Here we report the draft genome sequence (6.6 Mbp) of the type strain Clostridium magnum, an acetogen with two operons coding for two separate Rnf complexes. C. magnum grows on a broad range of organic substrates and converts CO2 and H2 to acetate using the Wood-Ljungdahl pathway. Copyright © 2016 Uhlig et al.
The methylation-dependent restriction endonuclease (REase) BisI (G(m5)C???NGC) is found in Bacillus subtilis T30. We expressed and purified the BisI endonuclease and 34 BisI homologs identified in bacterial genomes. 23 of these BisI homologs are active based on digestion of (m5)C-modified substrates. Two major specificities were found among these BisI family enzymes: Group I enzymes cut GCNGC containing two to four (m5)C in the two strands, or hemi-methylated sites containing two (m5)C in one strand; Group II enzymes only cut GCNGC sites containing three to four (m5)C, while one enzyme requires all four cytosines to be modified for cleavage. Another homolog, Esp638I cleaves GCS???SGC (relaxed specificity RCN???NGY, containing at least four (m5)C). Two BisI homologs show degenerate specificity cleaving unmodified DNA. Many homologs are small proteins ranging from 150 to 190 amino acid (aa) residues, but some homologs associated with mobile genetic elements are larger and contain an extra C-terminal domain. More than 156 BisI homologs are found in >60 bacterial genera, indicating that these enzymes are widespread in bacteria. They may play an important biological function in restricting pre-modified phage DNA.
Bacteria rarely inhabit infection sites alone, instead residing in diverse, multispecies communities. Despite this fact, bacterial pathogenesis studies primarily focus on monoculture infections, overlooking how community interactions influence the course of disease. In this study, we used global mutant fitness profiling (transposon sequencing [Tn-seq]) to determine the genetic requirements for the pathogenic bacterium Aggregatibacter actinomycetemcomitans to cause disease when coinfecting with the commensal bacterium Streptococcus gordonii Our results show that S. gordonii extensively alters A. actinomycetemcomitans requirements for virulence factors and biosynthetic pathways during infection. In addition, we discovered that the presence of S. gordonii enhances the bioavailability of oxygen during infection, allowing A. actinomycetemcomitans to shift from a primarily fermentative to a respiratory metabolism that enhances its growth yields and persistence. Mechanistically, respiratory metabolism enhances the fitness of A. actinomycetemcomitans in vivo by increasing ATP yields via central metabolism and creating a proton motive force. Our results reveal that, similar to cross-feeding, where one species provides another species with a nutrient, commensal bacteria can also provide electron acceptors that promote the respiratory growth and fitness of pathogens in vivo, an interaction that we term cross-respiration.Commensal bacteria can enhance the virulence of pathogens in mixed-species infections. However, knowledge of the mechanisms underlying this clinically relevant phenomenon is lacking. To bridge this gap, we comprehensively determined the genes a pathogen needs to establish coinfection with a commensal. Our findings show that the metabolism of the pathogen is low-energy-yielding in monoinfection, but in coinfection, the commensal improves the fitness of the pathogen by increasing the bioavailability of oxygen, thereby shifting the pathogen toward a high-energy-yielding metabolism. Similar to cross-feeding, this interaction, which we term cross-respiration, illustrates that commensal bacteria can provide electron acceptors that enhance the virulence of pathogens during infection. Copyright © 2016 Stacy et al.
Biological features can be inferred, based on genomic data, for many microbial lineages that remain uncultured. However, cultivation is important for characterizing an organism’s physiology and testing its genome-encoded potential. Here we use single-cell genomics to infer cultivation conditions for the isolation of an ectosymbiotic Nanoarchaeota (‘Nanopusillus acidilobi’) and its host (Acidilobus, a crenarchaeote) from a terrestrial geothermal environment. The cells of ‘Nanopusillus’ are among the smallest known cellular organisms (100-300?nm). They appear to have a complete genetic information processing machinery, but lack almost all primary biosynthetic functions as well as respiration and ATP synthesis. Genomic and proteomic comparison with its distant relative, the marine Nanoarchaeum equitans illustrate an ancient, common evolutionary history of adaptation of the Nanoarchaeota to ectosymbiosis, so far unique among the Archaea.
