Microbiology Research doi: 10.3390/microbiolres17050095

Authors: Dilbar Tursun Zulhumar Yakup Huifang Bao Faqiang Zhan Yingwu Shi Hongmei Yang Jiusheng Sun Shijie Fang Ning Wang

Soil salinization is a significant environmental factor limiting agricultural production. Developing salt–alkali-tolerant microbial resources is important for the improvement of saline–alkali land. Plant growth-promoting rhizobacteria stimulate crop growth by producing the plant growth hormone indole-3-acetic acid (IAA), but their fermentation process under salt stress still needs optimization. Single-factor experiments and response surface methodology (RSM) were used to systematically optimize the fermentation conditions of the salt–alkali-tolerant Vreelandella titanicae J113. Key influencing factors were screened using the single-factor experiment design, and optimal process parameters were determined using the Box–Behnken design. IAA production and cell biomass were used as evaluation indicators to study the interactions of carbon sources, nitrogen sources, inorganic salts, temperature, cultivation time, and inoculum size. The optimal fermentation process was obtained: starch concentration 17.5 g/L, NaCl concentration 32.5 g/L, yeast extract 5 g/L, cultivation temperature 30 °C, inoculum size 3%, and cultivation time 144 h. After optimization, IAA production reached 23.02 μg/mL, an increase of 115% compared with before optimization. Salt stress experiments showed that the strain could still maintain high IAA production under 3% NaCl, demonstrating good salt tolerance. Maize seed germination experiments demonstrated that the optimized fermentation broth significantly promoted seed germination and seedling growth under salt stress conditions, with root length, fibrous root number, and fresh weight increasing by 61–86%, 137–200%, and 25–57%, respectively, compared to the control group. This study established an efficient IAA fermentation process for the salt–alkali-tolerant Vreelandella titanicae J113, providing technical support for developing microbial plant growth regulators suitable for saline–alkali land. The optimized strain exhibits excellent growth-promoting potential under salt stress conditions, offering favorable application prospects.

Microbiology Research doi: 10.3390/microbiolres17050094

Authors: Silvana M. Tapia-Cabrera Adelfo Escalante Francisco Bolívar

Aromatic compounds derived from the shikimate (SHK) pathway constitute a diverse class of high-value molecules with applications in the pharmaceutical, food, cosmetic, and chemical industries. In microbial systems, particularly Escherichia coli, this pathway links central carbon metabolism (CCM) to the biosynthesis of L-tyrosine (L-Tyr), L-phenylalanine (L-Phe), and L-tryptophan (L-Trp), which serve as key precursors for structurally diverse metabolites. Over the past decades, metabolic engineering strategies have focused on increasing precursor availability, relieving feedback inhibition, and eliminating competing pathways. More recently, advances in synthetic biology have enabled dynamic control of metabolic flux through pathway modularization, genome-scale interventions, and regulatory circuit design. In this review, we provide a comprehensive overview of the engineering of E. coli for aromatic compound biosynthesis, highlighting key developments in the optimization of the SHK pathway and its major metabolic nodes chorismate, L-Tyr, L-Phe, and L-Trp. We examine emerging approaches, including CRISPR-based regulation, biosensor-driven dynamic control, membrane engineering, and synthetic microbial consortia. Despite significant progress, challenges related to pathway regulation, cofactor balance, metabolic burden, and product toxicity remain critical bottlenecks. Integrating metabolic engineering with synthetic biology is driving the development of programmable, scalable microbial platforms for the efficient bioproduction of aromatic compounds.

Microbiology Research doi: 10.3390/microbiolres17050093

Authors: Mandlenkosi Manika Lindiwe Modest Faye Ntandazo Dlatu Mojisola Clara Hosu

Background: The global burden of drug-resistant Mycobacterium tuberculosis continues to threaten tuberculosis control efforts, largely due to the emergence and transmission of resistance-associated genetic mutations. Molecular epidemiology provides critical insights into mutation profiles and resistance associations, yet the interplay among key mutations and their contributions to complex resistance patterns remains poorly understood, particularly in high-burden settings. Methods: A retrospective, cross-sectional, laboratory-based design was used to analyze 111 phenotypically confirmed drug-resistant isolates. Molecular drug susceptibility testing (DST) for first- and second-line anti-tuberculosis drugs was performed at the National Health Laboratory Service (NHLS) TB reference laboratory. Drug-resistance profiles were classified according to World Health Organization (WHO) definitions. Descriptive and inferential statistical analyses were conducted to determine mutation frequencies, co-occurrence patterns, and associations with resistance profiles. Results: rpoB (D435V 38.7%; S450L 36.0%) and katG (S315T 80.2%) mutations predominated, forming the core molecular basis of MDR-TB, while 15% harbored inhA promoter mutations associated with low-level isoniazid resistance. The most frequent combinations included rpoB S450L with katG S315T and rpoB D435V with katG S315T, consistent with multidrug-resistant tuberculosis (MDR-TB) profiles. Nearly 48% showed dual resistance to fluoroquinolones and second-line injectables. Conclusion: This study highlights the predominance of resistance-associated mutations and their co-occurrence patterns in shaping MDR-TB profiles in the study setting. The observed burden of second-line drug resistance underscores the importance of comprehensive resistance testing. These findings support the use of mutation profiling for rapid diagnosis and informed treatment decisions, while emphasizing the need for ongoing local surveillance to guide TB control efforts.

Microbiology Research doi: 10.3390/microbiolres17050092

Authors: Jessy T. Santana Berenice González-Rete Elia Torres-Gutiérrez Juliana Cordeiro Cardoso Cláudia Moura Melo Paz M. S. Salazar-Schettino

Trypanosoma cruzi exhibits complex genetic diversity, organized into seven distinct typing units. To complete its life cycle, the parasite must adapt to the digestive tract of various species of triatomine bugs. This systematic review aimed to understand the molecular adaptation mechanisms of T. cruzi in relation to different vector species, systematizing knowledge on vector competence. Following PRISMA guidelines, 18 experimental studies (published between 1995 and 2025) were selected from the ScienceDirect, PubMed, Scopus, and Web of Science databases, focusing on the parasite–vector interface and proteomic analyses. There was a predominance of studies conducted in Brazil (66.67%), using the Rhodnius prolixus model (72.22%) and the TcI strain (clone Dm28c). The evolution of methodological approaches reflects a transition from classical techniques, such as SDS-PAGE, to high-throughput omics strategies, including LC-MS/MS and gene editing tools such as CRISPR. The findings were organized into key biological processes, including parasite adhesion mediated by perimicrovillar membrane components, glycoinositolphospholipids (GIPLs), and mucins; the influence of the metabolic and nutritional microenvironment, particularly hemoglobin-derived peptides and glucose availability; and the role of intestinal redox conditions in triggering metacyclogenesis. Overall, the available evidence suggests that T. cruzi adaptation within triatomine vectors is a multifactorial process driven by proteomic reprogramming and post-transcriptional regulation in response to environmental signals within the vector gut. However, this understanding is largely derived from studies based on Rhodnius prolixus and TcI strains, which limits the generalization of these mechanisms across other triatomine species and parasite lineages.

Microbiology Research doi: 10.3390/microbiolres17050091

Authors: Nilton Luis Murga Valderrama Gleni T. Segura Jakson Ch Del Solar Hugo Frias Ana C. Romani Deiner J. Gongora-Bardales Ulises S. Quispe-Gutierrez Carla Maria Ordinola-Ramirez Richard C. Polveiro Dielson da S. Vieira Jorge Luis Maicelo Quintana Rainer M. Lopez Lapa

The uterine microbiome plays a critical role in maintaining pH balance, modulating the immune system, and influencing fertility, especially in artificial breeding contexts. This study examined the impact of uterine microbiota on pregnancy success in cows following embryo transfer (ET), using Illumina 16S rRNA gene sequencing of the V4 hypervariable region of samples collected from the uterine horn (UH) and the uterine body (UB) of cows during the estrous cycle preceding synchronization for ET in the Amazon region. Microbiomes from the uterine horn (UH) and the uterine body (UB) were analyzed before embryo transfer. Cows that became pregnant (UH-P and UB-P) and those that did not (UH-NP and UB-NP) were compared. Fifteen cows were grouped as follows: UB-P (three), UB-NP (five), UH-P (three), and UH-NP (four). Linear discriminant analysis effect size and heat tree analyses identified Sphingobacterium and Stenotrophomonas spp. as significantly enriched in the UB-P and UH-NP groups, respectively. Additionally, non-pregnant cows exhibited more distinctive genera than pregnant ones. These findings suggest that cows achieving pregnancy have lower microbial diversity and fewer potentially pathogenic genera. This study contributes to the emerging field of pre-pregnancy uterine microbiome research in cattle, offering evidence that microbial composition may influence reproductive success, and highlights specific taxa as potential biomarkers for pregnancy outcomes following embryo transfer.

Microbiology Research doi: 10.3390/microbiolres17050089

Authors: Egidia Costanzi Giovanna Traina Marco Misuraca Donia Msakni Giada Sgaravizzi Musafiri Karama Ebtesam Al-Olayan Saeed El-Ashram Marcelo Martinez-Barbitta Massimo Zerani Beniamino T. Cenci-Goga

The present study examined the effect of Enterococcus durans cell-free supernatant (CFS) on interleukin (IL) 8, 10 and 1β gene expressions in the intestinal cell line HT-29 treated with Staphylococcus aureus CFS. HT-29 cells were incubated with E. durans CFS or S. aureus CFS, or S. aureus CFS plus E. durans CFS. All concentrations of E. durans CFS did not show cytotoxicity, while the highest treatment (44.9 μg/mL) with S. aureus CFS induced significant cell death. S. aureus CFS did not modify IL-1β gene expression, while E. durans CFS alone or in combination with S. aureus CFS reduced it. Treatment with S. aureus CFS induced greater expression of the IL-8 gene compared to S. aureus CFS plus E. durans CFS. S. aureus CFS alone or in combination with E. durans CFS increased the expression of the IL-10 gene, while E. durans CFS alone did not modify it. These results suggest a potential protective role of the E. durans secretome in mitigating the inflammatory environment in intestinal cells. This treatment could be useful to protect against possible contact with dangerous soluble microbial products present in food.

Microbiology Research doi: 10.3390/microbiolres17050090

Authors: Petya Koeva Christova

Phytopythium species are water molds that have been divided as a separate group of oomycetes about 10 years ago. They are associated with diverse environments worldwide, but the ecological function of most of the species is still under investigation. In the present study, isolation and characterization of Pp. helicoides and Pp. palingenes are described. Both Phytopythium isolates originate from an aquatic environment and were derived from infected leaves. A host range of Pp. helicoides and Pp. palingenes using pathogenicity tests with leaves and cuttings from trees, bushes, perennial and herbaceous plants that belong to 15 different families was studied. Out of 21 tested plants, 18 were susceptible to infection with Pp. helicoides and nine were negatively affected by Pp. palingenes. Pp. helicoides is distinguished by higher aggressiveness and a wider range of potential host species. These results indicate that the impact of pathogenic species from the genus Phytopythium on forests and other natural ecosystems could be much more significant than is currently known.

Microbiology Research doi: 10.3390/microbiolres17050088

Authors: Stefano Petrini Mohamed Methnani Cecilia Righi Khaled El Hicheri Cristina Casciari Aida Tatli Ben Smida Boubaker Elena Tinelli Sana Kacem Claudia Pellegrini Roberto Sabato Francesco Feliziani Giovanni Pezzotti

Livestock farming represents a key economic activity in the Tataouine Governorate of southern Tunisia, where cattle and dromedary camels coexist. Varicellovirus bovinealpha1 (BoAHV-1), the etiological agent of infectious bovine rhinotracheitis (IBR), primarily affects cattle, while its circulation in camelids remains poorly understood. Following recent European Union regulations requiring BoAHV-1 surveillance in multiple animal species, this short communication reports serological findings from dairy cattle and dromedary herds in southern Tunisia. In March 2024, serum samples were collected from four non-vaccinated farms, including two intensive Friesian dairy cattle herds and two extensive dromedary herds (50 animals each). Serum samples from all animals were tested for BoAHV-1 antibodies using competitive commercial gB- and gE-based enzyme-linked immunosorbent assays (c-ELISA) and confirmed by virus neutralization test (VNT). Antibodies against BoAHV-1 were detected in cattle from both dairy farms, with low seroprevalence and neutralizing antibody titers, indicating past or ongoing exposure. In contrast, all dromedary samples tested seronegative by both c-ELISA and VNT. These findings confirm BoAHV-1 circulation in cattle in the Tataouine region and its absence in dromedaries at sampling. Further studies involving larger sample sizes and molecular investigations are required to clarify the potential role of camelids in BoAHV-1 epidemiology in southern Tunisia.

Microbiology Research doi: 10.3390/microbiolres17050087

Authors: Giorgia Piccinini Antonio Curtoni Alessandro Bondi Mattia Genco Fabio Longo Carlotta Polizzi Paolo Valesella Silvia Corcione Francesco Giuseppe De Rosa Cristina Costa

Antimicrobial resistance (AMR) represents one of the most urgent global health threats, significantly impacting patient outcomes, healthcare systems, and economic sustainability. Rapid and accurate antimicrobial susceptibility testing (AST) are essential to guide targeted therapy, reduce inappropriate antimicrobial use, and support antimicrobial stewardship programs. However, conventional phenotypic AST methods, including broth microdilution, disk diffusion, agar dilution, and gradient strip tests, remain labor-intensive and require prolonged turnaround times, often delaying optimal therapeutic decisions. Although automated commercial platforms such as VITEK 2, BD Phoenix, MicroScan WalkAway, and Sensititre ARIS have improved laboratory workflow and standardization, they still rely on culture-based approaches and typically require 16–36 h to generate minimum inhibitory concentration (MIC) results. In recent years, several innovative rapid phenotypic AST technologies have emerged, aiming to significantly shorten the time to susceptibility results while maintaining high accuracy. This review provides an overview of currently available rapid automated phenotypic platforms for MIC determination, including VITEK® Reveal™, ASTar, FASTinov®AST, QuickMIC®, and the Accelerate Pheno® system. These systems employ advanced technologies such as volatile organic compound detection, flow cytometry, microfluidics, real-time imaging, and morphokinetic cellular analysis to deliver susceptibility results within a few hours directly from positive blood cultures. We summarize their technical principles, antibiotics and pathogens included, performances, and current limitations. Overall, the implementation of rapid phenotypic AST tools has the potential to substantially improve clinical decisions, optimize antimicrobial therapy, and contribute to fight AMR.

Microbiology Research doi: 10.3390/microbiolres17050086

Authors: Waqas Ahmad Naeem Rasool Qurat ul Ain Usama Bin Naeem Muhammad Azeem Umbreen Anwar Tehreem Fayyaz Zeba Amjad Muhammad Yasin Tipu Mehmood Ahmad

Trypanosoma evansi (T. evansi) infection poses a significant health threat to equines. This study was aimed to assess the prevalence, risk factors, hematobiochemical alterations, and oxidative stress-mediated genotoxicity associated with equine trypanosomiasis in the Rahim Yar Khan District. This cross-sectional study was conducted on 384 equines from October 2024 to September 2025. Blood samples were collected for thin blood film microscopy and PCR assay using RoTat 1.2 primers. Hematological indices were analyzed with an automated hematology analyzer; serum biochemical parameters were quantified via standard assays. Oxidative stress markers, including malondialdehyde (MDA), catalase (CAT), superoxide dismutase (SOD), and reduced glutathione (GSH), were also measured. Genotoxicity was evaluated using the alkaline comet assay. Statistical analyses included the chi-square test, logistic regression, and independent t-tests. T. evansi was detected in 5.99% of samples by microscopy and 10.16% by PCR, with no significant association with species, age, or sex. Infected equines exhibited significant reductions in hemoglobin (5.4 ± 0.6 vs. 10.8 ± 0.5 g/dL; p < 0.0001), total serum protein (2.1 ± 0.3 vs. 5.8 ± 0.2 g/dL; p < 0.0001), albumin, and globulin, alongside elevated hepatic enzymes, blood urea nitrogen, and creatinine (all p < 0.01). Oxidative stress was confirmed by increased MDA (p < 0.0001) and decreased CAT activity (p < 0.001). Genotoxicity was significantly higher in infected animals (genetic damage index; 1.12 ± 0.08 vs. 0.40 ± 0.01; p < 0.01). This study provides the first integrated assessment of molecular epidemiology and oxidative stress-mediated genotoxicity in equines in this region, suggesting the pathogenic impact of the infection and targeted diagnostics for disease management strategies.

Microbiology Research doi: 10.3390/microbiolres17050085

Authors: Jun Iwahashi Akiko Shimizu Akinobu Togo Hiroshi Fuketa Kenji Gotoh Keisuke Ohta Norihiro Shinkai Naohisa Kawamura Hiroshi Watanabe

Background: Many of the pathogenic bacteria and fungi found in hospital environments form biofilms, which allow them to persist in the environment for long periods, posing a risk of hospital-acquired infections. Although the pathogens within biofilms often have reduced levels of drug susceptibility, the efficacy of disinfectants routinely applied against planktonic pathogens must be evaluated against biofilms as well. Our objective in this study was to determine the efficacy of treatment using slightly acidic hypochlorous acid water and to compare the results with sodium hypochlorite when both were used to disinfect Candida parapsilosis biofilms. Methods: C. parapsilosis in the planktonic or biofilm state was treated with each disinfectant. The number of viable cells that remained was determined, and scanning electron microscopy (SEM) of the disinfectant-treated biofilms was performed. Results: Compared with sodium hypochlorite, in a shorter period of time, hypochlorous acid water completely killed not only planktonic C. parapsilosis but also C. parapsilosis in a biofilm that had been formed for 72 h. SEM showed that both disinfectants were effective in removing the C. parapsilosis biofilm to some extent. Conclusions: Slightly acidic hypochlorous acid water appears to be an effective disinfectant against C. parapsilosis both in suspension and in biofilms.

Microbiology Research doi: 10.3390/microbiolres17040084

Authors: Linglin Tao Alessandro Ruan Shu Quan

Komagataella phaffii (formerly Pichia pastoris) is a prominent platform for recombinant protein production, yet secretion efficiency often remains a critical bottleneck. In this study, we validated three candidate genes—WHI3, CLPP, and PMP20—previously identified through genome-wide CRISPR activation screening, for their potential to enhance heterologous protein secretion. Overexpression of these factors under the control of the methanol-inducible AOX1 promoter increased the secretion of human serum albumin (HSA), with WHI3 and CLPP yielding improvements of 18.3% and 17.9%, respectively. Furthermore, applying this strategy to human lactoferrin (hLF) revealed that WHI3 overexpression robustly enhanced hLF secretion by approximately 70%. Comparative analysis of different promoters (AOX1, GAP, and CAT) indicated that the AOX1 promoter remains the most effective driver for these enhancers, suggesting a threshold-dependent regulatory mechanism. These results demonstrate the protein-dependent nature of secretion optimization and identify WHI3, CLPP, and PMP20 as novel, effective co-expression factors for improving recombinant protein yields in K. phaffii.

