Search Results (311)

Listeria monocytogenes is a persistent foodborne pathogen capable of surviving in food processing environments, often in association with diverse environmental microflora. This study examines genomic determinants of persistence, specifically stress adaptation and biofilm-associated traits, in environmental Listeria species and other environmental microflora from a dairy processing facility by analyzing whole-genome sequences of 6 environmental Listeria isolates, 4 ATCC reference strains, and 22 air and floor swab cultures, annotated using the RAST platform. Subsystem analysis revealed that Listeria isolates carried a defined set of genes linked to biofilm formation, antimicrobial resistance, and stress response, though in lower abundance than environmental cultures. Listeria exhibited fewer flagellar genes but greater consistency in core stress-related genes, including those for disinfectant and osmotic stress resistance, with SigB operon and RpoN genes highlighting strong stress tolerance. In contrast, environmental cultures exhibited broader transcriptional regulators (RpoE, RpoH) and greater diversity in acid and heat shock response genes, indicating distinct survival strategies. All examined Listeria species harbor biofilm and stress-resistance genes enabling independent survival, while environmental microbiota show greater genetic diversity that may promote persistence and multispecies biofilm formation. This study underscores the complex genetic landscape that may contribute to the persistence of Listeria and environmental microbiota in dairy processing environments, providing foundational insights for environmental cross contamination control strategies. Full article

The complex microbiota of cheese starters plays a key role in determining the structure and flavour of the final product, primarily through their acid-forming capacity, protease activity, and exopolysaccharide synthesis. However, the specific microbial communities underlying the unique qualities of artisanal cheeses remain poorly understood. This study presents the microbiological and molecular genetic characterisation of the microbiome isolated from an artisanal cheese starter in Kosh-Agach, Altai, Russia. Metagenomic analysis of this starter revealed the presence of three bacterial genomes corresponding to those of Lactococcus lactis. Pure cultures from this starter were obtained by sequential subculture, and seventeen colonies displaying distinct characteristics on differential media were selected. Genome sequencing was performed for each colony. Bioinformatic analysis based on the rpoB gene grouped the isolates into three clusters, each corresponding to a distinct strain of Lactococcus lactis subsp. diacetilactis. This classification was further confirmed by microbiological and microscopic analyses. A notable finding was that none of the strains produced the characteristic aroma compounds of L. l. subsp. diacetilactis, namely, diacetyl and CO₂. The functional properties and metabolic characteristics of this starter consortium are discussed. Full article

Rifampicin-resistant tuberculosis (RR-TB) remains a major threat to global TB control, primarily driven by mutations in the rpoB gene of Mycobacterium tuberculosis (Mtb). Most resistance-conferring mutations occur within the 81-base pair RIF resistance determining region (RRDR), particularly at codons S450L, H445Y/D, and D435V, which are strongly associated with high level resistance. However, increasing evidence of low-frequency and disputed variants both within and beyond the RRDR reveals a broader genetic spectrum that contributes to diagnostic uncertainty and variable phenotypic outcomes. This review summarizes current knowledge of high frequency, low frequency, and disputed rpoB mutations and their implications for molecular detection of RIF resistance. Structural analyses show that specific amino acid substitutions alter key hydrogen bonds or create steric hindrance in the RIF-binding pocket, leading to diverse resistance levels. Despite the success of molecular platforms such as Xpert MTB/RIF and line probe assays, their hotspot-based detection limits sensitivity to noncanonical variants. Lowering the minimum inhibitory concentration (MIC) breakpoint and integrating sequencing-based approaches, such as targeted and whole-genome sequencing, can enhance detection accuracy. A combined genomic and phenotypic framework will be essential to close existing diagnostic gaps and advance precision guided management of RIF-resistant and multidrug-resistant tuberculosis. Full article