Enterohemorrhagic E. coli (EHEC) O157:H7 strain EDL933 harbors multiple prophage-associated open reading frames (orfs) in its genome, which are highly homologous to the chromosomal nanS gene. The latter is part of the nanCMS-operon, which is present in most E. coli strains and encodes an esterase, which is responsible for the mono-deacetylation of 5-N-acetyl-9-O-acetyl neuraminic acid (Neu5,9Ac2). Whereas one prophage-borne orf (z1466) has been characterized in previous studies, the functions of the other nanS-homologous orfs are unknown.In the current study, the nanS-homologous orfs of EDL933 were initially studied in silico Due to their homology to the chromosomal nanS gene and their location in prophage genomes, we designated them nanS-p, and numbered the different nanS-p alleles consecutively from 1-10. The two alleles nanS-p2 and nanS-p4 were selected for production of recombinant proteins, their enzymatic activities were investigated and differences in their temperature optima were found. Furthermore, a function of these enzymes in substrate utilization could be demonstrated using an E. coli C600?nanS mutant in a growth medium with Neu5,9Ac2 as carbon source and supplementation with the different recombinant NanS-p proteins. Moreover, generation of sequential deletion of all nanS-p alleles in strain EDL933, and subsequent growth experiments demonstrated a gene-dose-effect on the utilization of Neu5,9Ac2Since Neu5,9Ac2 is an important component of human and animal gut mucus, and the nutrient availability in the large intestine is limited, we hypothesize that the presence of multiple Neu5,9Ac2-esterases provides them a nutrient supply under certain conditions in the large intestine, even if particular prophages get lost.In this study, a group of homologous prophage-borne nanS-p alleles and two of the corresponding enzymes of enterohemorrhagic E. coli (EHEC) O157:H7 strain EDL933 are characterized that may be important to provide alternative genes for substrate utilization. Copyright © 2016, American Society for Microbiology. All Rights Reserved.
The transfer of DNA between Enterococcus faecium strains has been characterized by both the movement of well-defined genetic elements and by the large-scale transfer of genomic DNA fragments. In this work we report on the whole genome analysis of transconjugants resulting from mating events between the vancomycin-resistant E. faecium C68 strain and vancomycin susceptible D344RRF to discern the mechanism by which the transferred regions enter the recipient chromosome. Vancomycin-resistant transconjugants from five independent matings were analysed by whole genome sequencing. In all cases but one, the penicillin binding protein 5 gene (pbp5) and the Tn5382-vancomycin resistance transposon were transferred together and replaced the corresponding pbp5 region of D344RRF. In one instance, Tn5382 inserted independently downstream of the D344RRF pbp5 Single nucleotide variants (SNV) analysis suggests that entry of donor DNA into the recipient chromosome occurred by recombination across regions of homology between donor and recipient chromosomes, rather than through insertion sequence-mediated transposition. Transfer of genomic DNA was also associated with transfer of C68 plasmid pLRM23 and another putative plasmid. Our data are consistent with transfer initiated by a cointegration of a transferable plasmid with the donor chromosome, with subsequent circularization of the plasmid/chromosome cointegrate in the donor prior to transfer. Entry into the recipient chromosome occurs most commonly across regions of homology between donor and recipient chromosomes. Copyright © 2016 García-Solache et al.
Myxobacteria are members of d-proteobacteria and are typified by large genomes, well-coordinated social behavior, gliding motility, and starvation-induced fruiting body formation. Here, we report the 10.33 Mb whole genome of a starch-degrading myxobacterium Sandaracinus amylolyticus DSM 53668(T) that encodes 8,962 proteins, 56 tRNA, and two rRNA operons. Phylogenetic analysis, in silico DNA-DNA hybridization and average nucleotide identity reveal its divergence from other myxobacterial species and support its taxonomic characterization into a separate family Sandaracinaceae, within the suborder Sorangiineae. Sequence similarity searches using the Carbohydrate-active enzymes (CAZyme) database help identify the enzyme repertoire of S. amylolyticus involved in starch, agar, chitin, and cellulose degradation. We identified 16 a-amylases and two ?-amylases in the S. amylolyticus genome that likely play a role in starch degradation. While many of the amylases are seen conserved in other d-proteobacteria, we notice several novel amylases acquired via horizontal transfer from members belonging to phylum Deinococcus-Thermus, Acidobacteria, and Cyanobacteria. No agar degrading enzyme(s) were identified in the S. amylolyticus genome. Interestingly, several putative ß-glucosidases and endoglucanases proteins involved in cellulose degradation were identified. However, the absence of cellobiohydrolases/exoglucanases corroborates with the lack of cellulose degradation by this bacteria. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.
