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PubMed2003

Complete genome sequence and comparative analysis of the industrial microorganism Streptomyces avermitilis.

Species of the genus Streptomyces are of major pharmaceutical interest because they synthesize a variety of bioactive secondary metabolites. We have determined the complete nucleotide sequence of the linear chromosome of Streptomyces avermitilis. S. avermitilis produces avermectins, a group of antiparasitic agents used in human and veterinary medicine. The genome contains 9,025,608 bases (average GC content, 70.7%) and encodes at least 7,574 potential open reading frames (ORFs). Thirty-five percent of the ORFs (2,664) constitute 721 paralogous families. Thirty gene clusters related to secondary metabolite biosynthesis were identified, corresponding to 6.6% of the genome. Comparison with Streptomyces coelicolor A3(2) revealed that an internal 6.5-Mb region in the S. avermitilis genome was highly conserved with respect to gene order and content, and contained all known essential genes but showed perfectly asymmetric structure at the oriC center. In contrast, the terminal regions were not conserved and preferentially contained nonessential genes.

Amino Acid Sequence↗
PubMed2001

Complete genome sequence of Salmonella enterica serovar Typhimurium LT2.

Salmonella enterica subspecies I, serovar Typhimurium (S. typhimurium), is a leading cause of human gastroenteritis, and is used as a mouse model of human typhoid fever. The incidence of non-typhoid salmonellosis is increasing worldwide, causing millions of infections and many deaths in the human population each year. Here we sequenced the 4,857-kilobase (kb) chromosome and 94-kb virulence plasmid of S. typhimurium strain LT2. The distribution of close homologues of S. typhimurium LT2 genes in eight related enterobacteria was determined using previously completed genomes of three related bacteria, sample sequencing of both S. enterica serovar Paratyphi A (S. paratyphi A) and Klebsiella pneumoniae, and hybridization of three unsequenced genomes to a microarray of S. typhimurium LT2 genes. Lateral transfer of genes is frequent, with 11% of the S. typhimurium LT2 genes missing from S. enterica serovar Typhi (S. typhi), and 29% missing from Escherichia coli K12. The 352 gene homologues of S. typhimurium LT2 confined to subspecies I of S. enterica-containing most mammalian and bird pathogens-are useful for studies of epidemiology, host specificity and pathogenesis. Most of these homologues were previously unknown, and 50 may be exported to the periplasm or outer membrane, rendering them accessible as therapeutic or vaccine targets.

Animals↗
PubMed2001

The complete genome sequence of the lactic acid bacterium Lactococcus lactis ssp. lactis IL1403.

Lactococcus lactis is a nonpathogenic AT-rich gram-positive bacterium closely related to the genus Streptococcus and is the most commonly used cheese starter. It is also the best-characterized lactic acid bacterium. We sequenced the genome of the laboratory strain IL1403, using a novel two-step strategy that comprises diagnostic sequencing of the entire genome and a shotgun polishing step. The genome contains 2,365,589 base pairs and encodes 2310 proteins, including 293 protein-coding genes belonging to six prophages and 43 insertion sequence (IS) elements. Nonrandom distribution of IS elements indicates that the chromosome of the sequenced strain may be a product of recent recombination between two closely related genomes. A complete set of late competence genes is present, indicating the ability of L. lactis to undergo DNA transformation. Genomic sequence revealed new possibilities for fermentation pathways and for aerobic respiration. It also indicated a horizontal transfer of genetic information from Lactococcus to gram-negative enteric bacteria of Salmonella-Escherichia group.

Amino Acids↗
PubMed1999

The identification and characterization of two promoters and the complete genomic sequence for the Wiskott-Aldrich syndrome gene.

