Microbial Genomics最新文献

We report the presence of the emerging plant pathogen 'Xanthomonas cannabis' in Australia through a comprehensive analysis of five historical isolates and all publicly available genomes of the species. Using comparative genomics, we characterized four isolates collected from Zinnia spp. and one from Cucurbita pepo. Our findings show that the Zinnia isolates form a distinct phylogroup with the pathotype strain of 'X. cannabis' pv. zinniae. This group possesses genes for the type 3 secretion system (T3SS) and effectors, a variety of genes unique within the species, and nine genomic islands associated with virulence and drug resistance. In contrast, the C. pepo isolate is genetically distinct and lacks the T3SS but contains its own genes unique within the species. Hypersensitivity response assays confirmed the pathogenic potential of all five isolates in black bean, eggplant, green bean, tomato, sunflower, zinnia and zucchini plants. These results highlight the genetic diversity and evolving threat of this pathogen in Australia, underscoring the critical need for ongoing biosecurity surveillance.

Wolbachia is a ubiquitous endosymbiont in arthropods that is maternally transmitted and affects host reproduction in various ways. Among these, skewing the host sex ratio towards females, either by killing males (male killing) or producing exclusively functional females (feminization or parthenogenesis), is considered advantageous for Wolbachia. In the butterfly Eurema mandarina, individuals harbouring the Wolbachia strain wFem exclusively produce female offspring. This occurs through a two-step mechanism in which Wolbachia blocks the transmission of the Z chromosome from Z0 females and feminizes the resultant Z0 offspring. Given the unique characteristics of wFem, understanding its genomic features is crucial to uncover the evolution and mechanisms of Wolbachia-induced reproductive manipulation. However, technical challenges in isolating wFem from co-infecting, closely related, non-male-killing/non-feminizing wCI Wolbachia strain have hindered genomic analyses of wFem. In this study, we established a closed circular genome of wFem by developing a series of Wolbachia purification, cell sorting and single-cell genome sequencing techniques. wFem genome, ~1.3 Mb in size, specifically encodes male-killing gene homologues (Em-oscar and wmk) and other putative virulence factors that are absent in wCI. In addition, wFem carried prophage elements that showed high similarity to previously characterized male-killing-associated prophages in Wolbachia strains. This study highlights the shared functional genomic features between feminizing and male-killing Wolbachia in Lepidoptera and suggests a mechanistic link between these two Wolbachia-induced reproductive phenotypes.

Pathogens adapting to the human host and to vaccination-induced immunity may follow parallel evolutionary paths. Bordetella parapertussis (Bpp) contributes significantly to the burden of whooping cough (pertussis) and shares vaccine antigens with Bordetella pertussis (Bp); both pathogens are phylogenetically related and ecological competitors. Bp vaccine antigen-coding genes have accumulated variation, including pertactin (PRN) disruptions, after the introduction of acellular vaccines in the 1990s. We aimed to evaluate evolutionary parallelisms in Bpp, even though pertussis vaccines were designed against Bp. We sequenced 242 Bpp isolates collected in France, the USA and Spain between 1937 and 2019, spanning pre-vaccine and two vaccines eras. We investigated the temporal evolution of Bpp sublineages using a Bayesian approach based on whole-genome SNPs and performed comparative genomic analyses focusing on antigen and virulence gene loci. The most recent common ancestor of all sequenced Bpp isolates was estimated around the year 1877, making it one of the youngest human pathogens, and the Bpp evolutionary rate we estimated (2.12×10^-7 substitutions per site per year) was remarkably similar to the one previously reported for Bp (2.24×10^-7). PRN antigen deficiency in Bpp was driven by 18 disruptive mutations, including deletion prn:ΔG-1895 estimated to have occurred around 1998 and observed in 73.8 % (149/202) of post-2007 Bpp isolates. In addition, we detected two early (year ~1900) mutations in the bvgA-fhaB intergenic region, which controls multiple virulence factors including the filamentous haemagglutinin antigen. Gene clusters for pertussis toxin and fimbriae showed a surprising lack of gene decay. Our findings suggest early adaptation of Bpp to humans through modulation of the bvgAS regulon, and a rapid adaptation through the loss of PRN expression, representing a late evolutionary parallelism concomitant with acellular vaccination against whooping cough.