The endophytic Streptomyces sp. PRh3 (PRh3), isolated from Dongxiang wild rice (DXWR), exhibited impaired biosynthetic capacity in the laboratory. To address this defect, rifampicin-based ribosome engineering was first applied to PRh3 to generate PRh3-r55, which acquired a characteristic H473Y rifampicin-resistant mutation in rpoB to activate the production of two β-carboline alkaloids JBIR-133 and JBIR-134. Then the biosynthetic gene cluster (BGC) ksl was introduced into PRh3-r55 for heterologous expression, generating PRh3-r55K. This combined approach achieved a synergistic effect, enabling the strain to produce not only the expected JBIR-133, JBIR-134, and kitasetaline, but also a novel sulfur-containing molecule, novkitasetaline. Structural elucidation identified novkitasetaline as a unique tryptamine-substituted kitasetaline derivative at the C-3 position of its pyridine ring. Notably, this structural modification conferred significant antimalarial activity to novkitasetaline, rendering it active against drug-sensitive Plasmodium falciparum 3D7 (IC₅₀ = 32.65 ± 2.93 μM) and three other drug-resistant P. falciparum strains: K13^C580Y, Dd2, and HB3 (IC₅₀ = 45.98 ± 4.17~59.67 ± 3.15 μM), primarily by disrupting late-stage parasite development. These efforts not only identified a promising antimalarial lead compound but also demonstrated that combining ribosome engineering with heterologous expression is an effective strategy for discovering bioactive natural products from Streptomyces. Full article

Clostridium perfringens is an opportunistic Gram-positive bacterium that causes necrotic enteritis and other severe infections in animals, as well as food poisoning in humans. In this study, we introduce a framework consisting of rpoB sequence typing (RSTing) and network analysis to investigate the evolutionary trajectories of C. perfringens. By analyzing 319 rpoB sequences—300 from public databases and 19 newly sequenced isolates from chicken and cattle sources—we identified 84 rpoB sequence types (RSTs). Among them, the early emerging RST 1-1 was the most prevalent (21.3%), while the putative ancestral type, RST 0, was the fifth most common (4.7%). The high RST diversity and the predominance of RST 1-1, mainly from chickens, suggest that chickens may serve as an important reservoir. By integrating virulence gene profiling, MLST, and comparative genomics, we separated identical RSTs into distinct genotypes and uncovered genomic evidence of possible interspecies transmission between chickens and cattle, two major food-producing species. These findings indicate that RSTing provides a useful complementary approach to investigating the evolutionary and epidemiological dynamics of C. perfringens. Full article

Previous studies have shown that AtMP1 and AtMP2, antimicrobial peptides found in the climbing perch (Anabas testudineus), possess antibacterial and anticancer properties. This study aimed to determine whether modified forms of the AtMP2 peptide could enhance its antimicrobial activity. In the research study, the epidermal mucus of Anabas testudineus was extracted and tested to contain antibacterial and anticancer properties. Two peptides were initially extracted from the freshwater fish Anabas testudineus; however, the focus was placed on AtMP2 to evaluate its potential for enhanced antibacterial activity. Systematic-directed evolution was used to generate AtMP2 varieties. The AtMP2 varieties were characterized using APD3, CAMP, and AMPFun. Based on the characterization, two peptides, AtMP2-1 and AtMP2-2, were selected for synthesis. AtMP2-1 and AtMP2-2 demonstrated higher antimicrobial activity against Gram-positive than Gram-negative bacteria compared to the AtMP2 peptide, based on Minimum Inhibitory Concentration (MIC) determination and Kirby–Bauer Disk Diffusion analysis. For the SRB cytotoxicity analysis using the HS-27 and Vero cell lines, both AtMP2-1 and AtMP2-2 are safe for human use at 20 μg/mL. ZDOCK and HPEPDOCK servers were used to evaluate the binding affinity between AtMP2-1 and AtMP2-2 peptides and proteins involved in the bacterial cell death cycle. The results showed varying docking scores, with more negative values indicating stronger binding interactions, particularly with proteins such as SecA, RpoB, GyrA, ClpP, and MetG. The study concluded that modified peptides derived from Anabas testudineus show enhanced antibacterial properties and minimal cytotoxicity, making them potential alternatives to traditional antibiotics. Future research should investigate the specific biochemical pathways affected by these AMPs to understand their mechanisms of action better and explore their potential as therapeutic agents, particularly in the treatment of bacterial infections, wound healing, and cancer therapy. Full article