The Wiskott-Aldrich syndrome (WAS) is an X-linked disorder characterized by immunodeficiency, eczema and thrombocytopenia. The gene responsible for WAS was identified through positional cloning, and the function of the encoded protein (WASP) is still the subject of much speculation. WASP is currently thought to be involved in the regulation of actin polymerization in hematopoietic cells. To study the elements that regulate the WASP gene, we have identified the sites for transcription initiation. We found that two promoters were responsible for controlling WASP expression. Multiple transcription initiation sites were found immediately adjacent to the translation start site, however an alternate exon with a second promoter region was identified 6 kb upstream. Examination of the 5' sequence adjacent to the initiation sites in both promoters failed to reveal a TATA or CCAAT box, but numerous putative transcription factor binding sites including Sp1, Ets, c-Myb and PU.1 were apparent. Reporter constructs generated from each promoter showed functional activity in the Jurkat T-cell and HEL erythro-megakaryocytic cell lines. Although the alternate exon sequence was extremely GC rich and contained several potential binding elements, the primary promoter was stronger than the upstream promoter in the cell lines assayed. The transcription factor binding site profiles within each promoter suggested that they may play different roles in regulating WASP expression depending on the stage of differentiation and development, and the cell lineage. In this study we have also reported the complete nucleotide sequence of the coding and intervening sequences for the WASP gene. A comprehensive knowledge of the genomic structure and the further characterization of WASP gene expression will facilitate the continued investigation of mutations in WAS patients, and the eventual prospect of gene therapy.

Base Sequence↗
PubMed2002

Complete genome sequence of Montana Myotis leukoencephalitis virus, phylogenetic analysis and comparative study of the 3' untranslated region of flaviviruses with no known vector.

Montana Myotis leukoencephalitis virus (MMLV), a virus isolated from bats, causes an encephalitis in small rodents reminiscent of flavivirus encephalitis in humans. The complete MMLV genome is 10690 nucleotides long and encodes a putative polyprotein of 3374 amino acids. The virus contains the same conserved motifs in genes that are believed to be interesting antiviral targets (NTPase/helicase, serine protease and RNA-dependent RNA polymerase) as flaviviruses of clinical importance. Phylogenetic analysis of the entire coding region has confirmed the classification of MMLV in the clade of the flaviviruses with no known vector (NKV) and within this clade to the Rio Bravo branch (both viruses have the bat as their vertebrate host). We have provided for the first time a comparative analysis of the RNA folding of the 3' UTR of the NKV flaviviruses (Modoc, Rio Bravo and Apoi viruses, in addition to MMLV). Structural elements in the 3' UTR that are preserved among other flaviviruses have been revealed, as well as elements that distinguish the NKV from the mosquito- and tick-borne flaviviruses. In particular, the pentanucleotide sequence 5' CACAG 3', which is conserved in all mosquito- and tick-borne flaviviruses, is replaced by the sequence 5' C(C/U)(C/U)AG 3' in the loop of the 3' long stable hairpin structure of all four NKV flaviviruses. The availability of this latter sequence motif allows us to designate a virus as either an NKV or a vector-borne flavivirus.

3' Untranslated Regions↗
PubMed2003

Complete genomic sequences of astroviruses from sheep and turkey: comparison with related viruses.

The genomes of astroviruses infecting sheep and turkey were sequenced. Detailed analyses of these sequences were performed, including comparison with the other complete astrovirus sequences available as well as with other RNA virus sequences, with focus on the non-structural proteins and RNA sequences. Earlier postulated functional astrovirus RNA motifs and protein domains could in most cases be recognised in the sheep and turkey astrovirus sequences. In addition, analyses of the available astrovirus sequences revealed: two protein regions with the potential for forming coiled coils, differences in the postulated transmembrane region, a similarity between the putative astrovirus nuclear localisation signal and calicivirus genome-linked proteins, and a stretch of a highly conserved RNA sequence with a possible role in the astrovirus capsid gene expression. The present analyses contribute to the deciphering of pertinent functions of the astrovirus genomes.

Amino Acid Sequence↗
PubMed2005

Complete genome sequence and biological characterizations of a novel goose paramyxovirus-SF02 isolated in China.

