Microbial Genomics最新文献

Clostridioides difficile infection (CDI) imposes a substantial clinical burden in paediatric populations. However, the high prevalence of asymptomatic colonization, especially in children under 2 years of age, complicates the distinction between true infection and non-pathogenic carriage. This diagnostic uncertainty hinders appropriate treatment decisions and complicates infection prevention efforts. A 6-year retrospective cohort study was performed at Shanghai Children's Hospital to characterize the epidemiology and clinical features of CDI and asymptomatic colonization in paediatric patients. Stool specimens were cultured for C. difficile, and isolates underwent whole-genome sequencing to perform multilocus sequence typing, identify SNPs and characterize functional gene content via Clusters of Orthologous Groups (COG) analysis. Mutations in genes associated with toxin production were analysed to assess genetic differences between clinical isolates from infected patients and asymptomatic carriers. In addition, comparative genomic analysis was performed to assess variations in virulence-associated genes, antimicrobial resistance (AMR) genes and genes involved in quorum sensing (QS). Colonization factors (CFs) were also characterized to elucidate potential mechanisms differentiating asymptomatic colonization from symptomatic infection. And we conducted in vitro experiments on toxin B-variant strains. A total of 23 sequence types (STs) were identified among isolates from 39 asymptomatic carriers and 61 symptomatic patients, with greater ST diversity observed in the infection group compared to the colonization group. COG analysis demonstrated an increased representation of uncharacterized functional categories in the infection group, suggesting a potential role for novel genes in pathogenesis. Patterns of virulence factor presence, AMR genes and QS gene distribution were comparable between the two groups, as were mutations in toxin regulation genes. Notably, six isolates belonging to ST37 and ST81, characterized by the absence of tcdA and presence of tcdB, exhibited a high frequency of mutations. In vitro experiments demonstrated that these strains exhibited higher biofilm formation capacity and elevated transcriptional levels of both tcdB and spoA. Additionally, no significant differences were detected in the distribution of CFs. Our findings contribute to the growing understanding of the genomic determinants and their functional roles underlying paediatric CDI severity, providing more evidence for improved diagnostic and therapeutic strategies.

Carbapenem resistance in Pseudomonas aeruginosa is a growing public health concern. Multiple mechanisms of antimicrobial resistance have been described. While surveillance often focuses on carbapenemase detection, non-carbapenemase mechanisms and their interplay with the genomic background remain underexplored. This study aimed to characterize how genomic background influences carbapenem resistance mechanisms and adaptive responses in carbapenem-resistant P. aeruginosa (CRPA). A total of 136 CRPA clinical isolates collected from 28 healthcare centres across Mexico were analysed through core genome phylogeny, sequence type (ST) assignment, resistome profiling, oprD variant analysis, bacterial growth kinetics under imipenem and meropenem exposure and qRT-PCR-based expression of oprD, mexA, mexC, mexE and mexY genes. Isolates clustered into PAO1 and PA14 phylogroups. ST309 was the most prevalent ST (29/136). Pseudomonas paraeruginosa lineage was also identified within these isolates. Phylogenetic clustering of antibiotic resistance genes was observed across phylogroups. In 36% (49/146) of the isolates, β-lactamases (bla _VIM 16%, bla _GES11% and bla _IMP 11%) were identified with carbapenemase activity previously reported. Potentially inactivating oprD variants were observed in 75% (102/136) of isolates, with nonsense and frameshift variants associated with resistance phenotypes. Isolates harbouring carbapenemase-encoding genes (CEGs) exhibited stable lag phases regardless of antibiotic exposure, whereas isolates lacking CEGs showed significantly prolonged lag phases. Overexpression of mexA, mexC and mexY genes was observed in 39% (7/18), 17% (3/18) and 39% (7/18) of isolates, respectively, under antibiotic-free condition, and increased under carbapenem exposure. mexA expression was significantly higher in PAO1 isolates (6/18) under antibiotic-free condition and in PA14 isolates under imipenem exposure (5/18). Carbapenem resistance in P. aeruginosa is shaped by both phylogenetic background and antibiotic-driven stress responses. This study provides an integrated analysis of resistance mechanisms - including gene expression and physiological adaptation - across major phylogenetic lineages in clinical isolates recovered in Mexico, underscoring the importance of considering non-carbapenemase resistance pathways in surveillance and treatment strategies.