Peptoclostridium difficile (Clostridium difficile) is the major pathogen associated with infectious diarrhea in humans. Concomitant with the increased incidence of C. difficile infection worldwide, there is an increasing concern regarding this infection type. This study reports a draft assembly and detailed sequence analysis of C. difficile strain ZJCDC-S82. The de novo assembled genome was 4.19 Mb in size, which includes 4,013 protein-coding genes, 41 rRNA genes, and 84 tRNA genes. Along with the nuclear genome, we also assembled sequencing information for a single plasmid consisting of 11,930 nucleotides. Comparative genomic analysis of C. difficile ZJCDC-S82 and two other previously published strains, such as M120 and CD630, showed extensive similarity. Phylogenetic analysis revealed that genetic diversity among C. difficile strains was not influenced by geographic location. Evolutionary analysis suggested that four genes encoding surface proteins exhibited positive selection in C. difficile ZJCDC-S82. Codon usage analysis indicated that C. difficile ZJCDC-S82 had high codon usage bias toward A/U-ended codons. Furthermore, codon usage patterns in C. difficile ZJCDC-S82 were predominantly affected by mutation pressure. Our results provide detailed information pertaining to the C. difficile genome associated with a strain from mainland China. This analysis will facilitate the understanding of genomic diversity and evolution of C. difficile strains in this region.
During 2013, ST278 Klebsiella pneumoniae with blaNDM-7 was isolated from the urine (KpN01) and rectum (KpN02) of a patient in Calgary, Canada. The same strain (KpN04) was subsequently isolated from another patient in the same unit. Interestingly, a carbapenem-susceptible K. pneumoniae ST278 (KpN06) was obtained 1 month later from the blood of the second patient. Next-generation sequencing (NGS) revealed that the loss of carbapenem-resistance in KpN06 was due to a 5-kb deletion on the blaNDM-7-harboring IncX3 plasmid. In addition, an IncFIB plasmid in KpN06 had a 27-kb deletion that removed genes encoding for heavy metal resistance. Phylogenetic analysis showed that the K. pneumoniae ST278 from patient 2 was likely a descendant of KpN02 and that KpN06 was a close progenitor of an environmental ST278. It is unclear whether KpN06 lost the blaNDM-7 gene in vivo. This study detailed the remarkable plasticity and speed of evolutionary changes in multidrug-resistant K. pneumoniae, demonstrating the highly recombinant nature of this species. It also highlights the ability of NGS to clarify molecular microevolutionary events within antibiotic-resistant organisms.© The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.
Bifidobacteria, often associated with the gastrointestinal tract of animals, are well known for their roles as probiotics. Among the dozens of Bifidobacterium species, Bifidobacterium bifidum, B. breve, and B. longum are the ones most frequently isolated from the feces of infants and known to help the digestion of human milk oligosaccharides. To investigate the correlation between the metabolic properties of bifidobacteria and their phylogeny, we performed a phylogenomic analysis based on 452 core genes of forty-four completely sequenced Bifidobacterium species. Results show that a major evolutionary event leading to the clade of the infant-adapted species is linked to carbohydrate metabolism, but it is not the only factor responsible for the adaptation of bifidobacteria to the gut. The genome of B. longum subsp. infantis, a typical bifidobacterium in the gut of breast-fed infants, encodes proteins associated with several kinds of species-specific metabolic pathways, including urea metabolism and biosynthesis of riboflavin and lantibiotics. Our results demonstrate that these metabolic features, which are associated with the probiotic function of bifidobacteria, are species-specific and highly correlate with their phylogeny. Copyright © 2016 Elsevier GmbH. All rights reserved.