Microbiology Research doi: 10.3390/microbiolres17040083

Authors: Natália Cristina de Melo Santos Evandro Neves Silva Leonardo Pereira de Araújo Carlos Roberto Prudêncio Rômulo Dias Novaes Patrícia Paiva Corsetti Leonardo Augusto de Almeida

Differences in the gut microbiota are directly reflected in lung–gut axis crosstalk, which may increase susceptibility to pulmonary infections, such as those caused by the bacterium Pseudomonas aeruginosa. Deficiency of the cytokine IL-10 leads to gut inflammation, and this pro-inflammatory environment is partly due to changes in the gut microbiota. To better understand the effects of IL-10 deficiency on the gut microbiota, the intestinal microbial composition of IL-10 KO mice was assessed, and an increase in the phyla Bacteroidetes and Proteobacteria and a decrease in the phylum Firmicutes were observed in the faeces compared with the wild-type group (WT). Additionally, IL-10 KO mice had a higher pro-inflammatory immunostimulatory caecal content. Furthermore, it was found that heterologous faecal microbiota transplantation (FMT) between groups reversed this gut imbalance. IL-10 KO mice showed greater susceptibility to acute pulmonary infection by P. aeruginosa, with a higher recovery of viable bacteria in the lung and spleen, greater tissue damage and increased expression of genes encoding pro-inflammatory cytokines in the lungs. This greater susceptibility was reversed after FMT. Taken together, these results demonstrate the role of endogenous IL-10 in the gut microbiota constitution and its importance in the pulmonary immune response against P. aeruginosa infection.

Microbiology Research doi: 10.3390/microbiolres17040082

Authors: Eleni Alaverntian Eugenia Papadaki

The gut microbiome plays a central role in human health, and probiotics are widely used to support microbial balance, though their efficacy depends on multiple factors. This study assessed the potential of commercial probiotics Saccharomyces cerevisiae var. boulardii, Weizmannia coagulans and Lacticaseibacillus rhamnosus by evaluating in vitro gastrointestinal kinetics, pharmaceutical stability, and antioxidant effects in chamomile tea. Growth across a broad pH range was modeled kinetically, while survival and inactivation were quantified in simulated gastric and intestinal fluids. Antibiotic and antifungal susceptibility was determined using disk diffusion, and antioxidant activity of fortified chamomile tea was assessed via DPPH radical scavenging. Results revealed distinct strain-dependent responses. S. cerevisiae var. boulardii and W. coagulans showed the highest gastrointestinal tolerance. The increase in fluid volume reduced survival during the gastric phase but improved survival in the intestinal phase, reflecting different stress responses. Antimicrobial susceptibility also varied, with S. cerevisiae var. boulardii exhibiting the highest resistance. Probiotic fortification enhanced chamomile tea’s antioxidant capacity, particularly for S. cerevisiae var. boulardii and L. rhamnosus. These findings provide quantitative insight into strain- and volume-dependent gastrointestinal performance, guiding the optimization of capsule formulations and the development of clean-label products combining probiotic and antioxidant benefits.

Microbiology Research doi: 10.3390/microbiolres17040081

Authors: Fatma A. Abo Nouh Ahmed M. Abdel-Azeem Tamer S. Abdelmoneim Nivien A. Nafady Saeed Mohammadi Najeeb Ur Rehman Hassan Moghtaderi Moosa Al Hamadani Saif Al-Housni Usama Qayum Abdullah M. S. Al-Hatmi

The emergence of antimicrobial resistance and the increasing incidence of cancer have highlighted the urgent need to develop new drugs; therefore, the discovery of new bioactive molecules is an important goal for future research. In this study, freshwater fungi isolated from submerged Phragmites australis from Egypt were screened for antimicrobial and cytotoxic activities. Using ITS1 and ITS4 primers, eight frequently occurring Sordariomycetes taxa were identified and were then selected for further evaluation of bioactivity. Ethyl acetate crude extracts (A–H) were evaluated for antimicrobial activity using the agar disk-diffusion method. Extracts A and E, derived from Chaetomium globosum SCUF0000404 (PX596738) and Chaetomium madrasense SCUF0000401 (PX596735), respectively, showed broad-spectrum activity at 100 mg/mL against bacterial pathogens, including Staphylococcus aureus ATCC 29213 (15.33 and 18.00 mm), Streptococcus pyogenes ATCC 19615 (11.00 mm), Escherichia coli ATCC 35218 (10.33 and 10.67 mm), Klebsiella pneumoniae ATCC 700603 (14.00 and 16.67 mm), and Pseudomonas aeruginosa ATCC 27853 (13.33 and 16.33 mm), and show antifungal activity against Candida albicans ATCC 14053 (20.33 mm), Candida krusei ATCC 6258 (15.67 and 15.33 mm), Trichosporon asahii AMS 187 (17.00 and 17.67 mm), Exserohilum rostratum AMS 1077 (34.00 and 33.67 mm), and Trichophyton indotineae AMS 180 (38.33 and 34.00 mm). Selective cytotoxic effects on the breast cancer cell line MDA-MB-231 were observed by extracts A and E at IC50 = 309 and 277 μg/mL, while non-selective cytotoxic effects on the normal HUVEC cell line were found with IC50 = 919 and 796 μg/mL, respectively. Characterization of the most effective extracts A and E by high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) shows that they have a wide range of secondary metabolites, including cytochalasans, azaphilone alkaloids, steroids, terpenoids, flavonoids, and phenols. These findings underscore the chemical diversity and therapeutic potential of freshwater fungi from Egypt.

Microbiology Research doi: 10.3390/microbiolres17040080

Authors: Claudia Berenice Tinoco-Cabral Luis Alfonso Muñoz-Miranda Manuel R. Kirchmayr Vianeth Martínez-Rodríguez Miguel Padilla-Rosas Maricarmen Iñiguez-Moreno Suchiquil Rangel-Velázquez Fabiola Berenice Hernández-Reyes Claudia Lisette Charles-Niño Cesar Arturo Nava-Valdivia

Candida species can shift from commensal organisms to opportunistic pathogens. Both Candida albicans and non-albicans Candida (NAC) species colonize oral biofilms and periodontal pockets, where they may contribute to inflammation and the progression of periodontal disease. This study aimed to determine the prevalence and antifungal susceptibility profiles of Candida species in individuals with different stages of periodontal disease. A cross-sectional study was conducted in 100 participants whose periodontal status was clinically evaluated. Saliva samples were cultured on chromogenic agar for yeast isolation, species identification was confirmed by MALDI-TOF MS, and antifungal susceptibility to fluconazole, clotrimazole, nystatin, and amphotericin B was assessed. Candida spp. was detected in 35% of participants, where C. albicans was the most prevalent species, followed by Nakaseomyces glabratus (formerly Candida glabrata), Candida parapsilosis, Candida dubliniensis, and Candida tropicalis. Species distribution varied according to periodontal status, with N. glabratus predominating in early periodontitis and C. albicans appeared more frequently in higher severe stages of periodontitis. Susceptibility testing showed resistance of C. albicans to clotrimazole (63.6%) and nystatin (22.7%), whereas amphotericin B and fluconazole remained effective. NAC species, particularly N. glabratus, exhibited resistance to nystatin and variable resistance to clotrimazole but remained susceptible to amphotericin B. These findings underscore the importance of early detection and personalized antifungal strategies for managing periodontal disease complicated by Candida colonization.

Microbiology Research doi: 10.3390/microbiolres17040079

Authors: Armando Navarro-Ocaña Armando Navarro-Cid del Prado Ricardo Ernesto Ahumada-Cota Ulises Hernández-Chiñas

Intestinal infections caused by Salmonella enterica serovar Typhi (S. Typhi) remain a global health concern, making preventive strategies and diagnostic tools essential. This study aimed to identify mimotope peptides of the Vi antigen using phage display and assess their recognition by rabbit and 46 human sera, as well as their potential for diagnosis and immunogen design. Rabbits were immunized with the Vi antigen (AgVi) from S. Typhi ATCC 6539, and sera-derived IgG was used for phage biopanning. DNA sequences from selected phagotopes were synthesized as Salmonella mimotope peptides (SMPs), either linear or KLH-conjugated. Their reactivity was tested with ELISAs against AgVi and SMPs, using both rabbit sera and 46 human serum samples. Ten phagotopes were identified, with a consensus motif (D/G–A/V–x–P–x–x–G–x–x–x–x–x), suggesting α-helix structures. Immunization with KLH-conjugated peptides generated specific antibodies, particularly SMPVi/5 and SMPVi/10, which recognized AgVi and their respective peptides. Competitive inhibition assays confirmed that SMPVi/5 reduced the anti-AgVi binding in a dose-dependent manner. In human sera, AgVi recognition occurred in 52% of samples, while SMPVi/5 and SMPVi/10 were recognized in 45%. Overall, SMPVi/5 demonstrated immunogenicity and functional mimicry, supporting its use as a synthetic reagent for serological assays and as a candidate for immunogen design.

Microbiology Research doi: 10.3390/microbiolres17040078

Authors: Shubhangi Kansal Kavita Gupta Shubhneet Kaur Mamik Neelam Taneja Archana Angrup

The rise in multidrug resistance in Klebsiella pneumoniae is an alarming issue, especially in invasive infections among patients with co-morbidities. With the gain of hypervirulence traits, multidrug-resistant K. pneumoniae has led to a significant increase in chronic infections and associated mortality. This study aims to explore the distribution of multidrug-resistant and hypervirulent (hv) K. pneumoniae in invasive infections in a tertiary care hospital. A total of 231 K. pneumoniae isolates were collected over a period of six months from invasive infections. These isolates were tested phenotypically and genotypically for the presence of antimicrobial resistance, along with molecular detection of hypervirulence determinants (iucA, rmpA, rmpA2, peg344, iroB). High levels of resistance to β-lactams, fluoroquinolones, and aminoglycosides were observed. Carbapenemase-encoding genes were widely distributed, and 22% showed the presence of at least one hypervirulence gene, most commonly iucA and rmpA. Co-carriage of resistance and hypervirulence determinants in K. pneumoniae was observed in nearly 20% of the isolates, indicating the emergence of MDR-hvKP phenotypes in the hospital setting. Mortality was significantly higher among patients infected with MDR isolates, whereas hypervirulence markers were not independently associated with mortality. The presence of MDR–hypervirulent strains remains clinically concerning and underscores the need for continued genomic surveillance.

Microbiology Research doi: 10.3390/microbiolres17040077

Authors: Edmar Gonçalves Pereira Filho Stéfanne Rodrigues Rezende Ferreira Amanda Veiga Paiva Simões Eli Júnior Pereira Rodrigues Iorrana Morais de Oliveira Marillia Lima Costa Adeliane Castro da Costa Berendina Elsina Bouwman Hanstter Hallison Alves Rezende

The objective of this study is to evaluate the efficiency of surgical instruments’ manual cleaning versus automated cleaning in an ultrasonic cleaner for the removal of biofilms on surgical forceps contaminated with Staphylococcus epidermidis and Pseudomonas aeruginosa. Subsequently, the residual microbial load was quantified through microbiological culture, aiming to evaluate the effectiveness of biofilm removal under different reprocessing conditions. Cleaning is an essential step in the processing of surgical instruments to ensure the effective removal of dirt and microorganisms. Through adhesion, microorganisms can attach to surfaces and form biofilms, organized structures surrounded by an extracellular matrix consisting of various components, which favor metabolic exchanges, adaptation, resistance, and bacterial dispersion. These biofilms increase the pathogenic potential of microorganisms, contributing to the occurrence of Healthcare-Associated Infections, and to avoid these, it is essential that preventive measures aimed at microbial reduction are adopted. Automated cleaning proved more effective than manual cleaning, and the combined approach achieved the greatest microbial reduction, though persistent contamination was still observed. The ability of adhesion and biofilm formation on the surfaces of surgical instruments is regarded as a challenge for complete microbial removal. These findings enhance the need for more rigorous reprocessing protocols and complementary strategies to ensure greater safety in the use of reusable instruments in clinical practice.

Microbiology Research doi: 10.3390/microbiolres17040076

Authors: Eugenia Guadalupe Ortiz-Lechuga Verónica Almaguer-Cantú Hiram Herrera-Barquín Karla Katiushka Solís-Arévalo Ramón Alberto Batista-García Katiushka Arévalo-Niño

Biosurfactant-producing microorganisms play an important ecological role in soils impacted by hydrophobic contaminants by enhancing substrate bioavailability and influencing microbial interactions. In this study, we critically evaluated the reliability of commonly used screening methods for biosurfactant detection. A total of 71 microbial isolates (16 bacteria and 55 fungi) were obtained from vegetable oil-contaminated soil and screened using a multi-step approach combining enzymatic assays (lipolytic and hemolytic activity) and physicochemical methods, including drop-collapse, oil spreading, emulsification index (E24), and surface tension reduction. Although 21 isolates exhibited lipolytic activity and 9 showed hemolysis, inconsistent responses among assays revealed significant limitations of individual screening methods. Only two bacterial isolates consistently tested positive across all criteria. When cultivated in mineral salt medium supplemented with hydrophobic substrates, both isolates produced stable emulsions and significantly reduced surface tension (from 54.26 mN/m to 31.46 mN/m). Substrate-dependent variation was observed for isolate C3, which showed reduced surface tension (39.63 mN/m) when grown with biodiesel. These findings highlight the risk of relying on single assays and emphasize the need for integrated screening strategies to ensure reliable detection of biosurfactant-producing microorganisms.

Microbiology Research doi: 10.3390/microbiolres17040075

Authors: Doris Evelyn Yah Hui Jong Siang Yin Lee Yun Khoon Liew Phyu Synn Oo Amar Harris Arifin Zi Ni Ngai Beek Yoke Chin Shamala Salvamani Rhun Yian Koh

In the original publication [...]

Microbiology Research doi: 10.3390/microbiolres17040074

Authors: Ajran Kabir Rosbelly Rios Mohamed Saleh Daniel Mallal Barbara L. Whitt Jaden Thompson Beatrice T. Sponseller Nathan M. Slovis Mats H. T. Troedsson Hossam El-Sheikh Ali Yosra A. Helmy

Klebsiella pneumoniae has been associated with reproductive infections in equines. The detection of β-lactam resistance determinants, especially extended-spectrum β-lactamase (ESBL) genes, within genomic regions linked to horizontal gene transfer (HGT), is of a particular concern. In this study, we characterize the whole-genome sequences (WGS) of three K. pneumoniae equine isolates harboring multiple antimicrobial resistance genes. Two isolates were recovered from uterine washes of mares: one with endometritis (YAH-KPEM1) and one clinically normal (YAH-KPSE1), and a third from the feces of a diarrheic foal (YAH-KPF132). WGS was performed using the Illumina MiSeq platform, and the reads were subsequently processed through hybrid assembly in Unicycler v0.5.1. Genome annotation was completed using PROKKA v1.14.5. Strain YAH-KPEM1 was classified as ST127, whereas YAH-KPSE1 and YAH-KPF132 belonged to ST1630 and ST224, respectively. Notably, K. pneumoniae ST1630 and ST224 have not been reported before in equines. All three genomes encoded multiple antimicrobial resistance (AMR) determinants, including two encoding ESBL genes (CTX-M-15), as well as virulence factors and regions associated with HGT. Additionally, two (YAH-KPEM1 and YAH-KPSE1) isolates were found to be multidrug resistant (MDR), harboring an IncFIB(K) plasmid replicon, and another isolate, YAH-KPF132, carried an IncFII replicon. The detection of AMR and virulence genes in equine Klebsiella isolates has important clinical implications for guiding antimicrobial selection and improving treatment success.

Microbiology Research doi: 10.3390/microbiolres17040073

Authors: Zhuliang Jiang Yukai Zeng Dingping Liu Yuanjing Li

Castanopsis hystrix (C. hystrix) is one of the most dominant and ecologically important species in subtropical evergreen broad-leaved forests of China. Interactions between its root and rhizosphere microorganisms play a pivotal role in nutrient acquisition and in mediating plant response s to environmental stresses. In this study, high-throughput 16S ribosomal RNA (16S rRNA) sequencing combined with untargeted metabolomics was employed to systematically characterize the rhizosphere microbial community and root exudates in C. hystrix. The results showed that, compared with non-rhizosphere soil, bacterial diversity in the rhizosphere of C. hystrix was significantly reduced, while several specialized and potentially efficient taxa were selectively enriched, particularly Candidatus_Solibacter, Candidatus_Xiphinematobacter, and Candidatus_Koribacter, thereby reshaping a distinct rhizosphere-specific community structure. Metabolomic analyses further revealed that 129 metabolites were significantly enriched in the rhizosphere, including four major classes of compounds associated with plant stress resistance: lipids and lipid-like molecules, organoheterocyclic compounds, organic acids and derivatives, and phenylpropanoids and polyketides. The enrichment of these metabolites likely contributes substantially to stress tolerance in C. hystrix. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis identified six defense-related metabolic pathways, including pyrimidine metabolism, steroid biosynthesis, nucleotide metabolism, plant hormone signal transduction, ATP-binding cassette transporter (ABC transporters), and the biosynthesis of various plant secondary metabolites. Further correlation analysis and co-occurrence network analysis suggested that C. hystrix may potentially influence the enrichment of beneficial microorganisms through rhizosphere metabolites selectively, which could reduce the reliance on external nutrient acquisition and enhance the stress resilience of C. hystrix. Our study provides a comprehensive perspective for elucidating rhizosphere interaction networks and their ecological functions in C. hystrix, thereby enhancing our understanding of the environmental adaptability of dominant tree species in subtropical forests.

Microbiology Research doi: 10.3390/microbiolres17040072

Authors: Sookyung Oh Bradd J. Haley Jitendra Patel

Whole-genome sequencing followed by comprehensive genomic analyses was used to characterize 16 Salmonella isolates from water-overlying sediments in Conococheague Creek (PA), an agricultural irrigation water source. Our goal was to characterize the genomic profiles and diversity of these Salmonella isolates. We identified eight distinct serotypes, including Newport, the most prevalent (43.8%), providing environmental context relevant to agricultural water systems. Genomic surveys showed various Salmonella Pathogenicity Island (SPI) profiles. Although widespread antimicrobial resistance (AMR) genes were not detected, the consistent presence of the aac(6’)-Iaa gene across all isolates and a parC (T57S) mutation in 14 isolates were identified as inherent genotypic markers. Six distinct plasmid replicon types were observed in over 60% of isolates. Replicons for IncF and IncI2 plasmids, frequently associated with β-lactamase genes, were found, documenting the presence of mobile genetic elements despite a lack of acquired AMR genes. Restriction-Modification (RM) systems and CRISPR/Cas loci were also detected, suggesting Salmonella genomic plasticity. Our study showed that sediment-associated Salmonella, notably serotype Newport, harbored diverse virulence-associated genomic features. These findings contributed to the genomic baseline for irrigation water quality and food safety.

Microbiology Research doi: 10.3390/microbiolres17040071

Authors: Xuanxuan Feng Zhenghua Xie Jianmin Wang Xinzhou Yang

While the gut microbiota of pollen-carrying bees (e.g., honeybees and bumblebees) has been well studied, the gut microbiota of vespine wasps remains poorly understood. Unlike pollen-carrying bees, which primarily consume pollen and nectar, vespine wasp larvae mainly feed on insects, suggesting that their gut bacterial communities may be different. Therefore, this study aimed to reveal the differences in larval gut bacterial communities between pollen-carrying bees and vespine wasps. Using datasets obtained from our own samplings and public resources released by other researchers, we compared the diversity and composition of larval gut bacterial communities between vespine wasps and pollen-carrying bees. Alpha diversity and beta diversity of bacterial communities were measured. Results showed that vespine wasp larvae harbored distinct gut bacterial communities from those of pollen-carrying bees, dominated by Leuconostoc, Hafnia-Obesumbacterium and Lactobacillus. Significant differences in bacterial composition were observed at both the community level and the dominant taxa level between pollen-carrying bee larvae and vespine wasp larvae. Moreover, significant differences were also found among larval gut bacteria of vespine wasps. These findings provide insights into the bacterial composition of aculeate wasps with different dietary habits.