Members of the class Clostridia, a polyphyletic group of pathogenic and beneficial Gram-positive, spore-forming anaerobes in the Bacillota (Firmicutes) phylum, are prevalent in the human gut. While this class includes select pathogens known to cause disease, many species are associated with beneficial functions, such as providing colonization resistance against pathogens. Despite a demonstrated value in maintaining Clostridial populations in the gut, functional strain diversity of most commensal Clostridial species remains understudied. Here, we isolated and characterized Clostridial isolates, focusing on the genomic diversity of Anaerostipes, a prevalent butyrate-producing genus within the gut microbiota. We conducted a genomic comparison across 21 Anaerostipes strains isolated from healthy human faecal samples (n=5) and publicly available genomes (n=105). Whole genome comparisons across the Anaerostipes genus demonstrated 12 species bins, clustering into three major functionally distinct clusters correlating with host origin. One cluster (representing mostly Anaerostipes caccae genomes) was distinguished by possessing a complete vitamin B12 biosynthesis pathway. Variability in genomic and phenotypic carbohydrate metabolism was demonstrated within dominant species of the human microbiota (Anaerostipes hadrus, A. caccae and Anaerostipes hominis). Collectively, these data indicate genomic metabolic variance across Anaerostipes species that may influence coexistence within the gut environment and variably influence health.

ssDNA viruses are important components of diverse ecosystems; however, it remains challenging to systematically identify and classify them. This is partly due to their broad host range and resulting genomic diversity, structure and rapid evolutionary rates. In addition, distinguishing genuine ssDNA genomes from contaminating sequences in metagenomic datasets (e.g. from commercial kits) has been an unresolved issue for years. Here, we present CRESSENT (CRESS-DNA Extended aNnotation Toolkit), a comprehensive and modular bioinformatic pipeline focused on ssDNA virus 'genome-to-analysis' and annotation. The pipeline integrates multiple functionalities organized into several modules: sequence dereplication, decontamination, phylogenetic analysis, motif discovery, stem-loop structure prediction and recombination detection. Each module can be used independently or in combination with others, allowing researchers to customize their analysis workflow. With this tool, researchers can comprehensively and systematically include ssDNA viruses in their viromics workflows and facilitate comparative genomic studies, which are often limited to dsDNA viruses, therefore leaving behind a crucial component of the microbiome community under study. Benchmarking analyses demonstrated that CRESSENT efficiently processes ssDNA virus datasets of varying scales, completing small family-level analyses within minutes and moderate comparative genomics studies within hours using standard computing resources. Its modular, parallelized design ensures scalability and low memory usage, making it accessible to research groups with diverse computational capacities.

To determine changes in the pneumococcal serotypes, sequence types (STs), clonal complexes (CCs) and the frequency of antimicrobial resistance genes after the introduction of pneumococcal conjugate vaccines (PCVs) in Lima, Peru. Retrospective multicentre study analysing whole-genome sequencing (WGS) data from three passive surveillance studies of invasive pneumococcal disease (IPD) in paediatric patients in Lima (2006-2020). Pneumococcal typing and antimicrobial resistance were analysed using in silico genomic tools. CCs were identified with eBURST and phylogenetic results were visualized using PHYLOViZ. 262 pneumococcal isolates were analysed (104 from IPD1, 70 from IPD2 and 88 from IPD3), 55.3% from children under 2 years old, 53.1% from patients with pneumonia and 28.5% with meningitis. After the introduction of PCVs, vaccine serotypes decreased, while serotype 19A and non-vaccine serotypes increased. The predominant STs were ST156 in IPD1 (n=25) and in IPD2 (n=7); and ST320 (n=38) and ST230 (n=15) in IPD3. Sixteen CC were identified, the most frequent were CC1421 (n=58) and CC156 (n=36). The overall penicillin non-susceptibility (NS) increased from 21.8% in IPD1 to 28.6% in IPD3, ceftriaxone-NS increased from 10% to 13.1% and macrolide-NS from 24.8% to 85.7% respectively. Resistance markers for macrolides, tetracycline and cotrimoxazole increased post-PCV13. WGS predicted antimicrobial resistance with high concordance, though some discrepancies were noted with phenotypic testing methods. Important changes in the distribution of serotype and ST, especially among vaccine serotypes, have been observed. These findings highlight the importance of monitoring vaccine effectiveness and tracking changes in bacterial populations to guide future vaccine implementation.

Pantoea allii, one of four Pantoea species known to cause onion centre rot, is infrequently isolated from onion compared to its closely related onion-pathogenic species in the same genus. To better understand the genomic diversity and genetic determinants of pathogenicity in this species, we analysed a collection of 38 P. allii strains isolated from 2 primary ecological niches, plants and rainwater, across North and South American and African continents using comparative genomics and phylogenetic approaches. Core-genome phylogeny, average nucleotide identity and gene presence-absence analyses revealed three genetically distinct lineages. All strains harboured conserved biosynthetic gene clusters (BGCs) for quorum sensing, carotenoid production, siderophores and thiopeptides. In contrast, two phosphonate BGCs, key determinants of onion pathogenicity, exhibited lineage-specific distributions. Onion-associated strains from lineages 1 and 2 carried the Halophos BGC responsible for onion tissue necrosis and also encoded the alt gene cluster that confers tolerance to thiosulfinates. Lineage 3 strains, isolated from both onion and rainwater, either lacked a phosphonate BGC entirely or possessed the HiVir phosphonate BGC. In addition, lineage 3 strains lacked the alt cluster altogether. The localization of these virulence genes in the genome varied, with Halophos integrated in the chromosome, HiVir encoded on the conserved Large Pantoea Plasmid, and alt located on small, variable plasmids (plasmid B). The type IV secretion system (T4SS) and type VI secretion system (T6SS) showed variable genomic architectures, with plasmid-borne T4SSs and two chromosomal T6SS loci differing in conservation and gene content. Additionally, conserved Pantailocin phage islands were detected in most genomes. Overall, this study reveals that while core metabolic and competitive traits are conserved across P. allii, virulence-associated loci display lineage-specific distribution, reflecting ecological differentiation and evolutionary plasticity within the species.