The aim of this study was to identify the causative agent of bacterial blight and basal glume rot of winter wheat that appeared in Serbia in 2023. To characterize the isolated bacteria (eight isolates in total), their cultural, biochemical, pathogenic, and genetic characteristics were examined. Based on the results of the LOPAT test, the isolates were classified into Pseudomonas Group Ia. The syrB and syrD genes were simultaneously detected in six wheat isolates—P0123, P0223, P0323, P0423, P0523, and P0823—while two isolates, P1123 and P1323, lacked both genes. Multilocus sequence typing (MLST) of the gapA, gltA, gyrB, and rpoD genes identified six isolates (P0123, P0223, P0323, P0423, P0523, and P0823) as Pseudomonas syringae pv. atrofaciens, whereas the remaining two isolates (P1123 and P1323) were most closely related to P. poae. Phylogenetic analysis revealed three genetically heterogeneous subgroups of P. syringae pv. atrofaciens among the wheat isolates from Serbia. Pathogenicity tests demonstrated that wheat isolates are able cause disease on wheat seedlings using three different inoculation methods: spraying the entire seedling, trimming the leaves before spraying, and wounding the leaves with multiple needles followed by spraying. Overall, isolates P0123 and P0423 were identified as the most virulent, inducing pronounced blight symptoms on wheat seedlings. In contrast, isolates P1123 and P1323 were weakly virulent and are therefore considered to be secondary or accompanying factors in plants already infected with more aggressive isolates, rather than primary pathogens responsible for disease development. This study contributes to a deeper understanding of the ecology, distribution, and pathogenic potential of bacterial communities associated with wheat blight disease in Serbia. Full article

Livestock is the backbone of the economy in an agricultural country like Pakistan, with cattle serving as a milk and protein source. In 2021/22, Pakistan was hit by the first major outbreak of lumpy skin disease (LSD) in cattle, in all four provinces. LSD is characterized by the development of skin nodules, leading to severe illness, decreased milk production, and mortality, causing huge economic losses. This study aimed to analyze and classify the lumpy skin disease virus (LSDV) strains involved in the outbreak in the Punjab province at the molecular and phylogenetic levels to develop effective control strategies. A combination of different real-time PCRs was used for the identification and differentiation between vaccine, wild-type, and recombinant LSDV strains. This was mented with the sequence determination and phylogenetic analysis of ten genomic loci from two selected isolates from the 2021/22 Pakistan outbreak. The combined data showed that these isolates belonged to LSDV clade 1.2 and were clearly different from the vaccine clade 1.1 (Neethling-like), as well as from the recombinant clade 2 strains. In addition, using a fit-for-purpose gel-based PCR, the isolates from the outbreak were also shown to be different from KSGP0240-based vaccines. Full article

Tomato is one of the most important vegetable crops in Costa Rica, where favorable environmental conditions enabled year-round production but also promote bacterial diseases. In recent years, pith necrosis has been frequently observed; nevertheless, the causal agents remain unidentified in the country. This study evaluated bacteria associated with symptomatic plants collected in the Central Valley of Costa Rica. From 32 plants, 61 bacterial isolates were obtained, described morphologically, and characterized through basic biochemical tests. Partial sequencing of the 16S rRNA gene revealed diverse bacterial taxa, predominantly belonging to the genus Pseudomonas. Thirteen isolates were selected for pathogenicity assays, which confirmed variable virulence levels. Multilocus sequence analysis based on concatenated sequences of the 16S rRNA, gyrB, rpoD, and rpoB genes identified Pseudomonas alliivorans LTM 13.1.2, P. flavescens LTM 14.2.2, and P. capsici LTM 78.3.2 as causal agents of pith necrosis. Additionally, P. straminea LTM 78.2.1 and Cedecea sp. LTM 72.2.1 caused tissue discoloration. Whole-genome sequencing of the two most virulent isolates (LTM 13.1.2 and LTM 78.3.2) supported their taxonomic classification and revealed virulence-associated genes and biosynthetic clusters. This study represents the first report of these Pseudomonas species as tomato pathogens in Costa Rica and expands their known distribution and host ranges. Full article