A paramyxovirus designated as APMV-1 (NDV) isolate SF02 (abbre. as SF02) was recently isolated from goose in China. SF02 was identified as a member of Newcastle disease virus (NDV) genotype VII. NDV strains are generally pathogenic only for fowls, including chicken and pigeon, and not for waterfowls such as goose and duck, whereas SF02 is highly pathogenic for both fowls and waterfowls. In the present study the complete genome consisting of 15, 192 nucleotides of SF02 was sequenced. Genomes of SF02 and all known APMV-1, Strains contain 6 ORFs in the order of NP-P-M-F-HN-L, and that of SF02 had an extra 6 nts between NP and P genes. Moreover, an anti-sense ORF consisting of 549 nt at the 1960 to 1412 and deduced 182 amino acids was found in SF02. The SF02 genome shared 83% identity and its 6 ORFs 81.9-86.1% identities with the reference APMV-1 strains. The possible mechanism determining different host range and pathogenicity is discussed based on genetic analyses.

Amino Acid Sequence↗
PubMed2026

Complete Genome Sequencing of Occult Hepatitis B Virus in Hemodialysis Patients Reveals Subgenotype D2 and Immune Escape Mutations in Bangladesh.

Hepatitis B virus (HBV) remains a major global health concern, and occult HBV infection (OBI) presents significant diagnostic and clinical challenges, particularly among hemodialysis (HD) patients. This study is aimed at characterizing complete HBV genomes from maintenance HD patients with OBI in Bangladesh to elucidate genetic features, mutational patterns, and clinical implications. Serum samples from two HBsAg-negative HD patients were screened by ELISA and quantitative PCR. Viral DNA was amplified by PCR across four overlapping open reading frames (ORFs) and sequenced on the Illumina platform. Genome assembly, phylogenetic analysis, and mutational profiling were performed using reference datasets and bioinformatics tools. Antigenicity and hydrophilicity of HBsAg were predicted in silico. Both patients were anti-HBc and anti-HBs positive with high HBV DNA loads (2.29 × 1010 and 2.53 × 1010 copies/mL). Full-length genomes (3182 bp) were successfully sequenced and phylogenetic analysis showed both HBV genomes clustered within Genotype D, Subgenotype D2, and subtype ayw3, consistent with previously reported Bangladeshi HBV genomes. Comparative mutational analysis identified substitutions such as T1753C in the basal core promoter, C1845T in preC, and D144E within the "a" determinant of HBsAg, suggesting potential roles in vaccine escape, immune escape, and diagnostic failure. Several nonsynonymous mutations were also detected in polymerase, though none were potentially associated with antiviral resistance. Antigenicity and hydrophilicity profiles of HBsAg and its major hydrophilic region remained largely conserved. These findings demonstrate the persistence of OBI in HD patients and provide an initial indication of the need for genomic surveillance to monitor immune-escape mutations and improve HBV diagnostic strategies in endemic regions.

HBV genome sequencing↗
PubMed2003

Complete genomic sequence of the Australian south-west genotype of Sindbis virus: comparisons with other Sindbis strains and identification of a unique deletion in the 3'-untranslated region.

Our previous studies have shown that two distinct genotypes of Sindbis (SIN) virus occur in Australia. One of these, the Oriental/Australian type, circulates throughout most of the Australian continent, whereas the recently identified south-west (SW) genetic type appears to be restricted to a distinct geographic region located in the temperate south-west of Australia. We have now determined the complete nucleotide and translated amino acid sequences of a SW isolate of SIN virus (SW6562) and performed comparative analyses with other SIN viruses at the genomic level. The genome of SW6562 is 11,569 nucleotides in length, excluding the cap nucleotide and poly (A) tail. Overall this virus differs from the prototype SIN virus (strain AR339) by 23% in nucleotide sequence and 12.5% in amino acid sequence. Partial sequences of four regions of the genome of four SW isolates were determined and compared with the corresponding sequences from a number of SIN isolates from different regions of the World. These regions are the non-structural protein (nsP3), the E2 gene, the capsid gene, and the repeated sequence elements (RSE) of the 3'UTR. These comparisons revealed that the SW SIN viruses were more closely related to South African and European strains than to other Australian isolates of SIN virus. Thus the SW genotype of SIN virus may have been introduced into this region of Australia by viremic humans or migratory birds and subsequently evolved independently in the region. The sequence data also revealed that the SW genotype contains a unique deletion in the RSE of the 3'UTR region of the genome. Previous studies have shown that deletions in this region of the SIN genome can have significant effects on virus replication in mosquito and avian cells, which may explain the restricted distribution of this genotype of SIN virus.

