Mycoplasma bovis is a major bacterial pathogen that can cause respiratory disease, mastitis, and arthritis in cattle. We report here the complete and annotated genome sequence of M. bovis strain 08M, isolated from a calf lung with pneumonia in China. Copyright © 2017 Chen et al.
We report here the complete genome of Vibrio coralliilyticus strain 58, which was originally isolated from inactive Pacific oyster (Crassostrea gigas) larvae in Japan. The assembled genome consisted of two chromosomes and one plasmid. These data will provide valuable information and important insights into the biodiversity of this organism. Copyright © 2017 Kim et al.
Bacillus subtilis J-5 was isolated from tomato rhizosphere soil and exhibited strong inhibitory activity against Botrytis cinerea To shed light on the molecular mechanism underlying the biological control on phytopathogens, the whole genome of this strain was sequenced. Genes encoding antimicrobial compounds and the regulatory systems were identified in the genome. Copyright © 2017 Jia et al.
Evolutionary shifts in bacterial virulence are often associated with a third biological player, for instance temperate phages, that can act as hyperparasites. By integrating as prophages into the bacterial genome they can contribute accessory genes, which can enhance the fitness of their prokaryotic carrier (lysogenic conversion). Hyperparasitic influence in tripartite biotic interactions has so far been largely neglected in empirical host-parasite studies due to their inherent complexity. Here we experimentally address whether bacterial resistance to phages and bacterial harm to eukaryotic hosts is linked using a natural tri-partite system with bacteria of the genus Vibrio, temperate vibriophages and the pipefish Syngnathus typhle. We induced prophages from all bacterial isolates and constructed a three-fold replicated, fully reciprocal 75 × 75 phage-bacteria infection matrix.According to their resistance to phages, bacteria could be grouped into three distinct categories: highly susceptible (HS-bacteria), intermediate susceptible (IS-bacteria), and resistant (R-bacteria). We experimentally challenged pipefish with three selected bacterial isolates from each of the three categories and determined the amount of viable Vibrio counts from infected pipefish and the expression of pipefish immune genes. While the amount of viable Vibrio counts did not differ between bacterial groups, we observed a significant difference in relative gene expression between pipefish infected with phage susceptible and phage resistant bacteria.These findings suggest that bacteria with a phage-susceptible phenotype are more harmful against a eukaryotic host, and support the importance of hyperparasitism and the need for an integrative view across more than two levels when studying host-parasite evolution.
Caldicellulosiruptor bescii is the most thermophilic cellulose degrader known and is of great interest because of its ability to degrade nonpretreated plant biomass. For biotechnological applications, an efficient genetic system is required to engineer it to convert plant biomass into desired products. To date, two different genetically tractable lineages of C. bescii strains have been generated. The first (JWCB005) is based on a random deletion within the pyrimidine biosynthesis genes pyrFA, and the second (MACB1018) is based on the targeted deletion of pyrE, making use of a kanamycin resistance marker. Importantly, an active insertion element, ISCbe4, was discovered in C. bescii when it disrupted the gene for lactate dehydrogenase (ldh) in strain JWCB018, constructed in the JWCB005 background. Additional instances of ISCbe4 movement in other strains of this lineage are presented herein. These observations raise concerns about the genetic stability of such strains and their use as metabolic engineering platforms. In order to investigate genome stability in engineered strains of C. bescii from the two lineages, genome sequencing and Southern blot analyses were performed. The evidence presented shows a dramatic increase in the number of single nucleotide polymorphisms, insertions/deletions, and ISCbe4 elements within the genome of JWCB005, leading to massive genome rearrangements in its daughter strain, JWCB018. Such dramatic effects were not evident in the newer MACB1018 lineage, indicating that JWCB005 and its daughter strains are not suitable for metabolic engineering purposes in C. bescii Furthermore, a facile approach for assessing genomic stability in C. bescii has been established. IMPORTANCE Caldicellulosiruptor bescii is a cellulolytic extremely thermophilic bacterium of great interest for metabolic engineering efforts geared toward lignocellulosic biofuel and bio-based chemical production. Genetic technology in C. bescii has led to the development of two uracil auxotrophic genetic background strains for metabolic engineering. We show that strains derived from the genetic background containing a random deletion in uracil biosynthesis genes (pyrFA) have a dramatic increase in the number of single nucleotide polymorphisms, insertions/deletions, and ISCbe4 insertion elements in their genomes compared to the wild type. At least one daughter strain of this lineage also contains large-scale genome rearrangements that are flanked by these ISCbe4 elements. In contrast, strains developed from the second background strain developed using a targeted deletion strategy of the uracil biosynthetic gene pyrE have a stable genome structure, making them preferable for future metabolic engineering studies. Copyright © 2017 American Society for Microbiology.
