly evolving molecular profiles. Locally adapted surveillance, rapid molecular diagnostics, and stewardship programmes are required since empirical therapy cannot be standardised across regions.

Microorganisms doi: 10.3390/microorganisms14091869

Authors: Hongyu Li Jiaqi Liu Jiashan Lu Jie Wei Yuying Bao Peng Zhang

Carminic acid (CA) is a high-value natural anthraquinone pigment used in foods, cosmetics, textiles, and pharmaceuticals, but its current industrial supply depends largely on extraction from the scale insect Dactylopius coccus, creating constraints in yield, cost, sustainability, and allergen control. This review summarizes recent progress from pathway elucidation to microbial production. We first outline the structure, occurrence, applications, and biosynthetic logic of CA, emphasizing the convergence of type III polyketide assembly with insect-associated tailoring reactions, especially C-glycosylation. We then compare heterologous production strategies in Escherichia coli, Saccharomyces cerevisiae, Yarrowia lipolytica, and Aspergillus nidulans, focusing on chassis-specific advantages, bottlenecks, precursor supply, malonyl-CoA engineering, dynamic regulation, enzyme compatibility, compartmentalization, and downstream processing. Structurally related anthraquinone pigments are further discussed to extract broader design principles for pathway diversification and synthetic biology. Finally, we highlight key challenges for industrial translation, including low titers, incomplete enzyme characterization, host–pathway incompatibility, and scalable purification, and propose integrated strategies combining precursor-pathway rewiring, AI-assisted enzyme engineering, biosensor-based regulation, and process optimization to develop competitive microbial cell factories.

Microorganisms doi: 10.3390/microorganisms14091868

Authors: Dimitar Parvanov Margarita Ruseva Rumiana Ganeva Teodora Tihomirova Maria Handzhiyska Stela Chapanova Jinahn Safir Sofia Koristashevskaya Ivan Pavlov Dimitar Metodiev Blaga Rukova Georgi Stamenov Savina Hadjidekova

Endometrial senescence and the endometrial microbiota have both been implicated in the regulation of endometrial receptivity, yet their relationship remains poorly understood. The aim of this study was to investigate associations between p16-positive endometrial cells and microbiota composition during the implantation window and to determine whether these relationships differ according to implantation outcome. Endometrial senescence was assessed by p16 immunohistochemistry and digital image analysis, whereas microbial composition was characterized by 16S rRNA gene sequencing in endometrial biopsies collected from 68 women prior to undergoing transfer of a single euploid embryo, which was performed within six months of biopsy under the same hormonal preparation protocol. No significant differences in luminal epithelial or stromal p16 abundance were observed according to subsequent implantation outcome. Although senescence was not directly associated with implantation success, stromal p16 expression demonstrated multiple associations with the endometrial microbiota. Increased stromal p16-positivity was associated with lower relative abundance of Lactobacillus and higher abundance of Delftia. Notably, in exploratory subgroup analyses more pronounced associations were observed in women who achieved pregnancy, including a negative correlation between stromal p16 expression and Lactobacillus abundance (ρ = −0.48, p = 0.005) and positive correlation with Delftia abundance (ρ = 0.42, p = 0.016). Species-level analyses revealed implantation outcome-specific associations involving Lactobacillus iners and Limosilactobacillus vaginalis. In addition, stromal p16 expression was associated with differences in microbial co-occurrence networks, indicating broader effects on microbial community organization. Together, these findings suggest that stromal, but not luminal, senescence is closely linked to endometrial microbiota composition and microbial community organization during the window of implantation. The observation that the strongest senescence–microbiota associations occurred in women who subsequently achieved successful implantation supports the hypothesis that coordinated senescence–microbiota relationships may represent a previously underrecognized feature of the receptive endometrial microenvironment.