Microbiology Research doi: 10.3390/microbiolres17040070

Authors: Anna-Marie Ilic Jan Werner Böhm Michael Kube

Increasing genomic data are driving changes in the selection of phylogenetic markers and analysis strategies. Databases enable the extraction of established markers, such as single and multilocus sequence typing (MLST), but are often limited by the number of informative sites or availability with respect to incomplete source data sets or reductive evolution in bacteria such as the Mollicutes. Genome-wide analyses like average nucleotide identity (ANI) often overcome these problems but also depend on the alignment percentage. Complementary analyses help validate results and address limitations of primary approaches. However, how genome-wide compositional signals and reduced core gene sets affect phylogenomic resolution across a large and taxonomically diverse dataset of complete Mollicutes genomes remains unclear. Therefore, we applied an advanced MLST approach based on single-copy orthologs (SCOs), alongside codon usage analysis. The reliability and impact of these approaches were first analyzed using Acholeplasmatales as the foundation, with 16S rRNA gene, ANI, SCOs, and codon usage. Codon usage analysis revealed lineage-associated compositional signatures across the 52 strains that were broadly consistent with current genus and subgroup assignments, whereas ANI and 16S rRNA gene identified species with ≥96.5% and ≥97%, respectively. Among these, SCOs showed the most matches to the current taxonomy, supporting the approach being extended to Mollicutes. Applied to 807 Mollicutes strains, the analysis revealed 16 shared SCOs. Concatenation of this core set significantly enhanced phylogenomic resolution, providing a robust framework for reconstructing evolutionary relationships within Mollicutes.

Microbiology Research doi: 10.3390/microbiolres17040069

Authors: Olga Jakovljević Željka Milovanović Miloš Stupar Željko Savković Marija Pećić Dragana Jerinkić Slađana Popović

Cave microbiota comprise metabolically diverse organisms, including microalgae, among which Bacillariophyta (diatoms) represent one of the most prominent groups, inhabiting a wide range of substrates within cave ecosystems. In contrast to aerophytic cave habitats, aquatic cave environments remain poorly studied. Therefore, the main aims of this study were to determine the diversity, spatial distribution, and seasonal dynamics of diatom assemblages in the Ponorac Stream flowing through Ravništarka Cave, and to assess the influence of environmental variables on diatom diversity and distribution. Samples were collected from six sites along the Ponorac stream in May and November 2023. Physical and chemical water parameters showed only minor variation among sampling sites. In total, 148 diatom taxa belonging to 54 genera were recorded, including several rare diatom taxa. Diatom assemblages in the Ponorac stream were characterized by high taxonomic richness, high α-diversity, and pronounced community heterogeneity. Many taxa occurred in both seasons and across multiple sites, whereas several were restricted to a single season or exhibited clear site specificity. Most diatom index values indicated generally high ecological status. This study highlights the importance of aquatic cave habitats as reservoirs of diatom diversity and their value in studying temporal and spatial variation of their communities.

Microbiology Research doi: 10.3390/microbiolres17040068

Authors: Sabīna Ribačuka Viktorija Bankoviča Ingus Skadiņš

The global rise in antimicrobial resistance has spurred increased interest in alternative antimicrobial agents, particularly essential oils (EOs). These oils are complex mixtures of volatile compounds that exhibit documented biological activity. This study evaluated antimicrobial and antibiofilm effects of selected EOs against clinically relevant bacterial and fungal pathogens. Antimicrobial activity against planktonic cells was assessed using disc diffusion assays with DMSO-diluted EO solutions against Escherichia coli (E.coli), Staphylococcus aureus (S.aureus), Pseudomonas aeruginosa, Klebsiella pneumoniae, and Candida albicans. Antibiofilm activity of E. coli and S. aureus was examined using ethanol-based EO formulations, with biofilm viability quantified by colony forming unit (CFU) enumeration. Cinnamon (Cinnamomum verum) oil showed the strongest and most consistent activity, inhibiting planktonic and biofilm models. Tea tree (Melaleuca alternifolia), lemongrass (Cymbopogon citratus), rosemary (Rosmarinus officinalis), rose (Rosa damascena), and jasmine (Jasminum officinale) oils showed significant planktonic antimicrobial effects, while jasmine oil (Jasminum officinale) demonstrated pronounced antibiofilm activity against S. aureus, including strong biofilm eradication in several replicates. In contrast, chamomile (Matricaria chamomilla) and sandalwood (Santalum austocaledonicum) oils showed limited or no activity. These findings highlight differences between planktonic and biofilm responses, emphasizing the importance of incorporating biofilm models into antimicrobial evaluation. Overall, Cinnamomum verum and Jasminum officinale oils may serve as complementary antimicrobial agents, warranting further investigation.

Microbiology Research doi: 10.3390/microbiolres17040067

Authors: Gagandeep Brar Ramandeep Kaur Mandeep Kaur Gill Navjot Singh Rupinderjeet Kaur

Climate change is intensifying winters in temperate regions, posing a serious threat to Apis mellifera health. The gut microbiome, a distinct community of core bacterial species, is central to overwintering success by supporting immune function, nutrient assimilation, and pathogen resistance, but is highly sensitive to environmental stressors such as cold temperatures and dietary shifts. Stress-induced perturbations can reshape the composition and relative abundance of the gut microbiome in honey bees, leading to adverse effects on host health, physiological functions, and overwinter survival. Cold temperatures and additional stressors further destabilize the microbiome, compounding these effects. This review is the first to synthesize current knowledge on how extrinsic factors, such as diet, antibiotics, and pathogens, and intrinsic factors, including age and strain, influence the composition and function of the honey bee gut microbiota during the overwintering period. Given the increasing severity of winter conditions under climate change, a deeper understanding of microbiome–host–environment interactions is essential for improving honey bee resilience. By integrating evidence on the microbiome’s roles in nutrient utilization, immune modulation, and pathogen defense, this review outlines a framework to guide future research aimed at sustaining pollinator health and nutrition in a changing global climate.

Microbiology Research doi: 10.3390/microbiolres17040066

Authors: Yaodong He Zhengwei Peng Fenglin Wang Peitao Liu Shirui Mu Yaqiong Wang Xiumei Zhang

Eukaryotic microorganisms, including microalgae, protists, fungi, and micro-metazoans, act as drivers of energy flow and nutrient cycling, collectively forming the microbial food loop, and also serve as important indicators of environmental health. To investigate the seasonal variation in eukaryotic microorganisms in a mussel farming area, a total of 96 seawater samples were collected from surface and bottom layers of water across different seasons. High-throughput sequencing of the 18S rRNA gene was employed to characterize shifts in microbial community structure and identify key influencing factors. Our results indicated significant seasonal differences in eukaryotic microbial communities between surface and bottom waters. Redundancy Analysis (RDA) revealed that seasonal variations in community structure were primarily driven by environmental factors such as temperature, dissolved oxygen (DO), and salinity. Co-occurrence network analysis indicated that surface water networks exhibited higher numbers of nodes and edges, as well as greater modularity, suggesting more distinct niche differentiation and higher natural connectivity within the community. These findings provide fundamental data for understanding the response mechanisms of eukaryotic microbial communities to seasonal changes in the mussel cultivation area of Gouqi Island.

Microbiology Research doi: 10.3390/microbiolres17040065

Authors: Mayur J. Nath Shubham R. Choudhury Priyanka Sourav Khan Manabendra Mandal Sanjay K. S. Patel

This study investigated the antibiofilm and anti-quorum-sensing (QS) potential of endophyte extracts isolated from medicinal plants and their validation against Pseudomonas aeruginosa. Endophytes were isolated from the plants using the serial dilution method, and the extracts produced by these endophytes were screened for antimicrobial and biofilm-inhibition activity using assays. The efficient extract was biochemically characterized, followed by validation of its secondary metabolite content. Furthermore, QS-regulatory gene expression levels and microscopy were used to confirm inhibition of biofilm formation. A total of 12 cultures, including 8 bacterial and 4 fungal, were isolated and screened, demonstrating efficient antimicrobial activity (zone of inhibition of 18.8 mm) and 64.3% antibiofilm activity. The efficient endophyte isolated from Paederia foetida was identified as Bacillus xiamenensis MM07 by 16S rRNA gene sequencing. MM07 extract analyses by biochemical and Fourier transform infrared methods revealed the presence of diverse biomolecules. A dose-dependent inhibition was observed, achieving up to 83.5, 60.3, 73.2, 82.7, 83.2, and 15.1 in biofilm formation and exopolysaccharide, violacein, pyocyanin, protease, and alginate production, along with 63.2% swimming ability at 30 µg/mL, respectively. Gas chromatography-mass spectrometry analyses validated the presence of major secondary metabolites, including 3,3-dimethyl-4-methylamino-butan-2-one, 6-amino-2-methyl-, 1-iodo-2-methylundecane, and hexadecanoic acid, with the potential to inhibit biofilm and QS activity. Quantitative real-time polymerase chain reaction of QS regulatory genes (lasI, lasR, rhlI, and rhlR) and microscopy analysis confirmed the anti-QS properties, evidenced by a 40.3% decline in gene expression and biofilm inhibition by MM07 extract. These findings highlight the potential of novel B. xiamenensis MM07 endophyte from P. foetida as a sustainable source of biomolecules for combating biofilm-associated infections.

Microbiology Research doi: 10.3390/microbiolres17030064

Authors: Shaun Cawthraw Andrew Wales Tom Huby Rob Davies

Introduction: Salmonella may contaminate livestock feed at several stages of production, transport and storage. Formaldehyde is an effective anti-Salmonella feed treatment, but it is now banned for this use in Europe. Organic acid-based additives are an alternative. Gap Statement: The efficacy of organic acid feed additives against natural Salmonella feed contamination is uncertain due to a paucity of reported work investigating low levels of infection that may be relevant for real-world situations. Aim: To compare the anti-Salmonella effects of feed additives based on formaldehyde versus those based on organic acids. Methodology: Experimental contamination of poultry feed with one of three Salmonella serovars at moderate (between 10 and 200 CFU/g) or low (around 1 CFU/g) levels was preceded (‘prevention’ mode) or followed (‘decontamination’ mode) by application of commercial antimicrobial additives. Storage at room temperature for 24 h was followed by pre-enrichment then culture. Results: Organic acid-based products at recommended application rates only eliminated detectable Salmonella from samples with the lowest degree of contamination. The effect was partial, with a proportion of samples still yielding Salmonella in most experiments, and only one such product showed efficacy above 50% of samples for the decontamination mode. The two formaldehyde-based products showed partial efficacy against moderate contamination, and one was entirely effective against low-level contamination even at its lower inclusion rate. Conclusions: Organic acid-based feed additives have a lesser anti-Salmonella effect than formaldehyde-based products at their respective recommended inclusion rates. However, some non-formaldehyde products may be substantially effective against a low, natural degree of contamination. Impact Statement: Chemical suppression of Salmonella in animal feed is an important element of measures to safeguard livestock health and, consequentially, public health too. The European ban on using formaldehyde for this purpose has necessitated the use of alternative products. The present work includes very low levels of Salmonella in feed, possibly mimicking natural contamination, to show that under these circumstances some such alternatives may be as efficacious as formaldehyde products.

Microbiology Research doi: 10.3390/microbiolres17030063

Authors: Khin Zar Linn Su Zar Chi Lwin Aye Thida Maung Marwa Nabil Sayed Abdelaziz Catherine Damaso Hofilena Yuzhi Lin Haomin Ye Yoshimitsu Masuda Takahisa Miyamoto Ken-ichi Honjoh

Nontyphoidal Salmonella (NTS) is a zoonotic pathogen that threatens public health worldwide. This study investigated the prevalence, serotype, virulence, and antimicrobial resistance of NTS isolated from chicken meat in Fukuoka, Japan. Of 50 samples, 64% were positive for Salmonella spp., and 32 NTS strains were isolated from positive samples. Serotyping identified three serotypes: S. enterica ser. Schwarzengrund (78.1%), S. enterica ser. Thompson (15.6%), and S. enterica ser. Oranienburg (6.3%). Multilocus sequence typing revealed three sequence types (STs), and MALDI-TOF MS analysis revealed six distinct clusters, reflecting heterogeneity in protein expression among isolates with the same STs. All isolates harbored the virulence genes hilA, spiC, and ssrB, but not spvC. Microplate assays showed that all S. enterica ser. Schwarzengrund and S. enterica ser. Thompson strains formed biofilms with varying strengths. Antimicrobial susceptibility tests demonstrated that S. enterica ser. Thompson and S. enterica ser. Oranienburg strains were sensitive to all the antimicrobials tested. However, S. enterica ser. Schwarzengrund strains showed resistance to multiple antibiotic classes, and 36% of the isolates were multidrug resistant. These findings suggest a potential public health concern, particularly from S. enterica ser. Schwarzengrund, and underscore the importance of continuous surveillance that integrates both genotypic and phenotypic methods.

Microbiology Research doi: 10.3390/microbiolres17030062

Authors: Zizipho Z. A. Mbulawa Laston Gonah Lindiwe M. Faye Charles B. Businge

Anal human papillomavirus (HPV) and cancer prevalence are increasing. Therefore, this study investigated the prevalence of anal HPV and associated risk factors, as well as HPV genotype-specific concordance at cervical and anal sites and associated risk factors among women of Eastern Cape Province, South Africa. A total of 326 women aged 18–60 were recruited from an Eastern Cape community health facility. HPV DNA was detected in cervical and anal specimens using the Seegene Anyplex™ and Allplex™ II HPV28 assay (Seegene Inc., Seoul, Republic of Korea), respectively. Anal HPV was detected in 68.1% (95% CI: 62.9–72.9) and independent predictors were cervical HPV positivity (AOR: 2.40, 95% CI: 1.39–4.14, p = 0.002), abnormal cytology (AOR: 3.12, 95% CI: 1.29–7.55, p = 0.012), single marital status (AOR: 3.55, 95% CI: 1.24–10.17, p = 0.018), and having more than three lifetime sexual partners (AOR: 1.75, 95% CI: 1.03–2.98, p = 0.039). Anal high risk (HR)-HPV types were detected in 50.9%, with HPV-58 (13.2%), HPV-68 (11.0%) and HPV-52 (9.2%) being the most dominant types. HPV genotype-specific cervical and anal concordance was observed in 33.5% of cases, with HPV-58 (7.1%), HPV-68 (4.9%), and HPV-35 (4.6%) being the most dominant. Women who were positive for cervical HPV infection (AOR: 3.24, 95% CI: 2.36–4.45, p < 0.001), anal HPV infection (AOR: 2.70, 95% CI: 2.01–3.63, p < 0.001) and abnormal cervical cytology (AOR: 2.01, 95% CI: 1.36–2.96, p < 0.001) had substantially higher odds of anal–cervical HPV concordance compared to those who were negative. High anal HPV prevalence and HPV genotype-specific anal and cervical concordance were observed among Eastern Cape women. Understanding anal HPV, HPV genotype-specific anal–cervical concordance, and associated factors can contribute to strategies towards anal HPV and associated disease prevention. These findings warrant further longitudinal investigation in future studies.

Microbiology Research doi: 10.3390/microbiolres17030061

Authors: Vincenzo Verdura Pasquale Bisceglia Luigi Annacontini Luigi Cagiano Francesca Sanguedolce Martina Miracapillo Fabrizia Fusco Sergio Lo Caputo Francesco Drago Gaetano Serviddio Aurelio Portincasa Giulia Ciccarese

Dermatofibrosarcoma protuberans (DFSP) is a rare tumor presenting as a slow-growing, plaque-like or multinodular, brownish lesion on the trunk in adult patients. Diagnosis is established by histological examination and surgical excision is the primary treatment. Typically, DFSP has an indolent course and local spread. In the present work, we describe the clinical–histologic features, surgical treatment and follow-up of a case of DFSP in a patient living with HIV infection (PLWH). A 40-year-old man was referred to us with confluent lesions on the left shoulder, present for about 3 years. His medical history was positive for HIV-1 infection, for which he was taking antiretroviral therapy. Microscopic examination showed dermal and hypodermic proliferation of spindle cells in a storiform pattern, confirming the clinical diagnosis of DFSP. A wide excision was performed with 3 cm clinically healthy tissue margins, and the defect was repaired using an artificial bilaminar dermal matrix. The histological examination revealed tumor-free margins, and a split-thickness skin graft was harvested from the same arm. After 10 months, the patient was free from the disease. As observed with other skin cancers, DFSP may have a higher incidence and greater aggressiveness in immunosuppressed than in immunocompetent patients. DFSP has been reported only twice in PLWH. Our case constitutes a third report, adding to the evidence that there may be an over-representation of this cancer in immunosuppressed individuals.

Microbiology Research doi: 10.3390/microbiolres17030060

Authors: Jianmin Liu Mingliang Geng Yi Chen Zhenhui Wang

Catalpol, one of the primary bioactive components in Rehmannia glutinosa, is an iridoid glycoside with significant pharmacological activities. To expand the microbial sources of catalpol, endophytic bacteria were isolated from R. glutinosa (cultivated in Jiaozuo, China) using the dilution plating method combined with vanillin–sulfuric acid colorimetric assay. High-performance liquid chromatography (HPLC) and liquid chromatography–mass spectrometry (LC-MS) were employed for screening and identification. The isolated strain was identified through morphological characterization and 16S rDNA gene sequence analysis, while single-factor experiments coupled with response surface methodology were utilized to optimize its fermentation conditions. Results indicated that the strain DH14 formed circular, cream-white, opaque colonies and was Gram-negative. It was identified as Brevundimonas olei. The optimal fermentation conditions were determined to be 190 rpm, pH 7.6, 31 °C, and 0% NaCl. Meanwhile, the results revealed a positive correlation between the pH of the fermentation broth and catalpol production. Under the optimized conditions, the maximum catalpol yield reached 0.142 mg/mL after 3 days of cultivation. This study provides a promising microbial resource and optimized fermentation parameters for the microbial production of catalpol.

Microbiology Research doi: 10.3390/microbiolres17030059

Authors: Kaloyan D. Georgiev Deyan Dzhenkov Marieta Georgieva Antoaneta Tsvetkova

Probiotics are live microorganisms that, when administered in adequate amounts, confer health benefits to the host. The present study aimed to evaluate the protective effects of a probiotic against experimentally induced gastric ulcers in rats. Male Wistar rats were allocated into five groups (n = 6 per group): Control, Model (indomethacin-induced ulcers, IND), Positive control (IND + famotidine), IND + Laktera Nature (LN) probiotic (800 mg/kg), and IND + LN probiotic (1600 mg/kg). The study duration was 14 days. On day 15, rats in groups 2–5 received 1 mL of indomethacin (40 mg/kg) orally. All rats were euthanized four hours later via cervical dislocation under diethyl ether anesthesia. LN probiotics have demonstrated dose-dependent protection against indomethacin-induced gastric ulcers, as assessed by both macroscopic and microscopic evaluation. Therefore, LN probiotics may represent a valuable adjunctive strategy in the management of gastric ulcers.