Acetylene (C2H2) is a trace constituent of the present Earth’s oxidizing atmosphere, reflecting a mix of terrestrial and marine emissions from anthropogenic, biomass burning, and unidentified biogenic sources. Fermentation of acetylene was serendipitously discovered during C2H2-block assays of N2O reductase, and Pelobacter acetylenicus was shown to grow on C2H2 via acetylene hydratase (AH). AH is a W-containing, catabolic, low redox potential enzyme that unlike nitrogenase (N2ase) is specific for acetylene. Acetylene fermentation is a rare metabolism that is well-characterized only in P. acetylenicus DSM3246 and DSM3247, and Pelobacter sp. strain SFB93. To better understand the genetic controls on AH activity, we sequenced the genomes of the three acetylene-fermenting Pelobacter strains. Genome assembly and annotation produced three novel genomes containing gene sequences for AH, with two copies being present in SFB93. In addition, gene sequences for all five compulsory genes for Mo-Fe nitrogenase were also present in the three genomes, indicating the co-occurrence of 2 acetylene-transformation pathways. Nitrogen fixation growth assays showed that DSM3426 could ferment acetylene in the absence of ammonium, but no ethylene was produced. However, SFB93 degraded acetylene, and in the absence of ammonium, produced ethylene indicating an active N2ase. Diazotrophic growth was observed under N2 but not in experimental controls incubated under Ar. SFB93 exhibits acetylene fermentation and nitrogen fixation, the only known biochemical mechanisms for acetylene transformation. Our results indicate complex interactions between N2ase and AH and suggest novel evolutionary pathways of these relic enzymes from early Earth to modern day.Importance Here we show that a single Pelobacter strain can grow via acetylene fermentation and carry out nitrogen fixation, using the only 2 enzymes known to transform acetylene. These findings provide new insights into acetylene transformations and adaptations for nutrient (C, N) and energy acquisition by microorganisms. Enhanced understanding of acetylene transformations in modern environments (i.e., extent, occurrence, rates, etc.) is important for using acetylene as a potential biomarker for extraterrestrial life and degradation of anthropogenic contaminants. Copyright © 2017 American Society for Microbiology.
Folsomia candida is a model in soil biology, belonging to the family of Isotomidae, subclass Collembola. It reproduces parthenogenetically in the presence of Wolbachia, and exhibits remarkable physiological adaptations to stress. To better understand these features and adaptations to life in the soil, we studied its genome in the context of its parthenogenetic lifestyle.We applied Pacific Bioscience sequencing and assembly to generate a reference genome for F. candida of 221.7 Mbp, comprising only 162 scaffolds. The complete genome of its endosymbiont Wolbachia, was also assembled and turned out to be the largest strain identified so far. Substantial gene family expansions and lineage-specific gene clusters were linked to stress response. A large number of genes (809) were acquired by horizontal gene transfer. A substantial fraction of these genes are involved in lignocellulose degradation. Also, the presence of genes involved in antibiotic biosynthesis was confirmed. Intra-genomic rearrangements of collinear gene clusters were observed, of which 11 were organized as palindromes. The Hox gene cluster of F. candida showed major rearrangements compared to arthropod consensus cluster, resulting in a disorganized cluster.The expansion of stress response gene families suggests that stress defense was important to facilitate colonization of soils. The large number of HGT genes related to lignocellulose degradation could be beneficial to unlock carbohydrate sources in soil, especially those contained in decaying plant and fungal organic matter. Intra- as well as inter-scaffold duplications of gene clusters may be a consequence of its parthenogenetic lifestyle. This high quality genome will be instrumental for evolutionary biologists investigating deep phylogenetic lineages among arthropods and will provide the basis for a more mechanistic understanding in soil ecology and ecotoxicology.
Bacteria that produce the broad-spectrum Carbapenem antibiotic New Delhi Metallo-ß-lactamase (NDM) place a burden on health care systems worldwide, due to the limited treatment options for infections caused by them and the rapid global spread of this antibiotic resistance mechanism. Although it is believed that the associated resistance gene blaNDM-1 originated in Acinetobacter spp., the role of Enterobacteriaceae in its dissemination remains unclear. In this study, we used whole genome sequencing to investigate the dissemination dynamics of blaNDM-1-positive plasmids in a set of 21 clinical NDM-1-positive isolates from Colombia and Mexico (Providencia rettgeri, Klebsiella pneumoniae, and Acinetobacter baumannii) as well as six representative NDM-1-positive Escherichia coli transconjugants. Additionally, the plasmids from three representative P. rettgeri isolates were sequenced by PacBio sequencing and finished. Our results demonstrate the presence of previously reported plasmids from K. pneumoniae and A. baumannii in different genetic backgrounds and geographically distant locations in Colombia. Three new previously unclassified plasmids were also identified in P. rettgeri from Colombia and Mexico, plus an interesting genetic link between NDM-1-positive P. rettgeri from distant geographic locations (Canada, Mexico, Colombia, and Israel) without any reported epidemiological links was discovered. Finally, we detected a relationship between plasmids present in P. rettgeri and plasmids from A. baumannii and K. pneumoniae. Overall, our findings suggest a Russian doll model for the dissemination of blaNDM-1 in Latin America, with P. rettgeri playing a central role in this process, and reveal new insights into the evolution and dissemination of plasmids carrying such antibiotic resistance genes.© The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.