Animals↗
PubMed2002

Complete genome sequence of Clostridium perfringens, an anaerobic flesh-eater.

Clostridium perfringens is a Gram-positive anaerobic spore-forming bacterium that causes life-threatening gas gangrene and mild enterotoxaemia in humans, although it colonizes as normal intestinal flora of humans and animals. The organism is known to produce a variety of toxins and enzymes that are responsible for the severe myonecrotic lesions. Here we report the complete 3,031,430-bp sequence of C. perfringens strain 13 that comprises 2,660 protein coding regions and 10 rRNA genes, showing pronounced low overall G + C content (28.6%). The genome contains typical anaerobic fermentation enzymes leading to gas production but no enzymes for the tricarboxylic acid cycle or respiratory chain. Various saccharolytic enzymes were found, but many enzymes for amino acid biosynthesis were lacking in the genome. Twenty genes were newly identified as putative virulence factors of C. perfringens, and we found a total of five hyaluronidase genes that will also contribute to virulence. The genome analysis also proved an efficient method for finding four members of the two-component VirR/VirS regulon that coordinately regulates the pathogenicity of C. perfringens. Clearly, C. perfringens obtains various essential materials from the host by producing several degradative enzymes and toxins, resulting in massive destruction of the host tissues.

Amino Acids↗
PubMed2026

Complete genome sequence of the first alphahypovirus from the phytopathogenic fungus Curvularia americana on Poa pratensis in China.

In the present study, a novel positive-sense single-stranded RNA virus was identified from Curvularia americana strain 32 - 2, a phytopathogenic fungus infecting Poa pratensis in China, which was designated as "Curvularia americana hypovirus 1" (CaHV1). The complete genome of CaHV1 consists of 14,386 nucleotides (nt) with a single large open reading frame (ORF) (nt position 593 to 14035). The ORF encodes a putative polyprotein (4,480 amino acids) containing two potential conserved domains, which includes a Domain of Unknown Function 3525 (DUF3525) and Helicase. BLASTp analysis showed that amino acid sequence of the polyprotein of CaHV1 shared the highest amino acid sequence identity with that of Bipolaris oryzae hypovirus 1 (63.52%). Phylogenetic analysis based on amino acid sequence of the polyprotein indicated that CaHV1 clustered together with members of the genus Alphahypovirus within the family Hypoviridae. To the best of our knowledge, this is the first report of an alphahypovirus infecting C. americana.

Genome, Viral↗
PubMed1994

Complete genomic sequence and patterns of transcription of a member of an unusual family of closely related, chromosomally dispersed Ig gene clusters in Raja.

Ig genes in cartilaginous fish are organized in clusters. This unique form of organization suggests major differences in the regulation of the segmental rearrangement mechanism from that found in mammals and other higher vertebrates. The complete DNA sequence of an IgX-type cluster in the species Raja eglanteria is defined, and shown to consist of four rearranging segmental elements and three constant region exons. Using fluorescence in situ hybridization it is shown that Raja clusters are present at multiple sites within the genome, and that there is no apparent relationship between the chromosomally dispersed IgX clusters and a second isotype (IgM type) in this species. Comprehensive examination of sequence motifs associated with transcription regulation reveals an abundance of short sequences closely resembling those found in higher vertebrate Ig and other genes. However, the linear relationship of these motifs differs markedly from that associated with regulation of expression of the mammalian Ig gene locus. Additional studies of the transcription products of the IgX gene loci emphasize the diversity of transcription and processing of these genes. Considerable variation was noted in the processing of putative IgX transcripts, including the detection of a heretofore unrecognized form containing at least four additional Ig-like domains. These results have profound significance in terms of understanding the selective expression and the evolutionary diversification of Ig genes.