Microbiology Research doi: 10.3390/microbiolres17030058

Authors: Christopher T. Piatnichouk Joshua V. Ruppel Nicole L. Snyder

Tuberculosis, caused by Mycobacterium tuberculosis, remains a global health challenge, particularly due to multidrug-resistant strains. Photodynamic therapy using porphyrin-based photosensitizers offers a promising alternative by targeting the trehalose-rich cell wall of the bacillus. Motivated by prior experimental observations that shorter linkers improve efficacy, this study probes the molecular basis of linker-length-dependent activity in trehalose–porphyrin glycoconjugates. Here, we show that shorter linker lengths are consistent with improved activity in vitro and, in an Ag85B docking model, constrain conformational flexibility, reduce solvent exposure, and promote tighter packing consistent with stronger predicted interactions. Using computational docking, we analyzed binding scores, RMSD variability, steric clashes, and protein–ligand interactions for conjugates docked into Ag85B, a key enzyme in cell wall synthesis. Shorter linkers (0–2 carbons) were found to exhibit superior binding scores, lower RMSD variability, and stronger interactions with residues such as ARG 43, including unique π–cation interactions. In contrast, longer linkers displayed increased flexibility, reduced binding specificity, and greater solvent exposure. These findings, which support our experimental observations, suggest a molecular basis for linker-dependent efficacy and provide a framework for designing next-generation porphyrin-based therapeutics for tuberculosis treatment.

Microbiology Research doi: 10.3390/microbiolres17030057

Authors: Valery M. Dembitsky Alexander O. Terent’ev Sergey V. Baranin

Siderophores are classically understood as microbial iron-acquisition metabolites: low-molecular-weight ligands secreted by bacteria to solubilize and transport Fe(III) under iron-limited conditions. In this review, we expand that paradigm by highlighting an emerging and underappreciated chemical axis—boron coordination by siderophores—that links terrestrial (soil/rhizosphere) and marine microbiomes. Across diverse bacterial taxa, siderophore production is widespread and central to competitive fitness because Fe(III) is poorly soluble and frequently sequestered in environmental or host matrices. Yet in boron-rich settings (seawater and borate-enriched soils), the same oxygen-donor architectures that support Fe(III) chelation can also engage boron chemistry. We synthesize evidence that carboxylate/α-hydroxyacid (dicitrate-type) and catecholate siderophores can form tetrahedral borate/boronate complexes, whereas hydroxamate siderophores generally lack the vicinal dianionic O,O motif required for stable boron binding. Structurally characterized examples—including vibrioferrin, rhizoferrin, and petrobactin—demonstrate that boron complexation is experimentally observable by ESI-MS and multinuclear NMR and can be modulated by pH and microenvironment. Integrating these findings with datasets on boron-tolerant bacteria, we propose that when iron is scarce and boron is available, boron–siderophore complexation becomes chemically feasible and may influence microbial physiology by altering ligand conformation, metal selectivity, and potentially extracellular signaling behavior—especially in marine systems where borate is abundant at oceanic pH. Overall, this review frames boron-binding siderophores as a cross-ecosystem phenomenon and a promising conceptual bridge between environmental boron geochemistry, microbial metal economy, and metalloid-mediated signaling.

Microbiology Research doi: 10.3390/microbiolres17030056

Authors: Gonçalo Melo Raquel Abreu Gonçalo Pereira Eva Cunha Lélia Chambel Luís Tavares Manuela Oliveira

Semi-aquatic turtles are popular companion animals and represent an important One Health interface in Portugal due to their potential to harbour zoonotic pathogens. This study aimed to characterize the virulence and antimicrobial resistance (AMR) profiles of Aeromonas spp. isolated from captive semi-aquatic turtles in Portugal. Cloacal swabs (n = 31) were collected from turtles under human care, and Aeromonas spp. were isolated using selective media and identified by multiplex PCR. Antimicrobial susceptibility was assessed by disk diffusion using eleven antibiotics, while the phenotypic virulence profile was evaluated by determining isolates’ ability to express five hydrolytic enzymes and to form biofilm. A total of 86 Aeromonas isolates were recovered, with A. hydrophila being the most prevalent (77.9%). Most isolates displayed high pathogenic potential: over 87% produced DNase, haemolysin, and lecithinase, while nearly all produced protease and gelatinase. Also, 44.2% of the isolates were resistant to at least one antibiotic, and 12 (14.0%) were multidrug resistant. Higher Multiple Antibiotic Resistance (MAR) indices were significantly associated with turtles housed in indoor aquariums (p < 0.05). These findings indicate that captive semi-aquatic turtles may act as reservoirs of virulent and antimicrobial-resistant Aeromonas spp., highlighting potential zoonotic risks and supporting the need for a One Health approach in their management.

Microbiology Research doi: 10.3390/microbiolres17030055

Authors: Zeyu Ai Ping Dai Kangnan He Ruilong Nie Shimin Zou Liang Chen

In recent years, the gut microbiome has been increasingly recognized as an important factor in regulating treatment responses and disease progression in prostate cancer (PCa). We synthesized literature published over the past five years, focusing on preclinical and clinical studies linking the microbiome to PCa treatment outcomes. There is accumulating evidence that gut microbiota dysbiosis and its associated metabolites can modulate key biological processes, such as androgen metabolism, inflammatory signaling pathways, and antitumor immune responses. These processes affect the sensitivity of PCa patients to androgen deprivation therapy (ADT) and other systemic treatments. The available evidence suggests that the gut microbiome has the potential to serve as a predictive biomarker for treatment response and could represent a novel target for interventional precision therapy in PCa. This narrative review summarizes the latest research on the “gut–prostate axis”, focusing on the role of the gut microbiome in regulating therapeutic responses in PCa and the underlying mechanisms. Finally, we address current limitations, including the predominance of preclinical evidence, methodological heterogeneity, and the critical need for longitudinal clinical validation to distinguish causality from association.

Microbiology Research doi: 10.3390/microbiolres17030054

Authors: Manuela Reyes-Estebanez Pedro Mendoza-de Gives

In the original publication [...]

Microbiology Research doi: 10.3390/microbiolres17030053

Authors: Yadi Liu Qiuyue Hu He Li

Colletotrichum fructicola is the predominant pathogenic agent responsible for anthracnose in Camellia oleifera. RGS2 is a GTPase-activating protein that negatively regulates G-protein signaling by inactivating Gα subunits. In this study, we characterized the ortholog of CfRGS2 in C. fructicola to explore its pathogenic roles. Seven canonical RGS genes were identified through BLASTp and keyword searches. Conserved domains and subcellular localizations were predicted bioinformatically. A CfRGS2 knockout mutant was generated via overlap-PCR and PEG-mediated transformation, verified by PCR, and complemented by reintroducing the wild-type gene. Phenotypic characterization showed that the growth rates of mutants ΔCfrgs2-1 and ΔCfrgs2-2 were significantly reduced compared with those of the wild-type and complemented strains. On both PDA and minimal medium, the mutant strains exhibited significantly smaller colony diameters of 3.3 cm and 3.1 cm, respectively, relative to the control strains. Moreover, conidiation in the mutants was only 4% of that in the wild-type and complemented strains, and appressorium formation was reduced to 6%, with statistical analyses confirming high significance. Under cell wall stress induced by 400 μg/mL Congo red, the growth inhibition rates of ΔCfrgs2-1 and ΔCfrgs2-2 were 44% and 48%, respectively, significantly higher than those of the control strains. Pathogenicity assays demonstrated that the mutants failed to induce lesions on unwounded leaves and caused 47% and 30% smaller lesion areas on wounded apple fruits, respectively. In summary, C. fructicola possesses seven canonical RGS proteins that regulate G-protein signaling, among which CfRgs2 is implicated in growth, conidiation, the stress response to cell wall perturbation, and virulence.

Microbiology Research doi: 10.3390/microbiolres17030052

Authors: Doris Evelyn Yah Hui Jong Siang Yin Lee Yun Khoon Liew Phyu Synn Oo Amar Harris Arifin Zi Ni Ngai Beek Yoke Chin Shamala Salvamani Rhun Yian Koh

Quebrachitol, an optically active cyclitol derived from plants, has recently gained attention as a potential natural product with therapeutic properties, though its antimicrobial effects remain unclear. This systematic review aims to determine, appraise, and consolidate evidence of the antimicrobial potential of quebrachitol. PRISMA-guided searches of PubMed, Scopus, and Google Scholar (2000–2024) identified English-language experimental in vitro, in vivo, and in ovo studies. Data on antimicrobial activity, dosage or treatment duration, and mechanisms were extracted, with study quality assessed using QUIN and SYRCLE tools. Of 866 studies screened, 11 met inclusion criteria: seven in vitro, one in vivo, one in ovo, and two combining both approaches. Quebrachitol demonstrated inhibitory effects against Salmonella sp., Candida albicans, infectious bursal disease virus (Avibirnavirus gumboroense), Newcastle disease virus, Plasmodium sp., and notably, biofilm formation by Staphylococcus epidermidis and methicillin-resistant Staphylococcus aureus (MRSA). Overall, quebrachitol exhibits promising antimicrobial potential, but rigorous in vivo studies are required to confirm its efficacy and safety in addressing antimicrobial resistance.

Microbiology Research doi: 10.3390/microbiolres17030051

Authors: Alexandra Gheorghiță Anca Pleșa Bianca Pop Vlad Stoian Roxana Vidican

Long-term soil fertility is governed by the metabolic plasticity of microbial communities, particularly during the decomposition of crop residues. This study investigated the straw-associated functional microbial profile associated with straw decomposition under the influence of 62 years of continuous management with mineral fertilization and liming. Using the Biolog EcoPlateTM approach combined with a modified litter-bag protocol, we assessed shifts in metabolic activity patterns of functional guilds and groups. PERMANOVA results revealed that the interaction between liming and fertilization (p < 0.05) was the primary driver of divergence in functional communities, rather than the individual effect of factors. Long-term treatments induced a significant reconfiguration of the functional niche, shifting from the native, generalist microbiome to specialized communities in treated variants, with carbohydrate (CH) guilds as dominant and indicators of community performance. Moderate levels of liming (L1) stimulated metabolic activity and maintained higher functional diversity across amino acid (AA) and polymers (Px) guilds. Intensive liming (L2), in contrast, restricted the activity of most microbial functional groups and favored amine (AM) and carboxylic acid (CX) guilds. Shifts from a generalist microbiome in native soil to specialized communities in treated soils show the capacity of microorganisms to adapt efficiently under agronomic management.

Microbiology Research doi: 10.3390/microbiolres17030050

Authors: Ana K. Rocha-Viggiano Saray Aranda-Romo Edgar R. Rocha-Lara Karla G. López-Macías Sergio Casas-Flores Nicolás Gómez-Hernández Daniel E. Noyola Cesaré Ovando-Vázquez Mariana Salgado-Bustamante

Pregnant women undergo a myriad of physiological changes, including important hormonal variations. Pregnancy gingivitis is a condition that affects up to 30% to 100% of women, is related to hormonal modifications, and could play an important role in gestational gut colonization and immunological training of the newborn. Nonetheless, oral health is not always included in routine prenatal care. In this study, we collected saliva samples of pregnant women with and without pregnancy gingivitis and analyzed the oral microbiota through 16S sequencing. In addition, meconium samples from the infants of participating women were analyzed. The oral microbiota of pregnant women with and without pregnancy gingivitis did not show significant differences in diversity. However, significant differences in microbiome composition were observed. Pathway analysis showed that, despite taxonomic similarity, the PG group had activated energy metabolism, bacterial growth, lipid metabolism, and virulence pathways with NOD-like receptor activation, indicating pro-inflammatory microbial activity. In contrast, the NPG group exhibited central metabolism and repair mechanisms, suggesting that PG could affect microbiome function rather than composition. In addition, it appears that the microbiome composition of offspring of mothers with gingivitis also differs from that of offspring from mothers without gingivitis, although the number of available samples did not allow for definite conclusions. As such, a larger cohort and deeper sequencing methods are needed to assess the oral microbiota of pregnant women with and without gingivitis and to explore the possibility of bacterial translocation from the maternal gingiva to the fetal gut.

Microbiology Research doi: 10.3390/microbiolres17030049

Authors: Hemshikha Rajpurohit Mark A. Eiteman

Phosphofructokinase 1 (PfkA) mediates the ATP-dependent phosphorylation of fructose-6-phosphate and is a key, controlling enzyme in glycolysis for Escherichia coli and other organisms. In this study, 22 chromosomally expressed PfkA variants were constructed in E. coli C. These variants, the wild-type strain, and the ∆pfkA strain were compared for growth rates using glucose as the sole carbon source. The majority of variants (14 of 22) attained a growth rate less than 20% of the growth rate of the wild-type strain (0.94 h−1) and thus similar to the knockout strain (0.12 h−1). Three variants (R171S, F76Y, and R77A), representing a range of growth phenotypes, and strains expressing the wild-type PfkA and the ∆pfkA deletion strain were additionally examined for key intracellular metabolites and gene expression under nitrogen-limited steady-state conditions. These five strains could be distinguished by two groupings: strains with relatively high growth rates under batch conditions (wild-type and R77A variant) showed the greatest glucose consumption rate and formed acetate, whereas strains with low growth rates (F76Y, R77A, and ∆pfkA) exhibited low glucose consumption and did not accumulate acetate. As the PfkA mutation severity increased, the intracellular concentrations of acetyl-CoA and fructose-1,6-bisphosphate and the sum of dihydroxyacetone and glyceraldehyde-3-phosphate greatly decreased. Although the mutation severity had a limited effect on the expression of maeB and icd genes expressing malic enzyme and isocitrate dehydrogenase, it correlated with reduced expression of zwf and pta genes expressing glucose-6P-dehydrogenase and phosphotransacetylase, respectively. The results highlight the great sensitivity of the enzyme to substitutions and the key role it plays in controlling glycolytic flux.

Microbiology Research doi: 10.3390/microbiolres17030048

Authors: Xiaoran Chu Kaili Wang Vincenzo Cuteri Cheng Liu Yubao Li Zhenshu Si

Fowl adenovirus serotype 4 (FAdV-4) is the major causative agent of hydropericardium-hepatitis syndrome (HHS), a disease responsible for considerable economic losses in poultry production. Although inactivated and live-attenuated vaccines reduce mortality, continued outbreaks highlight the need to optimize vaccination strategies. To address these limitations, we developed and evaluated a bivalent subunit vaccine composed of recombinant hexon-L1 and fiber-2 proteins, two major antigenic determinants associated with neutralization and pathogenicity. The proteins were expressed in Escherichia coli, purified under native conditions, confirmed for purity and antigenicity, and emulsified into a water-in-oil formulation. Chickens were immunized with either 10 μg or 20 μg doses, boosted after 14 days, and challenged with the homologous virulent FAdV-4 strain SDLC202009. The 20 μg dose conferred complete survival, eliminated histopathological lesions, prevented viral detection in tissues by PCR and immunohistochemistry, and fully blocked viral shedding. Similarly, the 10 μg dose induced a good protection with only minor pathological differences compared to the group treated with 20 μg. These results demonstrate that a bivalent hexon-L1 and fiber-2 subunit formulation elicits strong, dose-dependent humoral and tissue-level protection against homologous FAdV-4 challenge under the conditions tested. The experimental design did not include a monovalent fiber-2 comparator; therefore, conclusions regarding the relative contribution of each antigen are not drawn.

Microbiology Research doi: 10.3390/microbiolres17030047

Authors: Alisa Nakkaew Kotchakorn Praopring

Boletus griseipurpureus is an ectomycorrhizal mushroom characterized by a bitter flavor. In this study, specimens were collected from three host plants—Acacia auriculiformis (BgAa), Melaleuca cajuputi (BgMc), and Eucalyptus camaldulensis (BgEc)—and initially classified based on pileus morphology. Molecular biodiversity was investigated using internal transcribed spacer (ITS) DNA barcoding, and comprehensive phylogenetic analysis revealed that B. griseipurpureus populations in southern Thailand clustered according to their symbiotic host species. De novo transcriptome assembly of B. griseipurpureus associated with different hosts was performed to generate unigene datasets, followed by functional gene annotation. A total of 1157 differentially expressed genes (DEGs) were identified and linked to ectomycorrhizal symbiosis. The genes involved in biosynthesis and metabolic processes exhibited host-dependent expression patterns. Furthermore, expression profiles of five selected genes—major facilitator superfamily (MFS) substrate transporter, phosphatase II, hexose transporter, terpenoid synthase, and fungal hydrophobin—were consistent between DEG analysis and semi-quantitative RT-PCR validation. These findings suggest that these genes play important roles in ectomycorrhizal symbiosis and the biosynthesis of bioactive compounds in B. griseipurpureus, with expression influenced by host association. This study provides valuable insights into the molecular biodiversity and gene regulation underlying ectomycorrhizal symbiosis, contributing to a better understanding of the biological processes in B. griseipurpureus.

Microbiology Research doi: 10.3390/microbiolres17030046

Authors: Harshita Naithani Amélia Camarinha-Silva Thilo M. Fuchs Jana Seifert

Myo-inositol (MI) is an essential dietary polyol involved in host metabolism. However, the identity and diversity of MI-utilizing gut bacteria in poultry remain largely unknown. This study aimed to isolate anaerobically growing bacteria enriched under MI-based cultivation conditions from two commercial laying hen breeds. Digesta samples were cultured on minimal growth media containing MI as the sole (Trial 1) or principal (Trial 2) carbon source. Isolates were purified, screened by amplified ribosomal DNA restriction analysis (ARDRA), and identified using 16S rRNA gene sequencing. Among 42 sequenced isolates, ten representative strains were classified within the phyla Pseudomonadota (n = 5), Bacillota (n = 4), and Bacteroidota (n = 1). Members of the Escherichia-Shigella clade were most frequently recovered, followed by Clostridium, Enterococcus, Pediococcus, and Bacteroides. Selected Escherichia-Shigella isolates were screened negative for ipah and ial virulence genes, except for three isolates that tested positive only for the ipah gene. These findings provide the first culture-based framework for investigating MI-responsive bacteria in the chicken gut.

Microbiology Research doi: 10.3390/microbiolres17020045

Authors: Michael De Benedetto Grazia Cecchi Laura Cutroneo Simone Di Piazza Vincent Bertin Marco Capello Mirca Zotti

The deep sea is characterized by unique and extreme habitats. The absence of light, high salinity, hydrostatic pressure, low temperature, and high competition led to the evolution of physiological and biochemical adaptations necessary for survival. Marine fungi represent a significant part of deep-sea microbial communities. Studying bathypelagic sediment fungi helps us to understand their little-known communities and ecology, as well as their metabolic potential and ecophysiological properties, which have applications in pharmaceutical biotechnologies and bioremediation protocols. During an oceanographic campaign off the coast of Toulon (France, northwest Mediterranean Sea) in October 2021, as part of the KM3NeT Project, the Hybrid Remotely Operated Vehicle (HROV) Ariane collected a composite sediment sample at a depth of 2417 m. The sediment was physically, geochemically and mycologically characterized. Culturable fungi were isolated, and vital fungal strains were identified morphologically and molecularly. A total of 17 strains were isolated and identified in pure culture. The major taxa belonged to the Penicillium, Aspergillus, and Cladosporium genera, but widespread species such as Wallemia sebi were also found. This study also paves the way for further research into the advantages and disadvantages of using HROV technology for mycological cultural investigations at prohibitive depths.