Tiny photosynthetic microorganisms that form the picoplankton (between 0.3 and 3 µm in diameter) are at the base of the food web in many marine ecosystems, and their adaptability to environmental change hinges on standing genetic variation. Although the genomic and phenotypic diversity of the bacterial component of the oceans has been intensively studied, little is known about the genomic and phenotypic diversity within each of the diverse eukaryotic species present. We report the level of genomic diversity in a natural population of Ostreococcus tauri (Chlorophyta, Mamiellophyceae), the smallest photosynthetic eukaryote. Contrary to the expectations of clonal evolution or cryptic species, the spectrum of genomic polymorphism observed suggests a large panmictic population (an effective population size of 1.2 × 10(7)) with pervasive evidence of sexual reproduction. De novo assemblies of low-coverage chromosomes reveal two large candidate mating-type loci with suppressed recombination, whose origin may pre-date the speciation events in the class Mamiellophyceae. This high genetic diversity is associated with large phenotypic differences between strains. Strikingly, resistance of isolates to large double-stranded DNA viruses, which abound in their natural environment, is positively correlated with the size of a single hypervariable chromosome, which contains 44 to 156 kb of strain-specific sequences. Our findings highlight the role of viruses in shaping genome diversity in marine picoeukaryotes.
The extended-spectrum-ß-lactamase (ESBL)- and Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae represent serious and urgent threats to public health. In a retrospective study of multidrug-resistant K. pneumoniae, we identified three clinical isolates, CN1, CR14, and NY9, carrying both blaCTX-M and blaKPC genes. The complete genomes of these three K. pneumoniae isolates were de novo assembled by using both short- and long-read whole-genome sequencing. In CR14 and NY9, blaCTX-M and blaKPC were carried on two different plasmids. In contrast, CN1 had one copy of blaKPC-2 and three copies of blaCTX-M-15 integrated in the chromosome, for which the blaCTX-M-15 genes were linked to an insertion sequence, ISEcp1, whereas the blaKPC-2 gene was in the context of a Tn4401a transposition unit conjugated with a PsP3-like prophage. Intriguingly, downstream of the Tn4401a-blaKPC-2-prophage genomic island, CN1 also carried a clustered regularly interspaced short palindromic repeat (CRISPR)-cas array with four spacers targeting a variety of K. pneumoniae plasmids harboring antimicrobial resistance genes. Comparative genomic analysis revealed that there were two subtypes of type I-E CRISPR-cas in K. pneumoniae strains and suggested that the evolving CRISPR-cas, with its acquired novel spacer, induced the mobilization of antimicrobial resistance genes from plasmids into the chromosome. The integration and dissemination of multiple copies of blaCTX-M and blaKPC from plasmids to chromosome depicts the complex pandemic scenario of multidrug-resistant K. pneumoniae Additionally, the implications from this study also raise concerns for the application of a CRISPR-cas strategy against antimicrobial resistance. Copyright © 2017 American Society for Microbiology.
Trichoderma reesei (Ascomycota, Pezizomycotina) QM6a is a model fungus for a broad spectrum of physiological phenomena, including plant cell wall degradation, industrial production of enzymes, light responses, conidiation, sexual development, polyketide biosynthesis, and plant-fungal interactions. The genomes of QM6a and its high enzyme-producing mutants have been sequenced by second-generation-sequencing methods and are publicly available from the Joint Genome Institute. While these genome sequences have offered useful information for genomic and transcriptomic studies, their limitations and especially their short read lengths make them poorly suited for some particular biological problems, including assembly, genome-wide determination of chromosome architecture, and genetic modification or engineering.We integrated Pacific Biosciences and Illumina sequencing platforms for the highest-quality genome assembly yet achieved, revealing seven telomere-to-telomere chromosomes (34,922,528 bp; 10877 genes) with 1630 newly predicted genes and >1.5 Mb of new sequences. Most new sequences are located on AT-rich blocks, including 7 centromeres, 14 subtelomeres, and 2329 interspersed AT-rich blocks. The seven QM6a centromeres separately consist of 24 conserved repeats and 37 putative centromere-encoded genes. These findings open up a new perspective for future centromere and chromosome architecture studies. Next, we demonstrate that sexual crossing readily induced cytosine-to-thymine point mutations on both tandem and unlinked duplicated sequences. We also show by bioinformatic analysis that T. reesei has evolved a robust repeat-induced point mutation (RIP) system to accumulate AT-rich sequences, with longer AT-rich blocks having more RIP mutations. The widespread distribution of AT-rich blocks correlates genome-wide partitions with gene clusters, explaining why clustering of genes has been reported to not influence gene expression in T. reesei.Compartmentation of ancestral gene clusters by AT-rich blocks might promote flexibilities that are evolutionarily advantageous in this fungus’ soil habitats and other natural environments. Our analyses, together with the complete genome sequence, provide a better blueprint for biotechnological and industrial applications.