Animals↗
PubMed1999

Chromosomal localization and complete genomic sequence of the murine autoimmune regulator gene (Aire).

We have recently cloned the murine autoimmune regulator (Aire) gene, the homologue of human AIRE responsible for the autoimmune polyglandular syndrome type 1 (APS1) or autoimmune polyendocrinopathy candidiasis ectodermal dystrophy (APECED). Here, we report the genomic sequence (18,413 bp) for the entire Aire gene and its 5' flanking region, which contains putative regulatory sequences. Comparison of the genomic and cDNA sequences indicates that the Aire gene is composed of 14 exons and the coding sequence shares high similarities between mouse and human. The sizes of the homologous introns in the two species are conserved; however, the introns do not share significant sequence homologies except the sequences near the splice donor and acceptor sites. Sequence analyses of the 5' regulatory region and the complete coding region in three mouse strains (B6, NOD and SJL) did not reveal any sequence variation, suggesting sequence conservation between different inbred mouse strains. Using one of the six microsatellite markers identified by genomic sequencing and a B6 x Cast backcross mapping panel, we mapped the mouse Aire gene to chromosome 10, a syntenic region containing the Cdl18 and Pfkl genes on human chromosome 21q22.

Animals↗
PubMed2005

Complete genome sequence of the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1 and comparison with Pyrococcus genomes.

The genus Thermococcus, comprised of sulfur-reducing hyperthermophilic archaea, belongs to the order Thermococcales in Euryarchaeota along with the closely related genus Pyrococcus. The members of Thermococcus are ubiquitously present in natural high-temperature environments, and are therefore considered to play a major role in the ecology and metabolic activity of microbial consortia within hot-water ecosystems. To obtain insight into this important genus, we have determined and annotated the complete 2,088,737-base genome of Thermococcus kodakaraensis strain KOD1, followed by a comparison with the three complete genomes of Pyrococcus spp. A total of 2306 coding DNA sequences (CDSs) have been identified, among which half (1165 CDSs) are annotatable, whereas the functions of 41% (936 CDSs) cannot be predicted from the primary structures. The genome contains seven genes for probable transposases and four virus-related regions. Several proteins within these genetic elements show high similarities to those in Pyrococcus spp., implying the natural occurrence of horizontal gene transfer of such mobile elements among the order Thermococcales. Comparative genomics clarified that 1204 proteins, including those for information processing and basic metabolisms, are shared among T. kodakaraensis and the three Pyrococcus spp. On the other hand, among the set of 689 proteins unique to T. kodakaraensis, there are several intriguing proteins that might be responsible for the specific trait of the genus Thermococcus, such as proteins involved in additional pyruvate oxidation, nucleotide metabolisms, unique or additional metal ion transporters, improved stress response system, and a distinct restriction system.

Amino Acid Substitution↗
PubMed2004

Phylogenetic analysis of the complete genome sequence of Encephalitozoon cuniculi supports the fungal origin of microsporidia and reveals a high frequency of fast-evolving genes.

Microsporidia are unicellular eukaryotes living as obligate intracellular parasites. Lacking mitochondria, they were initially considered as having diverged before the endosymbiosis at the origin of mitochondria. That microsporidia were primitively amitochondriate was first questioned by the discovery of microsporidial sequences homologous to genes encoding mitochondrial proteins and then refuted by the identification of remnants of mitochondria in their cytoplasm. Various molecular phylogenies also cast doubt on the early divergence of microsporidia, these organisms forming a monophyletic group with or within the fungi. The 2001 proteins putatively encoded by the complete genome of Encephalitozoon cuniculi provided powerful data to test this hypothesis. Phylogenetic analysis of 99 proteins selected as adequate phylogenetic markers indicated that the E. cuniculi sequences having the lowest evolutionary rates preferentially clustered with fungal sequences or, more rarely, with both animal and fungal sequences. Because sequences with low evolutionary rates are less sensitive to the long-branch attraction artifact, we concluded that microsporidia are evolutionarily related to fungi. This analysis also allowed comparing the accuracy of several phylogenetic algorithms for a fast-evolving lineage with real rather than simulated sequences.