Microbiology Research doi: 10.3390/microbiolres17020044

Authors: Hideki Kakeya Yoshihisa Matsumoto

Background: The spike D614G substitution became globally dominant early in the COVID-19 pandemic, and reversion to ancestral D614 is expected to be rare once D614G is fixed. SARS-CoV-2 sequences lacking D614G detected later raise questions about the origin of these reversions. Methods: We analyzed spike protein amino-acid sequences from 22 SARS-CoV-2 Variants of Concern (VOCs) deposited in the NCBI GenBank database, screening for sequences carrying ancestral D614 and comparing their distributions across VOCs. Results: D614 reversions (reverse mutations of D614G) were not evenly distributed across VOCs but were strongly enriched in Delta (B.1.617.2) and Omicron BA.2, reaching levels statistically inconsistent with other VOCs. In both lineages, D614-containing sequences showed limited mutational diversity and pronounced geographic clustering within specific U.S. regions. Conclusions: These non-random patterns are difficult to reconcile with spontaneous reverse mutation arising and spreading through typical community transmission and are more consistent with localized reintroduction of an older genetic background. Further investigation is warranted to assess whether laboratory-associated events could be involved.

Microbiology Research doi: 10.3390/microbiolres17020043

Authors: Ming-Yue Zhang Xiao-Hui Zhang Xue-Ying Wang Jun-Hui An Dong-Hui Wang Rong Hou Yu-Liang Liu

A growing body of evidence indicates that the gut microbiota has a role in the mating preference process in mammals. This likely occurs through the modulation of various mating signals induced by symbiotic bacteria, thereby leading to variations in mating behavior. Given that giant pandas are solitary wild animals that rely on chemical signals for mate selection, it is relevant to explore whether the mating behavior of giant pandas is also affected by the gut microbiota. We hypothesize that antibiotic treatment-induced residual antibiotic resistance genes in captive giant pandas may disrupt intestinal microbiota homeostasis, diminish the abundance of beneficial microorganisms involved in short-chain fatty acid synthesis, and consequently impair nervous system function via the gut–brain axis. The ensuing physiological stress is likely to suppress innate mating behavior and compromise pheromone synthesis, thereby reducing an individual’s attractiveness to potential mates. To answer this question, we utilize fecal metagenomics technology to analyze the differences in gut microbes and antibiotic resistance genes (ARGs) between captive male adult giant pandas displaying natural versus non-natural mating behavior. The research findings suggest that, when compared with captive adult male giant pandas demonstrating natural mating behavior, those with non-natural mating behavior exhibit a significantly reduction in the abundance of beneficial gut microorganisms (s_Clostridium sp. and f_Ruminococcaceae) (p < 0.05). Concurrently, there is a significantly increase in the observed resistance genes tetO and mgtA, which are mainly associated with macrolide and tetracycline resistance (p < 0.05). Furthermore, Kegg functional analysis reveals a significant up-regulation of metabolic pathways related to sensory systems, such as taste and olfactory transduction, in the intestines of captive adult male giant pandas showing natural mating behavior. These results imply that changes in the abundance of gut microbiota and ARGs are correlated with the manifestation of natural mating behavior in captive adult male giant pandas. Consequently, to improve the success rate of natural reproduction within the male giant panda populations in captive environments, it is advisable to administer antibiotics judiciously and closely monitor the composition of beneficial bacteria in their gut microbiota. The findings of this study provide novel perspectives on the mechanisms by which captive conditions affect the decline in natural mating behavior observed in adult male giant pandas.

Microbiology Research doi: 10.3390/microbiolres17020042

Authors: Nande Ndamase Lindiwe Modest Faye Ntandazo Dlatu Teke Apalata Mojisola Clara Hosu

Background: Tuberculosis (TB) outcomes remain suboptimal in high-burden, resource-constrained settings. Clinical and socio-economic factors contribute to loss to follow-up, failure, and mortality, yet their relative importance remains underexplored. Methods: We analyzed a retrospective cohort of patients treated for pulmonary TB in the Eastern Cape, South Africa. Treatment outcomes were dichotomized as success (cured or treatment completed) versus unsuccessful (loss to follow-up, failure, or death), excluding transfers and patients still on treatment. Predictors included age, gender, income, occupation, comorbidities, HIV status, previous treatment history, patient category, and drug resistance status. Regularized logistic regression was used to estimate odds ratios, while the best decision tree model was applied to identify hierarchical risk profiles. Results: Logistic regression demonstrated high accuracy (86%) and identified drug susceptibility, age, income stability, and comorbidity burden as the strongest predictors of treatment success. The decision tree achieved lower accuracy (65%) but improved detection of unsuccessful outcomes, highlighting a clear hierarchy of risk: (1) drug resistance status, (2) age, (3) income source, and (4) comorbidities. Patients with drug-resistant TB, older age, no income or reliance on grants, and coexisting conditions were at the highest risk of poor outcomes. Conclusions: Drug resistance, age, income, and comorbidity burden shape a hierarchical risk profile for TB treatment outcomes in rural South Africa. Logistic regression offered robust overall classification, while the decision tree provided transparent stratification of at-risk groups. These findings underscore the need for integrated clinical and socio-economic support strategies to improve outcomes in high-burden settings.

Microbiology Research doi: 10.3390/microbiolres17020041

Authors: Sebastiano Rosati Martina Cofelice Nicolaia Iaffaldano Francesco Lopez Giuseppe Cinelli Alessandra Mazzeo Emanuele Antenucci Lucia Maiuro Elena Sorrentino

Fish are nutritionally valuable foods but are also highly perishable, representing a major research focus for the development of effective preservation strategies to delay spoilage while maintaining microbiological acceptability. In this context, edible coatings have gained increasing attention as clean-label tools to extend the shelf life of perishable foods. In this study, an alginate-based emulsion containing oregano essential oil (OEO) was applied to commercial frozen–thawed ready-to-cook (RTC) hake fillets, intentionally selected as a reproducible model system to evaluate coating performance under refrigerated conditions. Coated and uncoated fillets, stored at 4 °C for up to 7 days, were monitored over time for microbiological and physicochemical parameters, including microbial loads, pH, weight loss, and lipid oxidation (TBARS). Compared to uncoated samples, fillets treated with the alginate–OEO emulsion exhibited a marked delay in spoilage-related microbial growth, with a consistent inhibition of Pseudomonas spp. throughout the experimental period, while maintaining microbiological acceptability. Emulsion-based coated fillets also exhibited reduced lipid oxidation, a more uniform surface appearance with only minor visible color changes, and the absence of unpleasant off odors during the refrigerated storage. Overall, the results demonstrate that the alginate–OEO coating could represent an effective strategy for improving the microbiological and oxidative stability of RTC fish fillets under refrigerated conditions, with potential implications for extended shelf life and a reduction in food waste.

Microbiology Research doi: 10.3390/microbiolres17020040

Authors: Milena Branković Miroslava Jordović Pavlović Ana Tomic Natasa Opavski Ina Gajić

Antimicrobial resistance (AMR) is accelerating and driven by antimicrobial use. Hospital consumption and resistance data remain scarce in Serbia. This retrospective study assessed the prevalence of infectious agents among hospitalized patients (2019–2022), distribution and AMR rates of Escherichia coli and Klebsiella pneumoniae, antimicrobial consumption, and associations between AMR and antibiotic use (Spearman’s correlation). All bacterial isolates were included for species-frequency analyses. For AMR and specimen distribution, repeat isolates of E. coli and K. pneumoniae per patient were excluded. Among 10,780 isolates, E. coli remained most frequent (24.7% in 2019; 24.3% in 2022), whereas K. pneumoniae increased from 7.7% to 16.9% (p < 0.001). E. coli resistance to cephalosporins, fluoroquinolones, and piperacillin/tazobactam increased significantly from 2019 to 2022 (p < 0.05). K. pneumoniae resistance increased to most agents (p < 0.001), with meropenem resistance rising from 11.7% to 52.0%. Multidrug resistance rose from 27.3% to 35.4% for E. coli (2020–2022; p = 0.006) and from 62.4% to 82.9% for K. pneumoniae (2019–2022; p < 0.001). Total hospital antibiotic consumption nearly doubled from 57 to 111.8 Defined Daily Doses per 100 bed days (2019–2021) and decreased to 52.0 (2022). Temporal concurrence of increased antimicrobial use during COVID-19 and escalating resistance underscores the need for strengthened surveillance and stewardship.

Microbiology Research doi: 10.3390/microbiolres17020039

Authors: Alexey A. Ananev Olga A. Aleynova Nikolay N. Nityagovsky Alina A. Dneprovskaya Alexandra S. Dubrovina Haiping Xin Konstantin V. Kiselev

The search for antagonistic microorganisms as alternatives to chemical pesticides is an urgent priority in sustainable agriculture. Previously, we isolated several bacterial isolates from spruce plants, and one of them, identified as Bacillus atrophaeus R7PjV2-12, showed strong antagonistic properties against plant pathogens such as Magnaporthe oryzae, Fusarium avenaceum, and Erwinia billingiae. Given its strong fungicidal properties, we decided to sequence the complete genome of this bacterium to determine how it can inhibit fungal growth. The whole genome size of B. atrophaeus R7PjV2-12 was 4,127,644 bp with 4032 open reading frames. B. atrophaeus R7PjV2-12 genome possessed clusters of secondary metabolites with a complete set of genes with 100% similarity representing clusters of biosynthesis of bacillin, bacillibactin, subtilosin A, and fungicin, which indicates the studied strain’s ability to synthesize these substances. Thus, this paper has shown and discussed the potential importance of B. atrophaeus R7PjV2-12 for biocontrol of pathogenic microorganisms in agriculture.

Microbiology Research doi: 10.3390/microbiolres17020038

Authors: Tory M. Johnson Quanbo Xiong Zhengxin Wang

Thuja arborvitae are widely grown in North America and East Asia for their ornamental value, and their leaf oil extracts have been used to treat bacterial infections. This study aimed to identify antibacterial compounds from Thuja leaves. The methanol extract of Thuja leaves exhibited strong antibacterial activity against Gram-positive (Staphylococcus aureus and Streptococcus mutans) and Gram-negative (Acinetobacter baumannii, Pseudomonas aeruginosa) bacteria. The major compounds in the active fractions were isolated and identified as apigenin-7-di-p-coumarylglucoside and eicosapentaenoic acid. The identified compounds showed potent antibacterial activity against the four tested microorganisms with IC50 values of 10 to 50 µg/mL. More importantly, these compounds showed potent inhibitory activity (IC50: 10 μg/mL) against the multidrug-resistant bacterial strain Acinetobacter baumannii. Two antibacterial compounds are now being reported for the first time in Thuja arborvitae, and they may have potential for the treatment of bacterial infections.

Microbiology Research doi: 10.3390/microbiolres17020037

Authors: Kyung-A Hyun Ji-Hyun Kim Min Nyeong Ko Chang-Gu Hyun

Bombiscardovia sp. JNUCC 75 (=CH12) was isolated from the flowers of Prunus yedoensis on Jeju Island, representing one of the few known flower-associated members of the Bombiscardovia asteroides group. Whole-genome sequencing revealed a compact 2.28 Mb genome and a functional gene profile enriched in translation, amino-acid metabolism, and DNA repair. Although the strain contains a pseudogene content comparable to other B. asteroides-group members, the overall genomic architecture—together with the presence of stress-response and polyprenyl/terpenoid biosynthetic pathways—suggests adaptation to oxygen-variable, phenolic-rich, and UV-exposed floral environments. Comparative genomic analyses (OrthoANIu 97.16%; dDDH 72.7%) demonstrated that JNUCC 75 is closely related to B. polysaccharolytica yet forms a genetically distinct lineage within the group. Genome mining uncovered two previously unreported terpenoid/polyprenyl biosynthetic gene clusters, indicating a novel isoprenoid-derived metabolic repertoire with potential roles in membrane stabilization and oxidative-stress defense. These genomic features collectively position JNUCC 75 as a bridge between gut-associated and environmental bifidobacteria and highlight its potential as a promising microbial resource for postbiotic, antioxidant, and skin-barrier-enhancing applications. This study expands the ecological range of bifidobacteria and provides a genomic framework for evaluating flower-derived Bombiscardovia strains in cosmeceutical and functional food innovation.

Microbiology Research doi: 10.3390/microbiolres17020036

Authors: Ahmed Najm Abed Maroua Gdoura Ben Amor Radhouane Gdoura

Antimicrobial resistance among multidrug-resistant enteric pathogens, such as Salmonella enterica and Shigella flexneri, poses a critical global health challenge, necessitating novel therapeutic strategies from natural sources. This study evaluated the preliminary phytochemical profile and dual antibacterial and antibiofilm activities of methanolic leaf extract from Platanus orientalis Linn. Qualitative and quantitative phytochemical analyses were conducted to assess phenolic and flavonoid contents, while antibacterial activity against multidrug-resistant isolates was determined using broth microdilution to obtain minimum inhibitory concentrations (MICs). Antibiofilm effects were examined at sub-MIC concentrations via the crystal violet assay. The extract was found to be rich in phenolics and flavonoids and exhibited concentration-dependent antibacterial activity, with MICs of 64 µg/mL for S. enterica and 32 µg/mL for S. flexneri. Sub-MIC concentrations significantly reduced biofilm biomass by 60–80%, likely through interference with quorum sensing and extracellular polymeric substance production. These results highlight the extract’s potential as a phytotherapeutic agent targeting both planktonic cells and biofilms, providing insights for the development of natural antimicrobials against resistant enteric pathogens. Overall, the findings underscore the potential of Platanus orientalis methanolic leaf extract as a promising natural antibacterial and antibiofilm agent against multidrug-resistant enteric pathogens and support its further development as an alternative or complementary therapeutic strategy. It should be noted that the phytochemical results are qualitative and indicative, and further LC–MS or NMR analyses are required for definitive compound identification.

Microbiology Research doi: 10.3390/microbiolres17020035

Authors: Jingpeng Zhang Jinxiu Jiang Dingding Zhang Yusheng Lin Chunhe Wan Yongliang Che

Caseous lymphadenitis (CLA), an infectious disease caused by Corynebacterium pseudotuberculosis (C. pseudotuberculosis), poses a significant economic burden to the global small ruminant industry. This study aimed to investigate genetic variations in the CP40 gene of C. pseudotuberculosis and to develop a rapid detection assay for enhanced pathogen identification. Homology analysis was performed to compare the CP40 gene sequence of the FJ-PN strain with other Corynebacterium species. Specific primers targeting CP40 were designed, and a SYBR Green I-based real-time PCR protocol was optimized. The assay’s specificity, sensitivity, and reproducibility were subsequently validated. The FJ-PN strain exhibited ≥99.65% nucleotide identity and ≥98.94% amino acid identity with C. pseudotuberculosis biovar ovi reference strains, showing 90.18–91.84% nucleotide identity and 88.63–90.77% amino acid identity with C. pseudotuberculosis biovar equi, and ≤82.71% nucleotide identity and ≤78.63% amino acid identity with other Corynebacterium species. The established qPCR assay demonstrated high specificity, the limit of detection was 52 copies/μL, and it demonstrated good reproducibility (intra- and inter-assay CV < 1.0%). Clinical sample testing revealed 18.8% positivity rates in nasal swabs, which was higher than that detected by conventional PCR (16.3%). These results indicate that the CP40 gene is evolutionarily conserved and represents a specific molecular marker for the identification of C. pseudotuberculosis biovar ovis. The developed SYBR Green I real-time PCR assay enables the efficient detection of C. pseudotuberculosis and provides technical support for CLA surveillance and control.

Microbiology Research doi: 10.3390/microbiolres17020034

Authors: Yuan Gao Linli Ji Taya Tang Jiadi Zhu Shuyang Yu Junjie Niu Heng Li

The global rise of multidrug resistance (MDR) across bacterial pathogens poses a severe threat to public health, with the food chain serving as a critical reservoir and transmission route for resistant clones. This study investigated the genomic epidemiology of Staphylococcus aureus in retail pork from Beijing, China, and Copenhagen, Denmark, with a focus on MDR patterns and associated genetic elements. Among 134 isolates, the livestock-associated clonal complex CC398 was the dominant lineage (24.63%) and exhibited a high burden of MDR (48.48%), carrying resistance genes to β-lactams (blaZ and mecA), tetracyclines (tetM and tetK), and aminoglycosides. Notably, MRSA isolates displayed a significantly higher MDR prevalence (73.53%) compared to MSSA isolates (18.00%), underscoring methicillin resistance as a key marker for broader resistance phenotypes. Phylogenetic analysis revealed the segregation of CC398 into distinct sub-lineages, with the livestock-associated branch consistently linked to a characteristic tetracycline–β-lactam MDR profile. Furthermore, high frequencies of mobile genetic elements, such as the rep16 plasmid, were associated with MDR dissemination in CC398. These findings highlight retail meat as an important reservoir for MDR S. aureus and illustrate how livestock-adapted clones contribute to the environmental burden of antimicrobial resistance. This study underscores the need for integrated One Health surveillance that connects veterinary, food safety, and human health sectors to monitor and contain the spread of MDR bacteria across ecological niches.

Microbiology Research doi: 10.3390/microbiolres17020033

Authors: Monika Gothwal Flurina Suter Lamprini Karygianni Thomas Attin Thomas Thurnheer

The highly destructive pathogenic processes in patients with periodontitis are attributed to the presence of subgingival biofilms comprising key periodontal pathogens, such as Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia, as well as other periodontopathogens including Fusobacterium nucleatum, Selenomonas spp., Centipeda spp., and Campylobacter spp. Considering the vast microbial diversity in periodontitis, we aimed to analyze the presence of various bacterial species in subgingival dental plaque, with a special focus on Selenomonas spp. We first developed a phylogenetic tree for Selenomonas–Veillonella clusters, using in silico analysis followed by fluorescence in situ hybridization (FISH) on subgingival dental plaque samples from 22 patients with a history of chronic periodontitis, by using specific 16S rRNA oligonucleotide probes. These oligonucleotide probes’ specificity and hybridization conditions were determined on previously characterized bacterial strains. Qualitative and quantitative analysis of FISH slides was carried out by using an epifluorescence microscope. The majority of the patient samples showed high fluorescence signals with the oligonucleotide probes SEL1150, Sspu439, and SEL1469 (specific for Selenomonas spp.), ACI623 (identifying Selenomonas, Veillonella, and Dialister spp.), Tfor127 (T. forsythia) and L-Pgin1006-23 (P. gingivalis). SEL1150 showed specificity for bacterial species in the subclusters A, B, and C, namely S. dianae, S. infelix, S. flueggei, C. periodontii, S. artemidis, S. noxia, and S. sputigena; Sspu439 for S. sputigena; SEL1469 for subclusters A and B, and for S. sputigena; ACI623 for bacterial species in subclusters C and F, namely the S. sputigena and Veillonella species. The experimentally observed specificities of the oligonucleotide probes corresponded with our in silico analysis. Selenomonas spp. may play a role in the subgingival microbiome of periodontitis and contribute to the disease process. Targeting Selenomonas spp. with specific therapeutic strategies could offer new insights into the management of periodontitis. However, further studies are needed to determine a definite functional significance.