Enterococcus faecalis (E. faecalis) is a significant zoonotic pathogen, primarily causing opportunistic infections in humans while often existing as a commensal in animal reservoirs, facilitating its dissemination. Current understanding of the resistance profiles, virulence mechanisms, and host–pathogen interactions of E. faecalis from ruminants, particularly unique species such as the plateau yak, remains limited. This knowledge gap hinders the accurate assessment of their transmission risk and the development of effective control strategies. This study presents a comprehensive analysis of a multidrug-resistant E. faecalis isolate from yak feces, integrating whole-genome sequencing (WGS), an animal challenge model, and transcriptomic profiling. Antimicrobial susceptibility testing revealed resistance to β-lactams, aminoglycosides, glycopeptides, tetracyclines, and fluoroquinolones. WGS identified numerous resistance genes (e.g., parC, gyrA, rpoB) and virulence-associated genes (e.g., prgB/asc10, cpsA/uppS). Phylogenetic analysis indicated a close relationship with a human urinary tract isolate (ASM3679337v1). Mouse challenge experiments demonstrated that this strain induced significant intestinal histopathological damage. A subsequent transcriptomic analysis of infected tissues identified the differential activation of key signaling pathways, including NF-κB and MAPK. Our findings provide crucial insights into the resistance and pathogenic mechanisms of ruminant-derived E. faecalis and establish an experimental foundation for optimizing clinical antimicrobial therapy against such strains. Full article

Bacteria are promising sources of bioflocculants, yet their regulatory machinery for bioflocculant synthesis remains underexplored. This study focused on evaluating the biosynthetic genes, optimisation and assessment of bioflocculation efficiency in wastewater. The isolated bioflocculant producers were identified by 16S rRNA and rpoB gene analysis. Polymerase chain reaction was used to assess the presence of polyketide synthase I (PKS-1), polyketide synthase II (PKS-II), non-ribosomal peptide synthetase (NRPS), epsH and epsJ. A one-factor-at-a-time technique was utilised for optimisation of culture conditions. The bioflocculants’ efficiencies were evaluated in wastewater using the Jar test method. Among 31 isolates, Klebsiella michiganensis and Klebsiella pasteurii were the most potent bioflocculant producers. They both revealed the presence of PKS-II. K. pasteurii possessed the epsH gene. The optimal conditions for maximum bioflocculant production (95% activity) by K. michiganensis were a temperature of 35 °C, pH of 5, galactose, tryptophan and 84 h of incubation. K. pasteurii’s maximum bioflocculant production of 83% was obtained at a temperature of 35 °C and pH of 7, with galactose, a mixture of urea, yeast extract, and ammonium sulphate (NH₄)₂SO₄ and 96 h of fermentation. Their bioflocculants reduced the chemical oxygen demand and turbidity of wastewater by more than 70%. The bacteria had promising bioflocculant production with potential applicability in wastewater treatment. Full article

Tuberculosis (TB), caused by Mycobacterium tuberculosis, continues to be a leading cause of morbidity and mortality in Mexico, with more than 20,000 new cases annually and a rising proportion of drug-resistant strains. This work addresses the molecular epidemiology of TB in the Mexican context, emphasizing its role in understanding transmission, genetic diversity, and resistance mechanisms. To achieve this, we reviewed molecular typing approaches including spoligotyping, Mycobacterial Interspersed Repetitive Unit–Variable Number Tandem Repeat (MIRU-VNTR) analysis, and whole-genome sequencing (WGS), which have been applied to characterize circulating lineages and identify drug-resistance-associated mutations. The results indicate that the Euro-American lineage (L4) predominates across the country, although significant regional variation exists, with Haarlem, LAM, T, and X sub lineages dominating in different states, and occasional detection of Asian (L2) and Indo-Oceanic (L1) lineages. Key resistance mutations were identified in katG, rpoB, pncA, and gyrA, contributing to the emergence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains, particularly in border and marginalized regions. These findings highlight how social factors, such as migration, urban overcrowding, and comorbidities including diabetes and HIV, influence transmission dynamics. We conclude that integrating molecular tools with epidemiological surveillance is crucial for strengthening public health strategies and guiding interventions tailored to Mexico’s heterogeneous TB burden. Full article