Mycobacterium avium subsp. hominissuis (MAH) is a clinically important species of non-tuberculous mycobacteria that causes infections in a variety of hosts. This opportunistic pathogen is widespread in the environment, including natural and engineered water systems across the globe. To examine the current genetic diversity of this organism, we analysed 702 MAH genomes isolated from humans, pigs and environmental sources across six countries and three continents. Through Bayesian population structure analysis, we identified ten distinct global lineages, including seven previously described and three new ones. Several lineages exhibited regional clustering, including sequence cluster 6 (SC6) in Germany and SC8 in both the USA and Germany. Further analysis identified recombination hotspots in membrane proteins associated with virulence, antimicrobial resistance and immune modulation, driven by insertion sequences and other elements that frequently integrate at tRNA gene sites. This work demonstrates the remarkable genomic diversity of MAH and provides insight into the evolutionary mechanisms that contribute to its success as a pathogen in both humans and animals.

Background. Oxford Nanopore Technologies (ONT) enables direct methylome profiling using the Dorado basecaller, but the comparative performance of ONT-only versus hybrid-assembly reference-based methylation calling, particularly regarding inter-operator variability, remains understudied.Methods. Six operators independently prepared 15 sequencing libraries for nanopore (MinION R10.4.1 flow cells, Mk1D) and Illumina MiniSeq platforms for two Streptococcus dysgalactiae subsp. equisimilis strains (UT9728, 12 replicates; UT10237, 3 replicates). Methylation and motif profiles were identified with MicrobeMod v.1.0.3 using both Illumina-corrected hybrid reference assemblies (HRAs) and ONT-only reference assemblies (ORAs). A custom genome annotation-aware pipeline mapped methylation site calls to coding sequence, rRNA and tRNA features for reproducibility analysis. We also performed site-wise analyses to quantify concordance of methylated fractions among ORA replicates and assess the influence of sequencing coverage.Results. Strain UT9728 predominantly exhibited N6-methyladenine (6mA) at G6mATC motifs, whereas strain UT10237 displayed dual methylation patterns: C5-methylcytosine (5mC) at 5mCCWGG motifs and 6mA at G6mAGNNNNNTAA motifs. Motif identification concordance using HRAs and ORAs exceeded 99.9%. Reproducibility across replicates was high for G6mATC and 5mCCWGG motifs in both HRAs (Pearson's r>0.989) and ORAs (Pearson's r>0.993), but lower for the degenerate G6mAGNNNNNTAA motif (Pearson's r: HRA=0.80; ORA=0.78). ORA-based methylation site calls showed excellent precision and recall compared to HRA-based calls (F1-score>99.999%). Site-wise analysis of UT9728 G6mATC motifs revealed that discordant sites (absolute methylated fraction difference ≥0.15) were rare, with 44 of 66 pairwise replicate comparisons showing <1% discordance. Discordance was linked to low coverage (<70×), whereas sites sequenced >200× displayed complete concordance.Conclusion. Although limited to a single species, three motifs and a feature-based framework that does not capture promoter-proximal events, ONT-only sequence-based methylome profiling proved accurate and reproducible across multiple operators, with sequencing coverage emerging as the principal determinant of site-level concordance.

Polyomaviruses and papillomaviruses are icosahedral viruses with small circular dsDNA genomes. Limited information on their diversity and evolution in avian hosts is available, with even less known regarding Antarctic penguins. Prior to this study, only one polyomavirus and two papillomaviruses had been identified in Adélie penguins (Pygoscelis adeliae). To expand our knowledge of these viruses in Antarctic penguins, we collected faecal and cloacal swab samples from 246 Adélie penguins over 3 breeding seasons (2021-2024) and 10 emperor penguins (Aptenodytes forsteri) during the 2023-2024 season on Ross Island (Ross Sea). Additionally, we sampled 66 Adélie, 40 chinstrap (Pygoscelis antarcticus) and 71 gentoo (Pygoscelis papua) penguins during the 2022-2023 season across various sites on the Antarctic Peninsula. All samples were screened for papillomaviruses and polyomaviruses. We identified 31 polyomaviruses in Adélie, gentoo and chinstrap penguins and 4 papillomaviruses in Adélie penguins sampled in both eastern and western Antarctica. The 31 penguin polyomaviruses belong to a single species but form four distinct variants that are host species specific with strong geographic clustering. The four papillomaviruses represent three different types, of which two are new types from Adélie penguins sampled on Yalour Island in the West Antarctic Peninsula. Co-occurrence of two polyomavirus variants was identified in two individual gentoo penguins. Both of these variants appear to be circulating in gentoo penguins at Cierva Cove, Hope Bay in Trinity Peninsula along the Antarctic Peninsula, and at Hannah Point on Livingstone Island and Stranger Point on King George Island in the South Shetland Islands. Here, we expand the known diversity, host and geographical ranges of penguin polyomaviruses and, together with a previously identified polyomavirus on Ross Island from 2012 to 2013, show that they form five distinct lineages. The four papillomaviruses identified in this study, together with two previously identified from Ross Island in 2012 and 2013 breeding seasons, show substantial diversity reflecting four papillomavirus types across three viral species and two distinct genera. Continued surveillance and viral genomic analysis across a larger geographical framework will help understand the evolution, transmission and incidence rates of these viruses.