Microbiology Research doi: 10.3390/microbiolres17020032

Authors: Tamar Maghradze Alessandra Di Canito Carmen Cris De Oliveira Nobre Bezerra Mathabatha Evodia Setati Roberto Carmine Foschino Daniela Fracassetti Ileana Vigentini

Biocontrol, a practice for using living organisms to target plant pathogens, offers a promising, sustainable agricultural strategy. This study involves epiphytic yeasts isolated from Vitis vinifera ssp. sylvestris and ssp. vinifera as natural antagonists against Aspergillus carbonarius, Botrytis cinerea, and Penicillium expansum. Twenty-one of 37 yeasts were chosen based on the Pathology Intensity (PA) score during preliminary in vivo screening. Following identification, dual-culture assays, VOC production, copper tolerance, and commercial fungicide resistance were assessed. On YPD and GJ medium, Saccharomyces isolates were the strongest antagonists, whereas P. terricola UMY197 inhibited Penicillium and Aspergillus. H. uvarum UMY1473 was notably effective against B. cinerea. VOC analysis confirmed that S. cerevisiae UMY1430 was the most effective against Aspergillus, likely owing to its production of oxalic acid, while S. cerevisiae UMY1438 was a producer of various esters and phenylethyl alcohol. C. intermedia UMY189, M. pulcherrima UMY1472, H. uvarum UMY1473, and S. cerevisiae UMY1436 were the most copper-resistant. Yeast activity on chemical fungicide SWITCH (up to 1 g/L) depended on culture media usage; in fact, a higher viability on YPD than on GJ was observed, where only 4 yeasts were able to grow. Thus, since several yeasts exhibit promising inhibitory activity through various mechanisms and against different molds, the use of synthetic consortia could represent a powerful and essential tool in field trials to limit fungicide use while preventing the emergence of resistance.

Microbiology Research doi: 10.3390/microbiolres17020031

Authors: Renata Danielle de Souza Bartolomeu Ederson da Conceição Jesus Rafael Augusto da Costa Parrella Ivanildo Evódio Marriel Maria Lúcia Ferreira Simeone

Obtaining enzymes through bioconversion depends on a complex relationship between the microorganisms and the biomass used. Here, we evaluate xylanase production by diverse actinobacterial species, cultivated using xylan as the sole carbon source and complex media containing sorghum as the substrate. Fifty-three actinobacteria were tested for xylanase production in a solid medium. Seventeen strains produced xylanase and were tested for their ability to produce xylanase, total cellulases (filter paper activity, FPase), and endoglycanase in submerged culture using a defined liquid medium. The best xylanase-producing species was Streptomyces capoamus, yielding 24 IU·mL−1. For FPase, Streptomyces sp. showed the highest yield (1.12 IU·mL−1); for endoglycanase, the best producer was Streptomyces ossamyceticus (0.99 IU·mL−1). When sweet sorghum was used alone, S. curacoi, S. ossamyceticus, and S. capoamus showed xylanase activities of 4.5 IU·mL−1, 4.4 IU·mL−1, and 0.8 IU·mL−1, respectively. However, FPase activity was not detected under the assay conditions. The results showed that there is an intraspecific difference in xylanase, endoglucanase, and FPase production by actinobacteria, with the species S. curacoi, S. ossamyceticus, and S. capoamus able to use sorghum as a carbon source, demonstrating biotechnological potential.

Microbiology Research doi: 10.3390/microbiolres17020030

Authors: Fernando M. Rodrigues Paula Prieto-Oliveira Jean P. Zukurov Wagner T. Alkmim Michel M. Soane Michelle Camargo Sabri S. Sanabani Esper G. Kallas Maria Cecília Sucupira Ricardo Sobhie Diaz Denis Jacob Machado Luiz Mario Janini

The considerable HIV-1 genetic diversity has several implications for viral adaptive and evolutionary capabilities. Its genetic diversity is due to its high mutational rates derived from the error-prone viral reverse transcriptase activity, which generates highly heterogeneous viral populations. Moreover, genetic diversity can also increase due to intra- or intersubtype viral genomic recombination following multiple infections. This study examines HIV-1 intersubtype recombinant viruses and their increased genomic diversity over a 12-month period in five individuals from São Paulo state, Brazil. We collected peripheral blood mononuclear cells once every three months from selected participants at five distinct visits. Molecular clones of 1.15 Kbp fragments of the Pol polyprotein, spanning the protease and a portion of the reverse transcriptase (RT) genes, were generated by bulk PCR. Pol sequences were used for evolutionary analysis, including phylogenetics (using TnT), genetic diversity (using Highlighter), and hypermutation frequency (using Hypermut). Recombination detection experiments were conducted with a jumping profile-hidden Markov model (jpHMM), SimPlot++, and RDP5. We observed great genetic diversity and frequent recombination events in all patients. Furthermore, most of the patients presented hypermutations. These findings highlight the dynamic nature of HIV-1 genetic diversity, driven by frequent recombination and hypermutation, which can accelerate viral adaptation and diversification, underscoring the challenges for treatment, prevention, and disease control.

Microbiology Research doi: 10.3390/microbiolres17020029

Authors: Andrea Tommasi Cesare Bolla Laura Curci Serena Penpa Giovanni Genga Cristina Sarda Elisabetta Svizzeretto Andrea Salvaderi Giorgia Piceni Giuseppe Vittorio De Socio Daniela Francisci Antonio Maconi Guido Chichino Carlo Pallotto

Infective endocarditis (IE) due to methicillin-susceptible Staphylococcus aureus (MSSA) still represents a clinical and therapeutic issue. Discrepancies between guidelines, clinical studies and clinical practice have already been highlighted, especially regarding daptomycin use in MSSA cases. The aim of this study was to evaluate daptomycin’s impact on outcomes in this setting. This was a retrospective observational study. We enrolled all patients with MSSA IE admitted from 2015 to 2023. Patients were divided into two groups according to daptomycin administration. We enrolled 76 patients, with 49 in group A (standard treatment) and 27 in group B (treated with daptomycin). The in-hospital crude mortality was 14.3% and 29.6% in group A and B, respectively (p = 0.191). Only heart failure was significantly associated with negative outcome in the univariate and multivariate analyses (OR 6.424, 95% CI, 1.680–24.559; p = 0.007). In this study population, daptomycin treatment for IE due to MSSA was not associated with a reduced mortality rate. Heart failure was the only independent risk factor associated with in-hospital mortality.

Microbiology Research doi: 10.3390/microbiolres17010028

Authors: Elmostafa Gagou Hanae El Yeznasni Wissame Chafai Khadija Chakroune Mahmoud Abbas Touria Lamkami Mondher El Jaziri Abdelkader Hakkou

Arbuscular mycorrhizal fungi (AMF) play a pivotal role in plant adaptation to arid ecosystems, yet their widespread agricultural use is constrained by the scarcity of high-quality, locally adapted inoculum. This study established a reliable monoxenic culture system for mass-producing an indigenous AMF isolate from the date palm (Phoenix dactylifera L.) rhizosphere in the Figuig oasis, southeastern Morocco. The isolate was identified as Rhizophagus irregularis based on spore morphology and Large Subunit ribosomal DNA (LSU rDNA) phylogeny. Two propagule types, surface-sterilized spores and mycorrhizal root fragments of Plantago lanceolata L., were compared for initiation of in vitro cultures on Ri T-DNA-transformed carrot (Daucus carota L.) hairy roots. By week 16, cultures initiated from mycorrhizal root fragments produced 1414 ± 65 spores per plate and showed significantly higher performance than spore-derived cultures in terms of propagule viability, root colonization, and hairy root growth. Propagule viability reached 84% and 68%, root colonization frequencies were 95% and 72%, and hairy root lengths averaged 81 and 63 cm in root fragment- and spore-derived cultures, respectively (p < 0.01). In a subsequent whole-plant assay using P. lanceolata, in vitro-produced spores induced markedly higher mycorrhizal colonization frequency (91.0 ± 1.6% compared with 74.8 ± 1.9%) and intensity (70.0 ± 1.6% compared with 55.0 ± 1.6%) than spores obtained from conventional trap cultures (p < 0.001). These results demonstrate that monoxenic root-organ culture using root fragments is a robust, reproducible method for generating abundant, contaminant-free, and functionally superior inoculum of native R. irregularis. This advance provides a solid platform for developing tailored bio-inoculants to enhance crop resilience and sustainability in arid and semi-arid agroecosystems.

Microbiology Research doi: 10.3390/microbiolres17010027

Authors: Kayo Bianco Thereza Cristina da Costa Vianna Samara Santanna de Oliveira Kaylanne Montenegro Claudia Flores Ana Paula Alves do Nascimento Alexander Machado Cardoso Maysa Mandetta Clementino

Pseudomonas aeruginosa is an opportunistic pathogen commonly associated with nosocomial infections and environmental dissemination. Among its high-risk clones, ST244 is notable for its global distribution and distinctive genomic traits. This study reports whole-genome sequencing of ten ST244 isolates from hospitalized patients and wastewater in a healthcare complex in Southeastern Brazil. Genomic comparisons revealed a highly conserved clonal group, with nine isolates forming a tight monophyletic cluster based on rMLST, SNP phylogeny, and average nucleotide identity (>99.5%). One isolate showed close phylogenetic proximity to strains from Asia and North America, suggesting international dissemination. Serotype analysis revealed both O5 and O12 variants, indicating intra-lineage antigenic diversity. Resistance profiling identified multidrug-resistant phenotypes carrying carbapenemase genes (blaOXA-494, blaOXA-396) and diverse insertion sequences (ISPa1, ISPa6, ISPa22, ISPa32, and ISPa37), facilitating horizontal gene transfer. Virulence gene analysis showed conserved elements related to adhesion, iron uptake, secretion systems, and quorum sensing, while the cytotoxin gene exoU was absent. These results highlight clonal persistence, possible intra-hospital transmission, and links to globally circulating ST244 sublineages. Our findings underscore the importance of genomic surveillance to track high-risk P. aeruginosa clones at the clinical–environmental interface.

Microbiology Research doi: 10.3390/microbiolres17010026

Authors: Nadine Yossa Roma Adu Osei Travis Canida Anna Laasri Qing Jin Pascal Iraola Thomas Hammack Mei-Chiung Jo Huang Goran Periz Mi Sun Moon Rachel Binet

Cosmetic wipes are made for multiple functions, baby care, hand washing, feminine and personal cleansing, removing makeup, and applying products such as deodorants and sunless tanners among other uses. Despite the presence of preservatives, cosmetic wipes can become contaminated during processing steps and usage, which may lead to skin infections and other health issues for consumers. No validated method exists for the microbiological testing of cosmetic wipes. The goal of this study was to develop and validate a specific sample preparation method for the quantitative detection of microorganisms in cosmetic wipes for inclusion in the FDA Biological Analytical Manual (BAM). Ten wipe types differing in their composition and preservative combinations were inoculated with Bacillus cereus spore suspensions at three concentration levels and aged for 14 days. Three extraction methods were compared: mBAM1g (reference method using 1 g samples), mBAMww (whole wipe method based on BAM Chapter 23), and ISOww (whole wipe method based on ISO method without Tween 80). For commercial wipes, mBAMww and ISOww, using whole wipes, performed similarly (p ≥ 0.05) or significantly better (p < 0.05) than mBAM1g. For laboratory-made wipes, 1 g samples showed higher recovery rates than whole wipes, likely due to cell loss during aging. Inoculation method and preservatives affect microbial distribution, survival, and recovery rates. T80 may have a positive effect on the recovery of B. cereus from wipes. This study recommends mBAMww for the microbiological analysis of cosmetic wipes.

Microbiology Research doi: 10.3390/microbiolres17010025

Authors: Niyati Desai Nilam Vaghamshi Komal Antaliya Ashaka Vansia Arpan Tapaniya Anjana Ghelani Rajesh Chaudhari Rajesh Patel Pravin Dudhagara Douglas J. H. Shyu

Cow’s milk, urine, dung, ghee, and curd possess significant medicinal value in Ayurveda and have been integral to traditional Indian clinical practices for centuries. The cow-derived fermented product (CDFP), a formulation rooted in Ayurvedic tradition, combines these five components as a panchgavya and is believed to offer multifaceted health benefits. In this preliminary single-subject case study, we evaluated the microbial composition of CDFP itself and assessed its effects on the human gut microbiome before and after 7 and 15 days of administration. A single healthy male subject consumed CDFP daily for seven consecutive days. Using 16S rRNA metagenomic sequencing, we observed a prominent increase in gut microbial diversity and a rise in beneficial bacterial genera such as Bifidobacterium, Faecalibacterium, and Akkermansia during and after treatment. Functional profiling revealed significant enhancements in pathways associated with amino acid metabolism, vitamin biosynthesis (e.g., folate, riboflavin), and energy metabolism, along with transient boosts in secondary metabolite synthesis. Metabolomic analysis identified 171 bioactive compounds within CDFP, with 33 exhibiting interactions with human proteins involved in immune modulation, oxidative stress response, and gut barrier integrity. Although conducted on a single participant, this study is the first to elucidate the distinct changes observed in gut microbial composition and function following the seven-day CDFP regimen and provides initial insights that warrant further investigation in larger, controlled studies. These findings highlight the potential of CDFP as a microbiota-targeted intervention with health-supportive properties.

Microbiology Research doi: 10.3390/microbiolres17010024

Authors: Ding Lu Jixia Guo Xin Yan Quan Yang Xilong Zheng

To address the challenges of low land use efficiency, soil degradation, and high management costs in Ilex asprella cultivation, this study established an I. asprella–Grona styracifolia intercropping system and systematically evaluated its effects on soil nutrient cycling, microbial communities, and crop growth. Field experiments were conducted in Yunfu City, Guangdong Province, with monoculture (LCK for I. asprella, DCK for G. styracifolia) and three intercropping densities (HDT, LDT, MDT). Combining 16S rRNA sequencing and metagenomics, we analyzed the functional profile of the rhizosphere microbiome. The results showed that intercropping significantly increased the biomass of G. styracifolia, with the medium-density (MDT) treatment increasing plant length and fresh weight by 41.2% and 2.4 times, respectively, compared to monoculture. However, high-density intercropping suppressed the accumulation of medicinal compounds. In terms of soil properties, intercropping significantly enhanced soil organic carbon (SOC), total nitrogen (TN), total phosphorus (TP), and available nitrogen (AN) in the rhizosphere of both plants. Specifically, AN in the I. asprella rhizosphere increased by 18.9%. Soil urease and acid phosphatase activities were also elevated, while pH decreased. Microbial analysis revealed that intercropping reshaped the rhizosphere microbial community structure, significantly increased the Shannon diversity index of bacteria in the G. styracifolia rhizosphere, and enhanced the complexity of the microbial co-occurrence network. Metagenomic analysis further confirmed that intercropping enriched functional genes related to carbon fixation, nitrogen cycling (nitrogen fixation, assimilatory nitrate reduction), and organic phosphorus mineralization (the phoD gene), thereby driving the transformation and availability of soil nutrients. These findings demonstrate that the I. asprella–G. styracifolia intercropping system, particularly at medium density, effectively improves soil fertility and land use efficiency by regulating rhizosphere microbial functions, providing a theoretical basis for the sustainable ecological cultivation of I. asprella.

Microbiology Research doi: 10.3390/microbiolres17010023

Authors: Kyung-A Hyun Ji-Hyun Kim Min Nyeong Ko Chang-Gu Hyun

Vagococcus species have been isolated from diverse environments, including aquatic, terrestrial, food-associated, and clinical sources; however, plant- and flower-associated representatives remain poorly characterized at the genomic level. In this study, we report the complete genomic sequence and analysis of Vagococcus sp. JNUCC 83, isolated from flowers of Camellia japonica collected on Jeju Island, Republic of Korea. The genome comprises a single circular chromosome of 2,472,896 bp with a GC content of 33.5 mol% and was assembled at high depth (555.43×), resulting in a high-quality complete genome. Genome-based phylogenomic analysis using the Type (Strain) Genome Server (TYGS) showed that strain JNUCC 83 forms a distinct lineage within the genus Vagococcus. Digital DNA–DNA hybridization (dDDH) values were far below the 70% species threshold, and 16S rRNA gene-based phylogeny consistently supported its independent placement, suggesting that JNUCC 83 represents a previously undescribed genomic species. Functional annotation based on EggNOG/COG analysis indicated the enrichment of genes involved in core metabolism and genome maintenance, while antiSMASH analysis identified a terpene-precursor-type biosynthetic locus encoding a polyprenyl synthase. Overall, this study expands the genomic understanding of flower-associated Vagococcus lineages and provides a foundation for future investigations into their ecological roles and potential applications as plant-derived microbial resources.

Microbiology Research doi: 10.3390/microbiolres17010022

Authors: Yang Wang Yue Li Jingyi Wu Mengge Shen Aoqi Zhan Yuxin Wang Baobao Liu

Zinc (Zn), a ubiquitous environmental transition metal primarily existing as Zinc ions (Zn2+), plays a critical role in various biological processes. Its extensive application in agriculture, industry, and healthcare has led to significant environmental contamination. However, the mechanistic contribution of Zn2+ to bacterial antibiotic resistance and virulence remains insufficiently understood. This review explores the sources, cycling, and environmental accumulation of Zn2+ in a One Health context, emphasizing their impact on bacterial antibiotic resistance and virulence. Zn2+ promote bacterial antibiotic resistance by regulating efflux pumps, biofilm formation, expression and transfer of antibiotic resistance genes, as well as synergistic effects with other heavy metals and antibiotics. Meanwhile, Zn2+ promote bacterial virulence by regulating quorum sensing, secretion and metal homeostasis systems, as well as oxidative stress response and virulence factor expression. Additionally, it highlights the potential of targeting Zn homeostasis as a strategy to combat environmental antibiotic resistance. Collectively, these findings provide key insights into the mechanisms by which Zn2+ regulate bacterial antibiotic resistance and pathogenicity, offering valuable guidance for developing strategies to mitigate the global threat of antibiotic resistance.

Microbiology Research doi: 10.3390/microbiolres17010021

Authors: Laura Haleva Tiane Martin de Moura Luciana Costa Teixeira Horst Mitteregger Júnior Evgeni Evgeniev Gabev Adriana Ambrosini da Silveira Fabrício Souza Campos

Quality control of drinking water is essential for safeguarding public health, particularly in densely populated urban environments. Environmental microbiological monitoring can complement conventional surveillance by providing deeper insights into the dissemination of pathogens and antimicrobial resistance genes within aquatic systems. In this study, we assessed the quality of wastewater and treated water from two urban water supply systems, representing the southern and northern regions of Porto Alegre, Rio Grande do Sul, Brazil, across four climatic seasons between 2024 and 2025. Fifteen water samples were analyzed, including raw water from Guaíba Lake and treated water collected from public distribution points. The Water Quality Index was calculated, microbiological indicators were quantified, and carbapenem resistance genes were detected using molecular assays. Most treated water samples complied with established bacteriological standards; however, the blaOXA-48-like gene was recurrently detected in both wastewater and treated water. No resistance genes were identified during the summer, whereas the blaVIM gene was detected exclusively in spring samples. The presence of carbapenem resistance genes in the absence of cultivable coliforms suggests the persistence of extracellular DNA or viable but non-culturable bacteria, highlighting limitations inherent to conventional microbiological monitoring. Integrating classical microbiological methods with molecular assays enables a more comprehensive assessment of water quality and strengthens evidence-based decision-making within a One Health framework.