Records of Sarcocystis spp. diversity in rodents are relatively extensive; however, the increasing application of molecular approaches indicates that our current knowledge of these parasites remains incomplete. In the present study, morphological, genetic and phylogenetic data are provided on Sarcocystis arvalis n. sp. from the common vole (Microtus arvalis). Using light microscopy, the observed sarcocysts had a relatively thin (<1 μm) and smooth cyst wall. Via transmission electron microscopy, the sarcocyst wall thickness ranged from 0.7 to 1 μm, and the parasitophorous vacuolar membrane exhibited small knob-like blebs and was slightly wavy, type 1a. Based on 18S rRNA, 28S rRNA, cox1 and rpoB loci, S. arvalis n. sp. showed the highest similarity with Sarcocystis myodes from the bank vole (Clethronomys glareolus). According to the phylogenetic placement, S. arvalis n. sp. is the most closely related to Sarcocystis spp. with a rodent–mammal lifecycle. Morphologically, S. arvalis n. sp. forms sarcocysts that share a lot of similarities with those of S. myodes, Sarcocystis ratti and Sarcocystis cernae and molecular analysis is necessary for accurate species identification. Based on the abundance of the hosts and the proportion of voles in their diet, the most likely definitive hosts of S. arvalis n. sp. are red foxes, stone martens, least weasels, and domestic cats. Full article

Upon reacting with cellular components, Hg(II) ions elicit the production of reactive oxygen species (ROS). While the ROS-promoted cytotoxic and genotoxic effects induced by Hg(II) have been widely described in eukaryotes, such effects have been less studied in bacteria. In this work, the prokaryotic environmental model Bacillus subtilis was employed to evaluate the cytotoxic and genotoxic impact of Hg(II) over strains proficient or deficient in SOS, general stress and antioxidant responses, as well as the global transcriptional response elicited by this ion. The exposure to HgCl₂ significantly increased the mutation frequency to rifampicin resistance (Rif^R) in WT and mutant strains, suggesting a major contribution of these pathways in counteracting the genotoxic effects of Hg(II). Detection of A → T and C → G transversion mutations in the rpoB gene of Hg(II)-exposed cells suggested the generation of 8-oxo-guanines (8-OxoGs) and other oxidized DNA bases. The RNA-seq study revealed upregulation of genes involved in efflux and/or reduction of metal ions, synthesis of sulfur-containing molecules, and downregulation of genes implicated in iron metabolism and cell envelope stress. Therefore, our results indicate that metal extrusion and scavenging of Hg(II) by thiol-rich molecules may constitute a line of defense of B. subtilis that counteracts the noxious effects of ROS resulting from an imbalance in iron metabolism elicited by this ion. Full article

In recent years, the cultivated area of hazelnuts in Chile has increased significantly. Along with this rapid expansion, biotic constraints that affect the optimal development of the crop have been identified. Among these, bacterial blight disease caused by Xanthomonas arboricola pv. corylina has been particularly relevant. This pathogen has a global distribution and is present in all hazelnut-producing countries. In the spring of 2023, hazelnut orchards were sampled from the Maule to Biobío Regions of Chile. The Chilean isolates recovered from hazelnut tissues showing symptoms of bacterial blight were characterized by their ability to grow on different semi-selective media, their carbohydrate utilization profiles, hypersensitivity response in tobacco plants, and biochemical tests. Additionally, the isolates were identified based on the 16S rRNA gene and multilocus sequence analysis (MLSA) on the rpoD, gyrB, and atpD genes. The results showed that the X. arboricola pv. corylina Chilean isolates differed from previously reported isolates in other geographic areas as they are capable of metabolizing sorbitol and mannitol. Using MLSA and average nucleotide identity (ANI) comparison, these isolates were grouped into four and five phylogenetic clades, respectively, representing a significant difference from what has been reported in similar international studies. Full article