Tomato yellow leaf curl disease is a major viral disease severely affecting tomato crops in the Mediterranean region, leading to reduced crop yield and significant economic losses. The disease is caused by monopartite begomoviruses belonging to the Geminiviridae family, primarily tomato yellow leaf curl Sardinia virus (TYLCSV) and tomato yellow leaf curl virus (TYLCV), which often co-infect tomato plants, promoting the emergence of recombinant viral genomes. To investigate the diversity and evolutionary dynamics of these viruses, symptomatic plants collected from agricultural sites in Sicily between 2020 and 2022, along with archived plant samples from 1994 to 1999, were analysed. For each collection site, leaves from symptomatic plants were pooled to form representative samples. Total nucleic acids were extracted and subjected to rolling circle amplification to enrich circular viral genomes. The amplified products were sequenced using Oxford Nanopore Technologies (ONT) long-read sequencing to obtain full-length viral genomes. Bioinformatic analyses revealed that archived samples mainly contained TYLCSV-related sequences, confirming its historical predominance in Sicilian agroecosystems. Recent samples, by contrast, were dominated by TYLCV-derived recombinants such as TYLCV-IS141- and TYLCV-IS76-like variants, indicating a temporal shift in the structure of the viral population. Furthermore, a distinct group of newly emerged recombinants, provisionally referred to as TYLCV-IMS54, was identified in the most recent samples. Their genome comprises a TYLCV backbone, a 54-nt segment from TYLCSV located downstream of the stem-loop region and a 341-nt region derived from TYLCV-Mild. These results demonstrate the importance of continuous viral population monitoring through ONT-based sequencing to detect emerging variants that may influence disease management strategies in tomato crops and highlight the central role of recombination in shaping begomovirus populations.

Eimeria spp. are common coccidian parasites of a wide range of vertebrates, causing diarrhoea, poor weight gain and significant mortality in domestic animals and birds. However, there is a paucity of genomic data on these important pathogens. Of the 11 common Eimeria species in rabbits, only Eimeria stiedae invades biliary epithelial cells rather than the intestine, and the determinants of coccidian tissue tropism remain unclear. In this study, we sequenced the genomes of five common rabbit Eimeria species, including E. stiedae, Eimeria flavescens, Eimeria intestinalis, Eimeria magna and Eimeria media. Comparative genomic analysis reveals that the genome of E. stiedae appears more compact than the genomes of intestinal Eimeria species. It shows reductions in the numbers of rhoptry proteins, dense granule proteins, microneme adhesive repeats and TA4 surface antigens, suggesting that surface and invasion-associated proteins may be involved in the tissue tropism of Eimeria spp. In addition, E. stiedae-specific motifs are identified in a cluster of hypothetical surface antigens. These data provide not only new insights into the biological characteristics of coccidia but also valuable resources for functional research and drug and vaccine development.

Protists are polyphyletic single-celled eukaryotes that underpin global ecosystem functioning, particularly in the oceans. Most remain uncultured, limiting the investigation of their physiology and cell biology. MArine STramenopiles (MASTs) are heterotrophic protists that, although related to well-characterized photosynthetic diatoms and parasitic oomycetes, are poorly studied. The Nanomonadea (MAST-3) species Incisomonas marina has been maintained in co-culture with a bacterial consortium, offering opportunities to investigate the metabolic attributes and nutritional dependencies of the community. Employing a metagenomics approach, the 68 Mbp haploid genome of I. marina was retrieved to an estimated completeness of 93%, representing the most complete MAST genome so far. We also characterized the diversity of, and assembled genomes for, 23 co-cultured bacteria. Auxotrophy of I. marina for B vitamins (B₁, B₂, B₆, B₇ and B₁₂), but not vitamins C, B₃, B₅ and B₉, was predicted. Several bacteria also lacked complete B-vitamin biosynthesis pathways, suggesting that vitamins and/or their precursors are exchanged in the consortium. Moreover, I. marina lacked the ability to synthesize half the protein amino acids, although genes encoding the complete urea cycle were identified, like diatoms; this may play a role in recycling organic nitrogen compounds. Unexpectedly, we also identified the gene DSYB for dimethylsulphoniopropionate biosynthesis. Biosynthesis of this important stress protectant and bacterial chemoattractant is typically found in photosynthetic eukaryotes and has not been identified before in heterotrophic stramenopiles. Together, our study reveals the metabolic attributes of a hitherto understudied organism, advancing knowledge of the evolution and adaptations of the stramenopiles and informing future culturing efforts.