Salmonella enterica subspecies enterica serotype Minnesota (S. Minnesota) has recently emerged as a predominant serotype in poultry farming operations. Genomic surveillance efforts concentrated primarily in Europe have been used to evaluate food safety risks associated with S. Minnesota in imported poultry/poultry products. However, the burden imposed by S. Minnesota on consumers in sub-Saharan Africa is not understood. Here, we used whole-genome sequencing to characterize 36 S. Minnesota strains from raw poultry imported into South Africa, specifically (i) 11 strains isolated at port of entry and (ii) 25 strains from imported poultry in South African supermarkets. While all 36 strains belonged to the same sequence type (ST548), multiple ST548 lineages were present among poultry products. Comparison of the 36 strains sequenced here to all publicly available, high-quality ST548 genomes (n=460, from Enterobase) identified several public genomes differing by <30 core SNPs, including strains isolated previously from South American poultry imported into the UK. Notably, a cluster consisting of 14 highly similar genomes sequenced here (0 core SNPs) uniquely possessed bla _CTX-M-8. A search of plasmids in public databases, alongside antimicrobial resistance (AMR) genes from >1.9 million bacterial genomes, revealed that this cluster harboured bla _CTX-M-8 on an IncI1 plasmid-like region, which we hypothesize was acquired recently, from Escherichia coli. Overall, our study provides insight into the intercontinental dissemination of S. Minnesota and its associated AMR determinants via the global poultry trade.

Genetic exchange between Neisseria meningitidis (NM) and Neisseria gonorrhoeae (NG) has not been well studied. This study aimed to investigate evidence of genetic exchanges between these two species. All coincident paired NM and NG isolates cultured from pharyngeal swabs collected from a sexual health clinic in Sydney in 2021 underwent whole-genome sequencing. A gene-by-gene analysis of the 47 NM-NG pairs identified 184 instances where the ancestry of the gene revealed intermixing between the two species. Incorporating the gene phylogenies demonstrated that these events occurred across a wide range of timeframes. At the nucleotide level, 91 genes were found where paired isolates harboured identical sequences. Notably, one instance of unequivocal recent gene transfer events between the paired pharynx isolates was observed. This work provides new insights into the evolutionary dynamics of these bacteria and highlights the importance of genetic exchange in populations with high rates of pharyngeal gonorrhoea. The clinical implications of such exchanges call for continued vigilance and research to address the challenges posed by these bacteria.

Cleaning and disinfection of food production environments (FPE) are fundamental components of food safety programmes designed to control microbial pathogens and prevent food contamination. Yet, FPE can still harbour foodborne pathogens, including Listeria monocytogenes, a significant concern to food manufacturers and health authorities due to the high mortality rate associated with invasive listeriosis. Mechanisms contributing to L. monocytogenes persistence in FPE include biofilm formation and reduced susceptibility to biocides, such as benzalkonium chloride (BC), for which several mechanisms are known. We hypothesized that prolonged exposure to disinfectants and other FPE-associated stressors would drive L. monocytogenes adaptation, resulting in the accumulation of genetic mutations linked to biofilm formation and reduced biocide susceptibility. To test this, we developed a biofilm persistence model, which studied 30 consecutive passages of biofilm-associated cells grown on stainless steel under sub-inhibitory BC concentrations. Whole-genome sequencing of evolved populations identified mutations that were associated with biofilm lineages and/or BC exposure. Non-synonymous mutations were identified in genes and pathways involved in metal homeostasis, stress response and pyrimidine biosynthesis. In addition, reduced susceptibility to BC arose through multiple independent mutations within the fepRA operon, encoding FepR transcriptional repressor and FepA MATE efflux pump. These mutations were observed across both planktonic and biofilm lifestyles, resulting in a comparable level of reduced susceptibility to BC in both states. Several loci with fixed mutations associated with biofilm lineages were identified, including the ykoK riboswitch leader, the pyrimidine synthesis operon and the stress response-related gene rsbU. Collectively, these findings provide new insights into the genetic mechanisms underlying L. monocytogenes biofilm persistence and reduced biocide susceptibility in the context of FPE and reveal novel targets potentially exploited by L. monocytogenes to establish and maintain niches in unfavourable environments.