Animals↗
PubMed2005

Complete genome sequence and analysis of the multiresistant nosocomial pathogen Corynebacterium jeikeium K411, a lipid-requiring bacterium of the human skin flora.

Corynebacterium jeikeium is a "lipophilic" and multidrug-resistant bacterial species of the human skin flora that has been recognized with increasing frequency as a serious nosocomial pathogen. Here we report the genome sequence of the clinical isolate C. jeikeium K411, which was initially recovered from the axilla of a bone marrow transplant patient. The genome of C. jeikeium K411 consists of a circular chromosome of 2,462,499 bp and the 14,323-bp bacteriocin-producing plasmid pKW4. The chromosome of C. jeikeium K411 contains 2,104 predicted coding sequences, 52% of which were considered to be orthologous with genes in the Corynebacterium glutamicum, Corynebacterium efficiens, and Corynebacterium diphtheriae genomes. These genes apparently represent the chromosomal backbone that is conserved between the four corynebacteria. Among the genes that lack an ortholog in the known corynebacterial genomes, many are located close to transposable elements or revealed an atypical G+C content, indicating that horizontal gene transfer played an important role in the acquisition of genes involved in iron and manganese homeostasis, in multidrug resistance, in bacterium-host interaction, and in virulence. Metabolic analyses of the genome sequence indicated that the "lipophilic" phenotype of C. jeikeium most likely originates from the absence of fatty acid synthase and thus represents a fatty acid auxotrophy. Accordingly, both the complete gene repertoire and the deduced lifestyle of C. jeikeium K411 largely reflect the strict dependence of growth on the presence of exogenous fatty acids. The predicted virulence factors of C. jeikeium K411 are apparently involved in ensuring the availability of exogenous fatty acids by damaging the host tissue.

Anti-Bacterial Agents↗
PubMed2003

Chlorobium tepidum: insights into the structure, physiology, and metabolism of a green sulfur bacterium derived from the complete genome sequence.

Green sulfur bacteria are obligate, anaerobic photolithoautotrophs that synthesize unique bacteriochlorophylls (BChls) and a unique light-harvesting antenna structure, the chlorosome. One organism, Chlorobium tepidum, has emerged as a model for this group of bacteria primarily due to its relative ease of cultivation and natural transformability. This review focuses on insights into the physiology and biochemistry of the green sulfur bacteria that have been derived from the recently completed analysis of the 2.15-Mb genome of Chl. tepidum. About 40 mutants of Chl. tepidum have been generated within the last 3 years, most of which have been made based on analyses of the genome. This has allowed a nearly complete elucidation of the biosynthetic pathways for the carotenoids and BChls in Chl. tepidum, which include several novel enzymes specific for BChl c biosynthesis. Facilitating these analyses, both BChl c and carotenoid biosynthesis can be completely eliminated in Chl. tepidum. Based particularly on analyses of mutants lacking chlorosome proteins and BChl c, progress has also been made in understanding the structure and biogenesis of chlorosomes. In silico analyses of the presence and absence of genes encoding components involved in electron transfer reactions and carbon assimilation have additionally revealed some of the potential physiological capabilities, limitations, and peculiarities of Chl. tepidum. Surprisingly, some structural components and biosynthetic pathways associated with photosynthesis and energy metabolism in Chl. tepidum are more similar to those in cyanobacteria and plants than to those in other groups of photosynthetic bacteria.

Journal Article↗