Microbiology Research doi: 10.3390/microbiolres17010020

Authors: Maria Martínez-Ruiz Miriam Hernández-Porto Cintia Hernández-Sánchez Ángeles Arias José Carlos de Gracia Adolfo Perdomo-González Raquel Pérez-Reverón Francisco Javier Díaz Peña

Recycled wastewater is vital for the circular economy, especially on water-scarce islands. This study explored the presence of Carbapenem-Resistant Enterobacterales and other emerging pathogens in irrigation water on four Canarian Islands, applying a One Health perspective. Using membrane filtration and MALDI-TOF mass spectrometry, 69 bacterial isolates were identified. The findings revealed that 78% were Gram-negative bacilli like Pseudomonas aeruginosa, Acinetobacter spp., Enterobacteriaceae, etc., while 22% were Gram-positive bacteria, including Enterococcus spp. The main mechanisms of carbapenem resistance in Pseudomonas spp. and Acinetobacter spp. were oxacillinases, followed by metallo-β-lactamases (MBL). In Enterobacteriaceae, characterization of carbapenemase types was less frequent, with oxacillinase 48 (OXA-48) being the most prevalent. The detection of multidrug-resistant organisms in recycled wastewater highlights an urgent need for routine microbiological monitoring in water management to protect both public health and agricultural sustainability.

Microbiology Research doi: 10.3390/microbiolres17010019

Authors: Alla Mironenko Nataliia Muzyka Nataliia Teteriuk Larysa Radchenko Anastasia Popova Jonas Waldenström Denys Muzyka

Avian influenza viruses are predominantly associated with waterfowl and shorebirds, and are rarely detected in other avian hosts in nature. In 2021, an H1N1 virus was isolated from a Fieldfare Turdus pilaris in Zaporizhzhia Oblast, Ukraine. A phylogenetic analysis revealed that all eight gene segments belonged to the Eurasian low-pathogenic avian influenza lineages. The highest nucleotide identity of the HA gene was observed with viruses detected in Georgia, Sweden, and Ukraine (99.11%), while the NA gene showed the greatest identity to viruses from Western Europe (99.14–99.57%). Genetic analysis of the HA cleavage site showed a sequence (PSIQSR↓GLF) that contained a single basic amino acid. No deletions were detected in the stalk region of NA gene, and no specific mutations in PB2 protein were found. However, several amino acid substitutions were identified in the HA gene (D204E, S207T, and D239G) that may affect the binding affinity to specific antibodies. The occurrence of this virus in a wild, seemingly healthy thrush indicate that additional surveillance in poorly studied ecological groups such as Passeriformes is warranted.

Microbiology Research doi: 10.3390/microbiolres17010018

Authors: Chiara Storoni Silvia Preziuso Anna-Rita Attili Yubao Li Vincenzo Cuteri

Bovine Respiratory Disease (BRD) represents one of the largest causes of economic loss and animal morbidity in the global cattle industry, second only to neonatal diarrhea. Its etiology is complex, originating from a multifactorial combination of host susceptibility, environmental stressors, viral infections, and secondary bacterial pathogens. Although viruses are often the initial cause of disease, suppressing the host’s respiratory defense mechanisms, most of the severe pneumonic damage and clinical signs can be attributed to bacterial infections. This review provides an overview of the primary bacterial agents identified within the BRD complex, including Mannheimia haemolytica, Pasteurella multocida, Histophilus somni, and Mycoplasma bovis. We discuss their role as commensals that then become opportunistic pathogens, and further how they interact in a synergistic relationship with a primary viral insult, leading to the resulting pathogenesis and the development of pneumonia. This manuscript discusses in further detail some of the challenges in BRD management, such as the limitations of current diagnostic methodologies, overreliance on antimicrobial therapy, and the growing concern of antimicrobial resistance. Lastly, the need for integrated approaches in management, better husbandry and biosecurity, coupled with the development of novel therapeutic alternatives, is underlined as a means of assuring a sustainable control of this serious syndrome.

Microbiology Research doi: 10.3390/microbiolres17010017

Authors: César Ricardo Cortez-Álvarez Benjamín de Jesús Gutiérrez-García Pablo Ulises Romero-Mendoza María del Rosario Cabral-Medina Monserratt Abud-Gonzalez Susana Olivia Guerra-Martínez Livier Amalia Gutiérrez-Morales María Luisa Muñoz-Almaguer Santiago José Guevara-Martínez Daniel Osmar Suárez-Rico Marco Pérez-Cisneros Martin Zermeño-Ruiz

Antimicrobial resistance (AMR) continues to represent a significant global public health concern. Gram-negative bacilli (GNB) are the primary causative agents of severe nosocomial infections and possess a notable capacity to develop resistance mechanisms that restrict therapeutic options. The objective of this study was to characterize the antimicrobial susceptibility profiles of GNB isolated at a secondary-level hospital in Guadalajara, Mexico, with the aim of identifying predominant resistance patterns and the most effective therapeutic alternatives. A descriptive, retrospective, cross-sectional study was conducted using clinical isolates of Acinetobacter spp., Pseudomonas spp., Escherichia coli, Klebsiella spp., Morganella morganii, Proteus spp., and Enterobacter spp. collected during 2024. The identification and susceptibility testing were carried out using the VITEK® 2 automated system, and the results were interpreted in accordance with CLSI guidelines. High resistance rates were observed in Acinetobacter spp. and Pseudomonas spp., particularly to carbapenems (>50% and >40%, respectively). Escherichia coli and Klebsiella spp. demonstrated resistance to third-generation cephalosporins and trimethoprim/sulfamethoxazole, exhibiting high susceptibility to amikacin and carbapenems (>90%). New-generation β-lactam/β-lactamase inhibitor combinations, such as ceftazidime/avibactam and ceftolozane/tazobactam, have demonstrated high efficacy against resistant strains. Overall, GNB isolates in this secondary-level hospital demonstrated elevated resistance levels, particularly to β-lactams and carbapenems, which pose a significant therapeutic challenge. Nevertheless, amikacin, carbapenems, and new-generation β-lactams persist as valuable therapeutic options. In order to contain the spread of multidrug-resistant organisms, it is imperative to strengthen local surveillance, optimize antibiotic stewardship, and reinforce infection control measures.

Microbiology Research doi: 10.3390/microbiolres17010016

Authors: Grissana Pook-In Somsak Tammawong Chorpaka Phuangsri Khwanla Seansupa Sontaya Sookying Tomoko Takahashi Anchalee Rawangkan

Streptomyces is a versatile genus widely used in drug production and biotechnological applications. This study aimed to identify and characterize bioactive compounds produced by Streptomyces UP-AC4 and UP-3.2 strains and evaluate their antibacterial and anticancer activities. The strains were identified as Streptomyces californicus and Streptomyces purpurascens via chemotaxonomy, 16S rRNA sequencing, amplified ribosomal DNA restriction analysis, and phylogenetic analysis. Bioactive compounds were extracted using heat treatments at 63 °C for 30 min or 73–110 °C for 10 min. Antibacterial activity against Staphylococcus aureus, Bacillus cereus, and Escherichia coli was assessed by agar disc assay, with MICs of 0.024–0.195 mg/mL and MBCs of 0.098–0.391 mg/mL for the most effective extracts. Anticancer activity against A549, H1299, and Lu99 lung cancer cells was evaluated using the MTT assay, showing IC50 values of 0.23 ± 0.06 to 4.85 ± 0.64 mg/mL, while exhibiting no toxicity to normal fibroblast cells. HPLC analysis indicated that heat-assisted extraction of UP-AC4 at 73 °C for 10 min enriched a dinactin-associated compound as a predominant metabolite with antibiotic and anticancer activities. In conclusion, Streptomyces UP-AC4 and UP-3.2 produce promising low-cost bioactive compounds with strong potential for pharmaceutical and healthcare applications.

Microbiology Research doi: 10.3390/microbiolres17010015

Authors: Valery M. Dembitsky Alexander O. Terent’ev

Microorganisms have emerged as prolific and versatile producers of steroidal natural products, displaying a remarkable capacity for structural diversification that extends far beyond classical sterol frameworks. This review critically examines steroidal metabolites isolated from microbial sources, with a particular emphasis on marine-derived and endophytic fungi belonging to the genera Aspergillus and Penicillium, alongside selected bacterial and lesser-studied fungal taxa. Comparative analysis reveals that these organisms repeatedly generate distinctive steroid scaffolds, including highly oxygenated ergostanes, secosteroids, rearranged polycyclic systems, and hybrid architectures arising from oxidative cleavage, cyclization, and Diels–Alder-type transformations. While many reported compounds exhibit cytotoxic, anti-inflammatory, antimicrobial, or enzyme-inhibitory activities, the biological relevance of these metabolites varies considerably, highlighting the need to distinguish broadly recurring bioactivities from isolated or strain-specific observations. By integrating structural classification with biosynthetic considerations and bioactivity trends, this review identifies key steroidal frameworks that recur across taxa and appear particularly promising for further pharmacological investigation. In addition, current gaps in mechanistic understanding and compound prioritization are discussed. Finally, emerging strategies such as genome mining, biosynthetic gene cluster analysis, co-culture approaches, and synthetic biology are highlighted as powerful tools to unlock the largely untapped potential of microbial genomes for the discovery of novel steroidal scaffolds. Together, this synthesis underscores the importance of microorganisms as a dynamic and expandable source of structurally unique and biologically relevant steroids, and provides a framework to guide future discovery-driven and mechanism-oriented research in the field.

Microbiology Research doi: 10.3390/microbiolres17010014

Authors: Kareem Badr Fatima Al-Maadeed Alaa Azouz Zahoor Ul Hassan Quirico Migheli Samir Jaoua

Microbial communities found in arid environments often exhibit unique genetic and metabolic adaptations that enable them to synthesize potent bioactive compounds. Bacillus thuringiensis (Bt) is widely recognized for its biocontrol potential against various insects. This study aims to investigate the insecticidal potential of Bt strains isolated from Qatar’s soil against dipteran and lepidopteran larvae. The microscopic analysis identified distinct crystal types, including bipyramidal, cuboidal, spherical smooth, and spherical rough forms, with distinct cry, cyt, and vip genes. Strains producing bipyramidal crystals carry cry1A, cry2A, and vip3A genes, while only two strains producing spherical crystals carry cry4B and cyt1A genes. Bipyramidal crystal-producing strains (QBT552 and QBT877) showed potent insecticidal activity, achieving 100% mortality against Corcyra cephalonica larvae, with LC50 values of 25 µg/g. Spherical smooth crystal-producing strain (QBT862) exhibited high toxicity against Culex pipiens insect larvae (LC50 = 2 µg/L). The quantification of bipyramidal crystal protein production of strains QBT877 and QBT552 exhibited the highest δ-endotoxin yield (1334.4 ± 6.7 and 1188.7 ± 5.0 µg/mL, respectively), while smooth spherical crystal strains QBT758 and QBT862 were 577.5 ± 8.4 and 567.6 ± 8.4 µg/mL, respectively. These findings highlighted the potential of Bt QBT strains for biocontrol applications, with strains showing promise for producing effective δ-endotoxins.

Microbiology Research doi: 10.3390/microbiolres17010013

Authors: Seulgi Lee Fanbin Kong Jinru Chen

Salmonella enterica from low-moisture food has been found to have a higher thermal tolerance than from high-moisture food. However, the molecular mechanism underlying the association of thermal tolerance of this pathogen with low-moisture foods, such as peanut butter and peanut spread, has not been fully elucidated. We previously found that mutants of S. Tennessee with a defective gene encoding a cell membrane lipoprotein (Lpa) or cell division protein (ZapC) formed significantly (p ≤ 0.05) less biofilm than the wildtype strain. To assess the possible role of these genes in the thermal tolerance of S. Tennessee, this study compared the surviving populations of the wildtype S. Tennessee and its mutants defective in Lpa or ZapC in different types of peanut products (regular, reduced-fat, and natural) at 74 °C for 0, 2.5, 5, 10, 15, 20, 30, 40, or 50 min. Results showed that mutants with a defective lpa or zapC significantly affected the survival of Salmonella in peanut products during heat treatments. Significantly, a higher reduction in Salmonella population was observed in regular peanut butter, followed by natural and reduced-fat peanut spreads. The study provides new insight into one of the molecular mechanisms underlying the thermal tolerance of Salmonella enterica.

Microbiology Research doi: 10.3390/microbiolres17010012

Authors: Muhammad Halwani Manal Al Daajani Safa Boujemaa

Streptococcus pneumoniae (S. pneumoniae) is responsible for a wide range of infections. The aim of this study was to investigate the clonal diversity of S. pneumoniae in thirteen Arab countries. Multi-Locus Sequence Typing (MLST) data were extracted from PubMLST database. Genetic analysis was performed using DnaSP software version 6.0. A Minimum Spanning Tree (MST) analysis was conducted to evaluate the population structure of S. pneumoniae strains. Genetic data from 1008 Arab S. pneumoniae strains, collected over 22 years (1996–2018), were analyzed. MLST analysis identified a highly diverse population comprising 600 sequence types grouped into 87 clonal complexes and 295 singletons. Both internationally disseminated clones (e.g., ST156) and country-specific lineages (e.g., ST2307, Saudi Arabia) were observed, indicating substantial geographic structuring. Significant associations were detected between sequence types and geographical origin, decade of isolation, patient age, disease type, and serotype (p < 0.05). Although recombination events were presented, the population retained a predominantly clonal structure over time (ISA = 0.0715, p < 0.001). Overall, these findings demonstrated extensive genetic heterogeneity and spatiotemporal structuring of S. pneumoniae in the Arab region, providing valuable insights for regional surveillance and vaccine-related strategies.

Microbiology Research doi: 10.3390/microbiolres17010011

Authors: Dawid Jańczak Piotr Górecki Natalia Skrzypek Dominika Sobkiewicz Magda Paczocha Aleksander Chrzanowski Aleksandra Kornelia Maj Rafał Stryjek Aleksandra Anna Zasada Anna Golke

Bacterial urinary tract infections (UTIs) are common in dogs and cats. This study examined the correlations between routine urinalysis and culture-confirmed infections and described the etiologic agents and antimicrobial susceptibility to support stewardship. In 2023, 1787 urine samples (854 dogs, 933 cats) underwent urinalysis, aerobic culture with species identification, and disk-diffusion testing per Clinical and Laboratory Standards Institute standards; non-parametric statistics with effect sizes were applied. Pyuria was the strongest correlate of infection in both species. Low urine specific gravity was associated with infection and crystal detection, and urine pH correlated weakly with growth in dogs. Nitrite positivity was strongly associated with infection in dogs but showed no diagnostic value in cats. Hematuria showed a weak inverse association in dogs and no association in cats. Females and older animals were more frequently infected, and infections were slightly more common in summer. Most episodes were monomicrobial (85%), predominantly caused by Escherichia coli (48.4% of dogs; 51.5% of cats). E. coli remained broadly susceptible to nitrofurantoin and aminoglycosides. Fluoroquinolone activity was variable. Pseudomonas spp. showed the highest susceptibility to ceftazidime, cefepime, and aminoglycosides. These findings support culture when pyuria, dilute urine, or nitrite positivity is detected and favour short, targeted empiric therapy pending results, guided by a stepwise clinical decision pathway.

Microbiology Research doi: 10.3390/microbiolres17010010

Authors: Aziza Usmonkulova Massimo Pugliese Mukhiddin Juliev Ilkhom Khalilov Nafosat Kurbonova Nigora Tillyaxodjayeva Rixsiniso Karimova Wei Liu Feruza Khalilova Oysha Jabborova

This study involved a systematic literature review using bibliometric analysis to examine the evolution and current trends of Biological Remediation studies. The bibliometric analysis was used for the descriptive, intellectual, social, and conceptual network analyses, while systematic reviews were used to identify the application of the Biological Remediation. A total of 4835 papers were selected and extracted from Scopus between 2020 and 2025. The publication trends, most influential countries and articles, leading journals, collaboration networks, coupling networks, and application of the Biological Remediation in various disciplines were described. This study summarized the research agenda of the Biological Remediation field, which would be helpful for researchers and funding agencies. This article highlights four new research directions in Current Bioremediation Trends: (1) understanding the interactions between petroleum hydrocarbons and heavy metals in composite pollution systems; (2) exploring microbial community succession during bioremediation; (3) utilizing biosurfactants to enhance contaminant solubilization and biodegradation; and (4) developing integrative, multi-mechanistic remediation approaches.

Microbiology Research doi: 10.3390/microbiolres17010009

Authors: Žejko Savković Jelena Burazerović Marija Jovanović Sara Arsenijević Miloš Stupar

Cave ecosystems represent environmentally constrained habitats that host diverse and highly specialized fungal communities. Many cave-dwelling fungi act as decomposers, transient colonizers, or cave fauna symbionts. During a mycological survey of Sesalačka cave (Serbia), a previously undescribed species was isolated from the skin of a Miniopterus schreibersii. The aim of this study was to characterize this isolate using an integrative taxonomic approach combining morphology, physiology, and multilocus phylogenetics. The fungus was cultured on different media under and its morphophysiological traits were recorded. DNA sequences of ITS, LSU, SSU, and TEF1α were compared with existing Gamsia species and phylogenetic analysis placed the isolate within the Gamsia clade, forming a well-supported lineage the most closely related to G. aggregata, but differing from it by 8–12 base pairs across loci. Distinctive morphological features of this species include obovoid to pyriform polyblastic conidia, hyaline to pale-brown annelloconidia, and reduced conidiophores, clearly separating the species from described congeners. It is psychrotolerant and does not grow at 37 °C, suggesting it is a cave-associated saprobe rather than a mammalian pathogen. This study expands the known diversity of Gamsia species and contributes to the growing evidence that subterranean habitats harbor numerous undescribed fungal kingdom members.

Microbiology Research doi: 10.3390/microbiolres17010008

Authors: Geane Andriollo Paradynski Ronaldo dos Santos Machado Lucas Machado Sulzbacher Maicon Machado Sulzbacher Vítor Antunes de Oliveira Pauline Brendler Goettems Fiorin Mirna Stela Ludwig Thiago Gomes Heck Matias Nunes Frizzo

Inappropriate use of antibiotics can stimulate antimicrobial resistance, since bacteria are capable of circumventing pharmacological action through various resistance mechanisms. Recently, during the COVID-19 pandemic, there has been an increase in the use of antimicrobials. This is an analytical, quantitative, and retrospective study on bacterial resistance and mortality in Intensive Care Unit (ICU) patients from 2017 to 2022. This study analyzed sociodemographic aspects, clinical, and laboratory parameters in patients admitted to the ICU. A total of 221 medical records of patients with multidrug-resistant bacteria in the ICU were included, with an outcome of 95 discharges (42.98%) and 126 deaths (53.01%). An increase in the prevalence of bacterial resistance in the ICU was identified during the Pandemic period, when compared to the Pre-Pandemic period. It was identified that the increase in bacterial resistance of some pathogens was associated with death. It was also observed that age was a factor for an increased risk of mortality in the ICU, no matter the sex of the patient. Importance of careful analysis in the use of antimicrobials, as well as in the care of ICU patients and in the surveillance of bacterial infections by health professionals.