Microorganisms doi: 10.3390/microorganisms14091867

Authors: Seonhong Hwang Xuangao Wu Jang-Won Yoon In-Cheol Jeon Young-In Hwang Ki-Song Kim Sunmin Park

Exercise alters gut microbiome composition, but the temporal dynamics and diet-dependent metabolic interactions remain unclear. We investigated longitudinal changes in gut microbiota, functional pathways, and metabolite profiles during and after an exercise intervention. Twenty-four healthy adults completed a sequential three-phase protocol: an 8-week self-directed exercise intervention, an 8-week washout, and an 8-week no-exercise control period. Fecal samples were collected at T1 (baseline), T2 (post-exercise), T3 (post-washout), and T4 (post-control). Microbiota composition was assessed by 16S rRNA sequencing, functional pathways predicted using PICRUSt2, metabolites predicted using COBRA Toolbox, and fecal SCFAs and bile acids quantified by GC and HPLC. Temporal causal relationships were examined using Tigramite analysis with dietary pattern stratification, and microbiota-environment associations were assessed by redundancy analysis (RDA). Alpha diversity was significantly higher at T2 than at T4 (p < 0.05). Beta diversity differed significantly between T2 and both T3 and T4, with no difference between T1 and T4, indicating reversibility. Ruminococcus gnavus was significantly higher at T2 than at T4 (p < 0.001), with several additional taxa higher at T2 at a less stringent threshold. Propionate and butyrate were elevated at T2, while total bile acids were lower. Bacteroides thetaiotaomicron was positively associated with body weight in the total cohort and specifically under a balanced dietary pattern (BD), with no significant time-lagged associations detected under a Western-style diet (WSD). In RDA, taxa associated with body weight substantially overlapped with taxa found to increase during exercise, whereas physical performance measures showed no direct temporal association with microbiota composition in causal analysis. In conclusion, exercise-induced changes in the gut microbiome were not sustained after structured exercise ended, suggesting that continuous exercise may be required. Diet further shaped whether microbiota–host associations were detectable, underscoring dietary pattern as a factor for future microbiome-targeted exercise interventions.

Microorganisms doi: 10.3390/microorganisms14091865

Authors: Xinyi Zhuang Xiang Li Zhengle Yang Xiahong Dai

Metabolic syndrome (MetS) represents a growing global health burden characterized by obesity, insulin resistance, dyslipidemia, and hypertension. Increasing evidence suggests that gut microbiota-associated mediators may serve as signaling intermediates involved in host metabolic regulation. However, the mechanisms by which these mediators interact with host signaling pathways and influence metabolic responses remain incompletely understood. This review summarizes current advances in gut microbiota-associated mediators, focusing on short-chain fatty acids, bile acids, lipopolysaccharide, trimethylamine N-oxide, and branched-chain amino acids. We discuss their interactions with host metabolic and inflammatory pathways, including pathways implicated in FFAR2/3-mediated signaling, FXR/TGR5 signaling, TLR4/NF-κB-mediated inflammatory signaling, and mTORC1-associated nutrient-sensing. Furthermore, we propose a microbiota–mediator–host signaling network framework as an emerging conceptual model to integrate these molecular interactions and highlight the utility of multi-omics approaches in characterizing complex microbiota–host communication. Despite mechanistic advances, substantial challenges remain, including heterogeneous microbial signatures across populations, limited causal evidence, inter-individual variability in therapeutic responses, and barriers to clinical translation of microbiota-targeted interventions. A better understanding of microbiota-associated signaling networks may provide new insights into metabolic regulation and contribute to the rational development of microbiota-targeted strategies that complement established lifestyle interventions for MetS management.