Chaperonins are protein-folding machinery found in all cellular life. Chaperonin genes have been documented within a few viruses, yet, surprisingly, analysis of metagenome sequence data indicated that chaperonin-carrying viruses are common and geographically widespread in marine ecosystems. Also unexpected was the discovery of viral chaperonin sequences related to thermosome proteins of archaea, indicating the presence of virioplankton populations infecting marine archaeal hosts. Virioplankton large subunit chaperonin sequences (GroELs) were divergent from bacterial sequences, indicating that viruses have carried this gene over long evolutionary time. Analysis of viral metagenome contigs indicated that: the order of large and small subunit genes was linked to the phylogeny of GroEL; both lytic and temperate phages may carry group I chaperonin genes; and viruses carrying a GroEL gene likely have large double-stranded DNA (dsDNA) genomes (>70?kb). Given these connections, it is likely that chaperonins are critical to the biology and ecology of virioplankton populations that carry these genes. Moreover, these discoveries raise the intriguing possibility that viral chaperonins may more broadly alter the structure and function of viral and cellular proteins in infected host cells.
Single molecule sequencing (SMS) platforms enable base sequences to be read directly from individual strands of DNA in real-time. Though capable of long read lengths, SMS platforms currently suffer from low throughput compared to competing short-read sequencing technologies. Here, we present a novel strategy for sequencing library preparation, dubbed ConcatSeq, which increases the throughput of SMS platforms by generating long concatenated templates from pools of short DNA molecules. We demonstrate adaptation of this technique to two target enrichment workflows, commonly used for oncology applications, and feasibility using PacBio single molecule real-time (SMRT) technology. Our approach is capable of increasing the sequencing throughput of the PacBio RSII platform by more than five-fold, while maintaining the ability to correctly call allele frequencies of known single nucleotide variants. ConcatSeq provides a versatile new sample preparation tool for long-read sequencing technologies.
Botryococcus braunii has long been known as a prodigious producer of liquid hydrocarbon oils that can be converted into combustion engine fuels. This draft genome for the B race of B. braunii will allow researchers to unravel important hydrocarbon biosynthetic pathways and identify possible regulatory networks controlling this unusual metabolism. Copyright © 2017 Browne et al.
Ralstonia solanacearum is an important soil-borne plant pathogen with broad geographical distribution and the ability to cause wilt disease in many agriculturally important crops. Genome sequencing of multiple R. solanacearum strains has identified both unique and shared genetic traits influencing their evolution and ability to colonize plant hosts. Previous research has shown that DNA methylation can drive speciation and modulate virulence in bacteria, but the impact of epigenetic modifications on the diversification and pathogenesis of R. solanacearum is unknown. Sequencing of R. solanacearum strains GMI1000 and UY031 using Single Molecule Real-Time technology allowed us to perform a comparative analysis of R. solanacearum methylomes. Our analysis identified a novel methylation motif associated with a DNA methylase that is conserved in all complete Ralstonia spp. genomes and across the Burkholderiaceae, as well as a methylation motif associated to a phage-borne methylase unique to R. solanacearum UY031. Comparative analysis of the conserved methylation motif revealed that it is most prevalent in gene promoter regions, where it displays a high degree of conservation detectable through phylogenetic footprinting. Analysis of hyper- and hypo-methylated loci identified several genes involved in global and virulence regulatory functions whose expression may be modulated by DNA methylation. Analysis of genome-wide modification patterns identified a significant correlation between DNA modification and transposase genes in R. solanacearum UY031, driven by the presence of a high copy number of ISrso3 insertion sequences in this genome and pointing to a novel mechanism for regulation of transposition. These results set a firm foundation for experimental investigations into the role of DNA methylation in R. solanacearum evolution and its adaptation to different plants.
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