Microbiology Research doi: 10.3390/microbiolres17010007

Authors: Rabab A. D. Meshan Norah Ahmed AlOsaimi Abdulmohsen Redha Maribasappa Karched

Kuwait faces a significant public health challenge from obesity and Type 2 Diabetes Mellitus (T2DM), conditions known to disrupt the natural balance of oral bacteria. This imbalance, or dysbiosis, can promote gum disease and may worsen metabolic health. While the intragastric balloon (IGB) is a common, less invasive weight-loss procedure, its specific effect on the community of bacteria in saliva remains unclear, especially for high-risk groups. The objective of this study was to investigate changes in the salivary microbiota of obese prediabetic patients following IGB placement. We recruited 20 obese patients (11 female, 9 male; average age 31.5) from a clinic in Kuwait. Saliva samples were collected just before IGB (Allurion™) insertion and again 6 weeks after that. Using 16S rRNA gene sequencing, we identified the bacterial species present and used bioinformatic tools to analyze diversity and abundance. Our analysis revealed that the overall diversity and structure of the salivary microbial community remained stable after the procedure. However, we detected notable changes in specific types of bacteria. The relative abundance of several genera, including Veillonella, Porphyromonas, and Fusobacterium, shifted significantly. At the species level, Porphyromonas pasteri and Haemophilus parainfluenzae became less abundant, while certain Veillonella and Streptococcus species increased in number after the IGB was placed. In conclusion, for obese prediabetic patients in Kuwait, the salivary microbiome demonstrates remarkable stability in the weeks following IGB surgery. The procedure did not drastically alter the overall ecosystem, but it did trigger specific, subtle changes in certain bacterial populations. This suggests the oral microbiota is resilient, adapting to the new physiological conditions without a major upheaval.

Microbiology Research doi: 10.3390/microbiolres17010006

Authors: Hyun-Sol Jo Youl-Lae Jo Sun-Mee Hong

Pseudomonas extremorientalis PEY1, isolated from the intestinal contents of marine fish, was evaluated for the production and properties of antibacterial proteins active against Edwardsiella tarda, a major pathogen in aquaculture. Antibacterial production was maximized in a minimal medium supplemented with 1% yeast extract and 1% galactose under stationary cultivation at 25 °C and pH 7.0. Growth and bioactivity assays were conducted under varying carbon and nitrogen sources, temperatures, and pH levels. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed a distinct ~37 kDa protein band corresponding to antibacterial activity, exhibiting an inhibition zone of 2.4 ± 0.1 cm against E. tarda. The activity was completely abolished by papain digestion but remained detectable after exposure to 55 °C and pH 8, indicating that the active compound is a moderately heat-stable, proteinaceous antibacterial molecule. LC–MS/MS analysis identified the protein as a putative disulfide reductase with ~40% sequence coverage. The antibacterial factor exhibited strong physicochemical stability, retaining activity in the presence of surfactants and metal ions. Collectively, these findings demonstrate that P. extremorientalis PEY1 produces a thermostable, papain-sensitive antibacterial protein with selective activity against E. tarda, highlighting its potential as a promising natural biocontrol or postbiotic candidate for sustainable aquaculture.

Microbiology Research doi: 10.3390/microbiolres17010005

Authors: Burenjargal Otgonsuren Hangyu Lan Douglas L. Godbold

In forest soils, nitrogen (N) is present in inorganic and organic forms. The organic forms include monomeric amino acids, but also polymers such as chitin. Ectomycorrhizal fungi are known to take up both inorganic and organic N forms, and to depolymerize large organic compounds; however, it is unknown if the compounds are used for growth. The aim of this investigation was to determine the growth of a range of ectomycorrhizal fungi on inorganic and organic N sources. Seven ectomycorrhizal fungi and one endophyte originating from mountain regions either in Austria, Mongolia, or Slovenia were grown in in-vitro cultures containing ammonium, nitrate, or chitin. Four ectomycorrhizal fungi were used to investigate growth on amino acids. All fungi, except Paxillus involutus, utilized nitrate as a N source. All fungi also grew on both chitin and N-acetylglucosamine, the amino sugar precursor of chitin. Paxillus involutus and Melanogaster broomeanus showed enhanced growth on chitin-containing media. Amanita muscaria, Rhizopogon roseolus, and Suillus granulatus, but not Paxillus involutus, were able to utilize the amino acids glycine and glutamate, as well as the tripeptide triglycine. The ability to utilize the different N sources was independent of the origin of the fungi.

Microbiology Research doi: 10.3390/microbiolres17010004

Authors: Lei Fan Yuqing Lei Zhihui Qi Haiyang Zhang Lin Tian Fang Tang

Fumonisins, a major class of mycotoxins, pose significant health risks to humans and animals due to their widespread contamination and potent toxicity. Recent advances in molecular biology, biochemistry, and enzymology have greatly enhanced the understanding of fumonisin biosynthesis and its genetic regulation. The key biosynthetic genes are typically organized in clusters and regulated by specific transcription factors; increasing evidence also highlights the involvement of complex transcriptional and epigenetic mechanisms. Environmental factors such as nitrogen, carbon, and pH also modulate these regulatory networks. Despite substantial progress, critical gaps remain in fully elucidating the regulatory pathways that control fumonisin production. This review synthesizes current knowledge regarding fumonisin biosynthesis, gene clusters, and multi-level regulatory mechanisms, while emphasizing recent trends, existing challenges, and potential applications in food safety and biotechnology to enhance food security and promote sustainable development.

Microbiology Research doi: 10.3390/microbiolres17010003

Authors: Sinazo Kondlo Zizipho Z. A. Mbulawa

Ureaplasma species are associated with various reproductive health issues, while human papillomavirus (HPV) is associated with cervical, vaginal and vulvar cancers. Data on the association between Ureaplasma species and HPV are limited in South Africa. This study investigated the prevalence of Ureaplasma urealyticum (U. urealyticum), Ureaplasma parvum (U. parvum), and HPV coinfection and their associated factors, among adolescent girls and young women (AGYW) in the Eastern Cape Province, South Africa. A total of 214 participants were retrospectively recruited, and secondary data on HPV, U. urealyticum, U. parvum, demographics, and sexual behavior were used. HPV was detected using the Roche Linear Array HPV Genotyping Test, while U. urealyticum and U. parvum were detected using Allplex™ sexually transmitted infection (STI) essential Assay. Statistical analyses were performed using GraphPad Prism Version 8.0.1.244. The prevalence of U. urealyticum was 43.9% (94/214) and increased significantly with age (p = 0.036, R2 = 0.8497); while U. parvum prevalence was 68.7% (147/214) and was not influenced by age. Having four to six lifetime sexual partners (PR: 1.77, 95% CI: 1.04–3.00, p = 0.043) was associated with increased risk of U. urealyticum. A proportion of 36.3% (77/212) had HPV-U. urealyticum coinfection and its risk was increased among those with 3–6 lifetime sexual partners (PR: 1.59, 95% CI: 1.10–2.53, p = 0.017), 2–4 new partners past three months (PR: 2.14, 95% CI: 1.19–2.42, p = 0.021); vaginal sexual intercourse frequency past 1-month (2–3 vaginal intercourse: PR: 1.54, 95% CI: 1.06–2.53, p = 0.037; 4–10 vaginal intercourse: PR: 1.91, 95% CI: 1.83–1.91, p = 0.005) and alcohol consumption (PR: 1.85, 95% CI: 1.20–3.28, p = 0.004). U. urealyticum positives had a significantly higher risk of HPV types targeted by Cervarix® HPV vaccine than negatives (PR: 2.56, 95% CI: 1.23–5.37, p = 0.013), Gardasil®4 (PR: 2.16, 95% CI: 1.25–3.75, p = 0.006) and Gardasil®9 (PR: 1.70, 95% CI: 1.25–2.32, p = 0.001). AGYW of Eastern Cape Province, South Africa had high prevalence of U. urealyticum-HPV and U. parvum-HPV coinfections. Ureaplasma species coinfection was associated with HPV prevalence and distribution of genotypes. The U. urealyticum prevalence and its coinfection with HPV were associated with sexual behavior. Data from this study could contribute to the design of sexual health and STI interventions and could serve as a baseline for future epidemiological studies, which include ongoing surveillance of HPV genotype prevalence to evaluate the impact and effectiveness of HPV vaccination programs in the population.

Microbiology Research doi: 10.3390/microbiolres17010002

Authors: Fátima Jael Olvera-Muñoz Martina Hilda Gracia-Valenzuela Fabiola Lango-Reynoso Olaya Pirene Castellanos-Onorio Jesús Montoya-Mendoza Christian Reyes-Velázquez María de Lourdes Fernández-Peña Bani Mariana Ruesgas-Ramon María del Refugio Castañeda-Chávez

The Veracruz Reef System National Park (VRSNP), located in the central Gulf of Mexico, is one of the country’s most ecologically and economically significant coral systems. Despite its high biodiversity and ecosystem functionality, it is affected by anthropogenic inputs such as fluvial discharges, urban effluents, and port and tourism activities that contribute organic and bacteriological loads. This study aimed to identify the presence of Enterococcus spp. and Vibrio spp. during three climatic seasons—dry, rainy, and north winds—at two water column depths (surface and bottom) across three reefs (Enmedio, Chopas, and Gallega) within the VRSNP during the 2022 annual cycle. Samples were analyzed according to national and international standards. Results showed that Vibrio spp. were influenced mainly by temporal factors, with higher values during north winds and the dry season (>1100 MPN/100 mL); otherwise, rainy conditions reported the lowest load (184.89 ± 15.00 MPN/100 mL). While Enterococcus spp. exhibited greater spatial influence, particularly in surface waters, Enmedio Reef recorded the highest load (478.34 ± 37.28 CFU/100 mL); in addition, Chopas Reef reported the lowest at the bottom (12.43 ± 1.26 CFU/100 mL). The findings highlight the need to strengthen microbiological monitoring protocols in marine coastal ecosystems to assess water quality, public health risks, and the ecological integrity of coral reef environments, as well as the implementation of molecular identification techniques.

Microbiology Research doi: 10.3390/microbiolres17010001

Authors: José Luis Aguirre-Noyola Gustavo Cuaxinque-Flores Jorge David Cadena-Zamudio Marco A. Ramírez-Mosqueda Lorena Jacqueline Gómez-Godínez Juan Ramos-Garza

The green synthesis of silver nanoparticles (AgNPs) by bacteria is a strategic route for sustainable nanobiotechnology; however, the genomic and biochemical mechanisms that make it possible remain poorly defined. In this study, bacteria native to silver-bearing mine tailings in Taxco (Mexico) were isolated, capable of tolerating up to 5 mM of AgNO3 and producing extracellular AgNPs. Spectroscopic (430–450 nm) and structural (XRD, fcc cubic phase) characterization confirmed the formation of AgNPs with average sizes of 17–21 nm. FTIR evidence showed the participation of extracellular proteins and polysaccharides as reducing and stabilizing agents. Genomic analyses assigned the isolates as Lysinibacillus fusiformis 31HCl and L. xylanilyticus G1-3. Genome mining revealed extensive repertoires of genes involved in uptake, transport, efflux and detoxification of metals, including P-type ATPases, RND/ABC/CDF transporters, Fe/Ni/Zn uptake systems, and metal response regulators. Notably, homologues of the silP gene, which encode Ag+ translocator ATPases, were identified, suggesting convergent adaptation to silver-rich environments. Likewise, multiple nitroreductases (YodC, YdjA, YfKO) were detected, candidates for mediating electron transfer from NAD(P)H to Ag+. These findings support the role of Lysinibacillus as microbial nanofactories equipped with specialized molecular determinants for silver tolerance and AgNP assembly, providing a functional framework for microorganism-based nanobiotechnology applications.

Microbiology Research doi: 10.3390/microbiolres16120263

Authors: Hernando Maldonado-Pérez Juan Pablo Pinzón-Esquivel Gloria María Molina-Salinas Avel Adolfo González-Sánchez Haziel Eleazar Dzib-Baak Ángel Dzul-Beh Carlos Javier Quintal-Novelo Andrés Humberto Uc-Cachón

Background: Infections caused by Staphylococcus aureus are a health problem worsened by antibiotic resistance. New drugs, including those inhibiting virulence and resistance mechanisms, are needed. This study aimed to evaluate the anti-growth, anti-virulence, and anti-biofilm activities of Trametes villosa. (2) Methods: Fractions were obtained from the basidiomata of T. villosa. Anti-growth, anti-hemolysis, and anti-biofilm activities were tested against S. aureus strains using resazurin microtiter, blood cell lysis, and crystal violet assays, respectively. Cytotoxicity was evaluated in Vero and HaCaT cells using sulforhodamine B. The active fractions were subjected to GC-MS analysis and molecular docking with S. aureus quorum-sensing receptors. Results: The n-hexane and ethyl acetate (EtOAc) fractions exhibited anti-growth activity against all strains (MIC: 31.2–2000 µg/mL). These fractions also displayed anti-hemolysis (IC50 = 33.8 ± 1.1–53.8 ± 5.1 µg/mL) and anti-biofilm formation activity (IC50 = 106.6 ± 4.8–383.4 ± 31.4 µg/mL), while exhibiting low cytotoxicity in Vero and HaCat. GC-MS analysis revealed that both active fractions mainly contained alkanes, aldehydes, and fatty acids. Molecular docking revealed that isovanillic acid, identified in the EtOAc fraction, exhibited optimal interactions with S. aureus quorum-sensing receptors AgrA and SarA. (4) Conclusions: Our research highlights the potential of T. villosa as a source of bioactive compounds effective against S. aureus.

Microbiology Research doi: 10.3390/microbiolres16120262

Authors: Himanshu Jangid Chirag Chopra Atif Khurshid Wani

Carbapenem-resistant Enterobacterales (CREs) pose a critical threat to global public health, largely driven by the enzymatic activity of Klebsiella pneumoniae carbapenemase-2 (KPC-2), a class A serine β-lactamase that hydrolyzes most β-lactam antibiotics. While β-lactamase inhibitors like avibactam offer temporary relief, emerging KPC variants demand novel, sustainable inhibitory scaffolds. This study aimed to identify and characterize potential natural inhibitors of KPC-2 from Pongamia pinnata, leveraging a comprehensive in silico workflow. A curated library of 86 phytochemicals was docked against the active site of KPC-2 (PDB ID: 3DW0). The top-performing ligands were subjected to ADMET profiling (pkCSM), and 100 ns molecular dynamics simulations (GROMACS) to evaluate structural stability and interaction persistence, using avibactam as control. Ponganone V exhibited the most favorable binding energy (−9.0 kcal/mol), engaging Ser70 via a hydrogen bond and forming π–π interactions with Trp105. Glabrachromene II demonstrated a broader interaction network but reduced long-term stability. ADMET analysis confirmed high intestinal absorption, non-mutagenicity, and absence of hERG inhibition for both ligands. Molecular dynamics simulations revealed that Ponganone V maintained compact structure and stable hydrogen bonding throughout the 100 ns trajectory, closely mirroring the behavior of avibactam, whereas Glabrachromene II displayed increased fluctuation and loss of compactness beyond 80 ns. Principal Component Analysis (PCA) further supported these findings, with Ponganone V showing restricted conformational motion and a single deep free energy basin, while avibactam and Glabrachromene II exhibited broader conformational sampling and multiple energy minima. The integrated computational findings highlight Ponganone V as a potent and pharmacologically viable natural KPC-2 inhibitor, with strong binding affinity, sustained structural stability, and minimal toxicity. This study underscores the untapped potential of Pongamia pinnata phytochemicals as future anti-resistance therapeutics and provides a rational basis for their experimental validation.

Microbiology Research doi: 10.3390/microbiolres16120261

Authors: You-Zuo Chen Chieh-Ting Chen Tsung-Wei Shih Wei-Hsuan Hsu Bao-Hong Lee Tzu-Ming Pan

Gut microbiota dysbiosis is increasingly being recognized as a major contributor to host metabolic imbalance, immune dysfunction, and neurophysiological disorders. Probiotics are known to modulate intestinal metabolism and exert systemic effects through the gut–brain axis. Herein, we evaluated the safety and probiotic potential of Pediococcus acidilactici SWP-CGPA01 (SWP-CGPA01) under antibiotic-induced microbiota dysbiosis. Genomic and phenotypic analyses verified its safety profile, supporting its suitability for use in food and nutritional applications. In a mouse model of antibiotic-induced dysbiosis, SWP-CGPA01 supplementation alleviated diarrhea and restored hippocampal brain-derived neurotrophic factor expression. These findings demonstrate that SWP-CGPA01 is a safe and functionally active probiotic with the potential to maintain gastrointestinal and neurotrophic homeostasis under gut microbiota dysbiosis.

Microbiology Research doi: 10.3390/microbiolres16120260

Authors: Ikram Markaoui Meryem Idrissi Yahyaoui Abdeslam Asehraou Abdelkrim Daoudi Brahim Housni Houssam Bkiyar

Multidrug-resistant (MDR) pathogens and biofilm-associated infections represent a major global health concern, particularly in the context of medical devices such as catheters, tubing, and blood sampling devices. Biofilms, responsible for up to 85% of human infections, confer a high level of microbial resistance and compromise device performance and patient safety. In this study, the antibiofilm potential of Syzygium aromaticum (clove) essential oil was investigated through an in vitro assay. GC–MS analysis revealed eugenol (72.77%) as the predominant compound, accompanied by β-caryophyllene (14.72%) and carvacrol (2.09%). The essential oil exhibited notable antimicrobial activity, producing inhibition zones of 30.5 ± 4.5 mm against Staphylococcus aureus, 24.5 ± 0.5 mm against Micrococcus luteus, 16.0 ± 2.0 mm against Escherichia coli, 13.0 ± 1.0 mm against Pseudomonas aeruginosa, 23.5 ± 1.5 mm against Candida albicans, and 24.0 ± 2.0 mm against C. glabrata. A marked reduction in biofilm biomass observed on polyvinyl chloride (PVC) surfaces. The application of clove essential oil as a coating for PVC-based medical devices remains a future possibility that requires formulation and in vivo testing. This strategy is proposed as potentially eco-safe, although environmental toxicity and biocompatibility have not yet been evaluated. It could contribute to the prevention of biofilm formation in arterial sampling systems and other healthcare-related materials, thereby enhancing device safety and longevity.

Microbiology Research doi: 10.3390/microbiolres16120259

Authors: Kyung-A Hyun Ji-Hyun Kim Min Nyeong Ko Chang-Gu Hyun

Enterococcus alishanensis JNUCC 77 (=BLH10) was isolated from the flowers of Zinnia elegans collected at Ilchul Land, Jeju Island, Republic of Korea. Whole-genome sequencing was conducted to clarify its taxonomic position, genomic composition, and adaptive metabolic potential. The assembled genome comprised five contigs totaling 3.86 Mb, with a G + C content of 35.6% and 100% completeness. Genome-based phylogenomic analyses using the Type Strain Genome Server (TYGS) and digital DNA–DNA hybridization (dDDH) confirmed that strain JNUCC 77 belongs to E. alishanensis. Functional annotation revealed enrichment of genes related to transcriptional regulation, carbohydrate metabolism, replication, and DNA repair, suggesting a lifestyle adapted to oxidative and UV-exposed floral habitats rather than pathogenic competitiveness. Genome mining with antiSMASH identified two putative biosynthetic regions associated with terpenoid and isoprenoid metabolism, which are commonly linked to redox regulation and cellular protection. These genomic features indicate that E. alishanensis JNUCC 77 has evolved a metal-assisted, redox-regulated survival strategy suitable for floral microenvironments. Given its origin from vibrant flowers and its genomic potential for redox-protective metabolism, this strain represents an attractive microbial resource for future development of nature-inspired postbiotic and cosmeceutical ingredients that align with the clean and eco-friendly image of flower-derived biotechnologies.