Microorganisms doi: 10.3390/microorganisms14091866

Authors: Anna Dobrzeniecka Ewa Talarek Małgorzata Aniszewska Barbara Kowalik-Mikołajewska Agnieszka Ołdakowska Beata Krynicka-Czech Magdalena Marczyńska Maria Pokorska-Śpiewak

Following successful neonatal HBV vaccination and perinatal screening, pediatric hepatitis B has become uncommon in Central Europe. However, migration and complex clinical scenarios are reshaping its epidemiology. We aimed to characterize the clinical course, disease phases, and treatment eligibility of children with HBV infection. This retrospective single-center study included patients aged 0 to <18 years referred to a tertiary pediatric infectious diseases center in Warsaw, Poland, between January 2016 and June 2026. Demographic, clinical, and laboratory data were analyzed, and patients were classified according to the 2025 EASL guidelines. Sixteen children were included. New referrals tripled from 2016–2020 (n = 3) to 2021–2026 (n = 9), and 50% were refugees, predominantly from Ukraine. Vertical transmission accounted for 44% of infections, whereas 25% were iatrogenic. According to the 2025 EASL criteria, 38% (6/16) of patients were unclassifiable. Occult HBV infection was identified in 31%, while only 13% (2/16) received analogue therapy because of restrictive national treatment criteria. One adolescent developed liver fibrosis (7.6 kPa), likely related to metabolic steatohepatitis. Contemporary pediatric HBV management might require adaptation to changing epidemiology, recognition of the limitations of adult-derived disease phase classifications, and prevention of HBV reactivation during immunosuppression.

Microorganisms doi: 10.3390/microorganisms14081864

Authors: Fang-Chao Zhu Yan-Bin Yang Pei-Pei Liu Xin Liu Qun-Jian Yin Xu-Yang Chen Shuo Yu

Candidatus Hepatincolaceae is a poorly characterized family of obligate Alphaproteobacterial symbionts that are widely detected in ecdysozoans. They were previously assumed to play a nutrient-scavenging role in the gut lumen. In this study, two high-quality metagenome-assembled genomes (MAGs, 1.39 Mb and 1.48 Mb in size) were recovered from the gut of the snapping shrimp Alpheus brevicristatus via metagenomic sequencing. Phylogenetic and whole-genome similarity analyses confirm that these two MAGs represent two novel, undescribed genera within the family Ca. Hepatincolaceae. Metabolic reconstruction reveals that they not only retain the canonical nutrient-scavenging pathways conserved across all Hepatincolaceae members, but also encode previously undocumented functional modules for antioxidant defense, vitamin B1 and B2 biosynthesis, and short-chain fatty acid production. They maintain a high oxygen-affinity cytochrome bd terminal oxidase to thrive in the anoxic gut microenvironment. Consistent with their symbiotic lifestyle, their genomes exhibit typical signatures of reductive evolution, such as reduced genome size, low GC content, and gene loss in amino acid and nucleotide de novo biosynthesis pathways. This study presents the first reported high-quality genomes of marine Ca. Hepatincolaceae symbionts, which are predicted to possess multiple metabolic functions extending beyond nutritional mutualism.

Microorganisms doi: 10.3390/microorganisms14081863

Authors: Kazuya Nakamura Takeshi Nagai Hitoshi Ishiguro Keiichi Kobayashi Kazuhisa Nakagawa Masahiro Okanojo Akira Nukazuka

Airborne viruses are a key driver of infectious disease transmission, highlighting the importance of reliable detection in public health surveillance. As atmospheric viral concentrations are very low, a sampler with a high viral collection efficiency is essential. Although multiple approaches for evaluating collection efficiency have been applied using various samplers, no standardized sampler has yet been developed. We previously developed a wet-type electrostatic precipitator (WT-ESP) and successfully collected severe acute respiratory syndrome coronavirus 2 from the public environment. However, its efficiency for quantitative collection of airborne viruses remains unclear. This study aimed to clarify the collection efficiency of the WT-ESP. We evaluated collection efficiency via two different approaches: direct spray, where virus-containing aerosols were sprayed directly into the WT-ESP inlet, and indirect spray, where aerosols were dispersed into a closed space and then collected using the sampler. The direct spray tests achieved 20.1–50.2% collection efficiency, whereas the indirect spray test achieved an efficiency <12%. These findings highlight that electrostatic precipitation has an advantage of enhancing collection efficiency compared with values reported for impingers in previous studies and provide preliminary insights into the collection efficiencies under direct and indirect spray conditions, providing foundational data that bridge the gap between both methods.