Project description:The complete 1,751,377-bp sequence of the genome of the thermophilic archaeon Methanobacterium thermoautotrophicum deltaH has been determined by a whole-genome shotgun sequencing approach. A total of 1,855 open reading frames (ORFs) have been identified that appear to encode polypeptides, 844 (46%) of which have been assigned putative functions based on their similarities to database sequences with assigned functions. A total of 514 (28%) of the ORF-encoded polypeptides are related to sequences with unknown functions, and 496 (27%) have little or no homology to sequences in public databases. Comparisons with Eucarya-, Bacteria-, and Archaea-specific databases reveal that 1,013 of the putative gene products (54%) are most similar to polypeptide sequences described previously for other organisms in the domain Archaea. Comparisons with the Methanococcus jannaschii genome data underline the extensive divergence that has occurred between these two methanogens; only 352 (19%) of M. thermoautotrophicum ORFs encode sequences that are >50% identical to M. jannaschii polypeptides, and there is little conservation in the relative locations of orthologous genes. When the M. thermoautotrophicum ORFs are compared to sequences from only the eucaryal and bacterial domains, 786 (42%) are more similar to bacterial sequences and 241 (13%) are more similar to eucaryal sequences. The bacterial domain-like gene products include the majority of those predicted to be involved in cofactor and small molecule biosyntheses, intermediary metabolism, transport, nitrogen fixation, regulatory functions, and interactions with the environment. Most proteins predicted to be involved in DNA metabolism, transcription, and translation are more similar to eucaryal sequences. Gene structure and organization have features that are typical of the Bacteria, including genes that encode polypeptides closely related to eucaryal proteins. There are 24 polypeptides that could form two-component sensor kinase-response regulator systems and homologs of the bacterial Hsp70-response proteins DnaK and DnaJ, which are notably absent in M. jannaschii. DNA replication initiation and chromosome packaging in M. thermoautotrophicum are predicted to have eucaryal features, based on the presence of two Cdc6 homologs and three histones; however, the presence of an ftsZ gene indicates a bacterial type of cell division initiation. The DNA polymerases include an X-family repair type and an unusual archaeal B type formed by two separate polypeptides. The DNA-dependent RNA polymerase (RNAP) subunits A', A", B', B" and H are encoded in a typical archaeal RNAP operon, although a second A' subunit-encoding gene is present at a remote location. There are two rRNA operons, and 39 tRNA genes are dispersed around the genome, although most of these occur in clusters. Three of the tRNA genes have introns, including the tRNAPro (GGG) gene, which contains a second intron at an unprecedented location. There is no selenocysteinyl-tRNA gene nor evidence for classically organized IS elements, prophages, or plasmids. The genome contains one intein and two extended repeats (3.6 and 8.6 kb) that are members of a family with 18 representatives in the M. jannaschii genome.
Project description:Chlamydia trachomatis is a human bacterial pathogen that multiplies only within an intracellular membrane-bound vacuole, the inclusion. C. trachomatis includes ocular and urogenital strains, usually causing infections restricted to epithelial cells of the conjunctiva and genital mucosa, respectively, and lymphogranuloma venereum (LGV) strains, which can infect macrophages and spread into lymph nodes. However, C. trachomatis genomes display >98% identity at the DNA level. In this work, we studied whether C. trachomatis Inc proteins, which have a bilobed hydrophobic domain that may mediate their insertion in the inclusion membrane, could be a factor determining these different types of infection and tropisms. Analyses of polymorphisms and phylogeny of 48 Inc proteins from 51 strains encompassing the three disease groups showed significant amino acid differences that were mainly due to variations between Inc proteins from LGV and ocular or urogenital isolates. Studies of the evolutionary dynamics of inc genes suggested that 10 of them are likely under positive selection and indicated that most nonsilent mutations are LGV specific. Additionally, real-time quantitative PCR analyses in prototype and clinical strains covering the three disease groups identified three inc genes with LGV-specific expression. We determined the transcriptional start sites of these genes and found LGV-specific nucleotides within their promoters. Thus, subtle variations in the amino acids of a subset of Inc proteins and in the expression of inc genes may contribute to the unique tropism and invasiveness of C. trachomatis LGV strains.
Project description:BACKGROUND:Nanopore sequencing provides a rapid, cheap and portable real-time sequencing platform with the potential to revolutionize genomics. However, several applications are limited by relatively high single-read error rates (>10 %), including RNA-seq, haplotype sequencing and 16S sequencing. RESULTS:We developed the Intramolecular-ligated Nanopore Consensus Sequencing (INC-Seq) as a strategy for obtaining long and accurate nanopore reads, starting with low input DNA. Applying INC-Seq for 16S rRNA-based bacterial profiling generated full-length amplicon sequences with a median accuracy >97 %. CONCLUSIONS:INC-Seq reads enabled accurate species-level classification, identification of species at 0.1 % abundance and robust quantification of relative abundances, providing a cheap and effective approach for pathogen detection and microbiome profiling on the MinION system.
Project description:The complete nucleotide sequences of six IMP-4-encoding plasmids recovered from Enterobacteriaceae isolates of wildlife origin were characterized. Sequencing data showed that plasmids of different incompatibility groups (IncM, IncI1, IncF, and nontypeable [including an IncX5_2 and two pPrY2001-like]) carried the blaIMP-4-carrying integrons In809 or In1460. Most of the plasmids carried an mph(A) region, and chrA-like, aac(3)-IId, and blaTEM-1b genes. Finally, plasmid analysis revealed the involvement of two different IS26- and Tn1696-associated mechanisms in the mobilization of IMP-4-encoding integrons.