Clostridium pasteurianum is emerging as a prospective host for the production of biofuels and chemicals, and has recently been shown to directly consume electric current. Despite this growing biotechnological appeal, the organism’s genetics and central metabolism remain poorly understood. Here we present a concurrent genome sequence for the C. pasteurianum type strain and provide extensive genomic analysis of the organism’s defence mechanisms and central fermentative metabolism. Next generation genome sequencing produced reads corresponding to spontaneous excision of a novel phage, designated f6013, which could be induced using mitomycin C and detected using PCR and transmission electron microscopy. Methylome analysis of sequencing reads provided a near-complete glimpse into the organism’s restriction-modification systems. We also unveiled the chief C. pasteurianum Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) locus, which was found to exemplify a Type I-B system. Finally, we show that C. pasteurianum possesses a highly complex fermentative metabolism whereby the metabolic pathways enlisted by the cell is governed by the degree of reductance of the substrate. Four distinct fermentation profiles, ranging from exclusively acidogenic to predominantly alcohologenic, were observed through redox consideration of the substrate. A detailed discussion of the organism’s central metabolism within the context of metabolic engineering is provided.
We and others have shown the utility of long sequence reads to improve genome assembly quality. In this study, we generated PacBio DNA sequence data to improve the assemblies of draft genomes for Clostridium thermocellum AD2, Clostridium thermocellum LQRI, and Pelosinus fermentans R7. Copyright © 2016 Utturkar et al.
Clostridium sp. Bc-iso-3 is a cellulolytic strain isolated from a Swedish industrial-scale biogas digester. Here, we present the draft genome sequence of this strain, which consists of four contigs with a total length of 4,327,139 bp and an average coverage of 312.97×. Copyright © 2016 Sun and Schnürer.
Intestinimonas is a newly described bacterial genus with representative strains present in the intestinal tract of human and other animals. Despite unique metabolic features including the production of butyrate from both sugars and amino acids, there is to date no data on their diversity, ecology, and physiology. Using a comprehensive phylogenetic approach, Intestinimomas was found to include at least three species that colonize primarily the human and mouse intestine. We focused on the most common and cultivable species of the genus, Intestinimonas butyriciproducens, and performed detailed genomic and physiological comparison of strains SRB521(T) and AF211, isolated from the mouse and human gut respectively. The complete 3.3-Mb genomic sequences of both strains were highly similar with 98.8% average nucleotide identity, testifying to their assignment to one single species. However, thorough analysis revealed significant genomic rearrangements, variations in phage-derived sequences, and the presence of new CRISPR sequences in both strains. Moreover, strain AF211 appeared to be more efficient than strain SRB521(T) in the conversion of the sugars arabinose and galactose. In conclusion, this study provides genomic and physiological insight into Intestinimonas butyriciproducens, a prevalent butyrate-producing species, differentiating strains that originate from the mouse and human gut.© 2016 The Authors. Environmental Microbiology Reports published by Society for Applied Microbiology and JohnWiley & Sons Ltd.
Owing to gene transposition and plasmid conjugation, New Delhi metallo-ß-lactamase (NDM) is typically identified among varied Enterobacteriaceae species and STs. We used WGS to characterize the chromosomal and plasmid molecular epidemiology of NDM transmission involving four institutions in Singapore.Thirty-three Enterobacteriaceae isolates (collection years 2010-14) were sequenced using short-read sequencing-by-synthesis and analysed. Long-read single molecule, real-time sequencing (SMRTS) was used to characterize genetically a novel plasmid pSg1-NDM carried on Klebsiella pneumoniae ST147.In 20 (61%) isolates, blaNDM was located on the pNDM-ECS01 plasmid in the background of multiple bacterial STs, including eight K. pneumoniae STs and five Escherichia coli STs. In six (18%) isolates, a novel blaNDM-positive plasmid, pSg1-NDM, was found only in K. pneumoniae ST147. The pSg1-NDM-K. pneumoniae ST147 clone (Sg1-NDM) was fully sequenced using SMRTS. pSg1-NDM, a 90?103 bp IncR plasmid, carried genes responsible for resistance to six classes of antimicrobials. A large portion of pSg1-NDM had no significant homology to any known plasmids in GenBank. pSg1-NDM had no conjugative transfer region. Combined chromosomal-plasmid phylogenetic analysis revealed five clusters of clonal bacterial NDM-positive plasmid transmission, of which two were inter-institution clusters. The largest inter-institution cluster involved six K. pneumoniae ST147-pSg1-NDM isolates. Fifteen patients were involved in transmission clusters, of which four had ward contact, six had hospital contact and five had an unknown transmission link.A combined sequencing-by-synthesis and SMRTS approach can determine effectively the transmission clusters of blaNDM and genetically characterize novel plasmids. Plasmid molecular epidemiology is important to understanding NDM spread as blaNDM-positive plasmids can conjugate extensively across species and STs.© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.
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