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1.
Genome sequence of Avery's virulent serotype 2 strain D39 of Streptococcus pneumoniae and comparison with that of unencapsulated laboratory strain R6
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Lanie JA Ng WL Kazmierczak KM Andrzejewski TM Davidsen TM Wayne KJ Tettelin H Glass JI Winkler ME 《Journal of bacteriology》2007,189(1):38-51
2.
Beghetto E Gargano N Ricci S Garufi G Peppoloni S Montagnani F Oggioni M Pozzi G Felici F 《FEMS microbiology letters》2006,262(1):14-21
Streptococcus pneumoniae is a causative agent of otitis media, pneumonia, meningitis and sepsis in humans. For the development of effective vaccines able to prevent pneumococcal infection, characterization of bacterial antigens involved in host immune response is crucial. In order to identify pneumococcal proteins recognized by host antibody response, we created an S. pneumoniae D39 genome library, displayed on lambda bacteriophage. The screening of such a library, with sera either from infected individuals or mice immunized with the S. pneumoniae D39 strain, allowed identification of phage clones carrying S. pneumoniae B-cell epitopes. Epitope-containing fragments within the families of the histidine-triad proteins (PhtE, PhtD), the choline-binding proteins (PspA, CbpD) and zinc metalloproteinase B (ZmpB) were identified. Moreover, library screening also allowed the isolation of phage clones carrying three distinct antigenic regions of a hypothetical pneumococcal protein, encoded by the ORF spr0075 in the R6 strain genome sequence. In this work, Spr0075 is first identified as an expressed S. pneumoniae gene product, having an antigenic function during infection. 相似文献
3.
A functional dlt operon, encoding proteins required for incorporation of d-alanine in teichoic acids in gram-positive bacteria, confers resistance to cationic antimicrobial peptides in Streptococcus pneumoniae
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Kovács M Halfmann A Fedtke I Heintz M Peschel A Vollmer W Hakenbeck R Brückner R 《Journal of bacteriology》2006,188(16):5797-5805
Streptococcus pneumoniae is one of the few species within the group of low-G +C gram-positive bacteria reported to contain no d-alanine in teichoic acids, although the dltABCD operon encoding proteins responsible for d-alanylation is present in the genomes of two S. pneumoniae strains, the laboratory strain R6 and the clinical isolate TIGR4. The annotation of dltA in R6 predicts a protein, d-alanine-d-alanyl carrier protein ligase (Dcl), that is shorter at the amino terminus than all other Dcl proteins. Translation of dltA could also start upstream of the annotated TTG start codon at a GTG, resulting in the premature termination of dltA translation at a stop codon. Applying a novel integrative translation probe plasmid with Escherichia coli 'lacZ as a reporter, we could demonstrate that dltA translation starts at the upstream GTG. Consequently, S. pneumoniae R6 is a dltA mutant, whereas S. pneumoniae D39, the parental strain of R6, and Rx, another derivative of D39, contained intact dltA genes. Repair of the stop codon in dltA of R6 and insertional inactivation of dltA in D39 and Rx yielded pairs of dltA-deficient and dltA-proficient strains. Subsequent phenotypic analysis showed that dltA inactivation resulted in enhanced sensitivity to the cationic antimicrobial peptides nisin and gallidermin, a phenotype fully consistent with those of dltA mutants of other gram-positive bacteria. In addition, mild alkaline hydrolysis of heat-inactivated whole cells released d-alanine from dltA-proficient strains, but not from dltA mutants. The results of our study suggest that, as in many other low-G+C gram-positive bacteria, teichoic acids of S. pneumoniae contain d-alanine residues in order to protect this human pathogen against the actions of cationic antimicrobial peptides. 相似文献
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Yuki Iwasaki Takashi Abe Kennosuke Wada Masae Itoh Toshimichi Ikemura 《DNA research》2011,18(2):125-136
Influenza virus poses a significant threat to public health, as exemplified by the recent introduction of the new pandemic strain H1N1/09 into human populations. Pandemics have been initiated by the occurrence of novel changes in animal sources that eventually adapt to human. One important issue in studies of viral genomes, particularly those of influenza virus, is to predict possible changes in genomic sequence that will become hazardous. We previously established a clustering method termed ‘BLSOM’ (batch-learning self-organizing map) that does not depend on sequence alignment and can characterize and compare even 1 million genomic sequences in one run. Strategies for comparing a vast number of genomic sequences simultaneously become increasingly important in genome studies because of remarkable progresses in nucleotide sequencing. In this study, we have constructed BLSOMs based on the oligonucleotide and codon composition of all influenza A viral strains available. Without prior information with regard to their hosts, sequences derived from strains isolated from avian or human sources were successfully clustered according to the hosts. Notably, the pandemic H1N1/09 strains have oligonucleotide and codon compositions that are clearly different from those of human seasonal influenza A strains. This enables us to infer future directional changes in the influenza A viral genome. 相似文献
6.
The nutritional requirement that Streptococcus pneumoniae has for the aminoalcohol choline as a component of teichoic and lipoteichoic acids appears to be exclusive to this prokaryote. A mutation in the tacF gene, which putatively encodes an integral membrane protein (possibly, a teichoic acid repeat unit transporter), has been recently identified as responsible for generating a choline-independent phenotype of S. pneumoniae (M. Damjanovic, A. S. Kharat, A. Eberhardt, A. Tomasz, and W. Vollmer, J. Bacteriol. 189:7105-7111, 2007). We now report that Streptococcus mitis can grow in choline-free medium, as previously illustrated for Streptococcus oralis. While we confirmed the finding by Damjanovic et al. of the involvement of TacF in the choline dependence of the pneumococcus, the genetic transformation of S. pneumoniae R6 by using S. mitis SK598 DNA and several PCR-amplified tacF fragments suggested that a minimum of two mutations were required to confer improved fitness to choline-independent S. pneumoniae mutants. This conclusion is supported by sequencing results also reported here that indicate that a spontaneous mutant of S. pneumoniae (strain JY2190) able to proliferate in the absence of choline (or analogs) is also a double mutant for the tacF gene. Microscopic observations and competition experiments during the cocultivation of choline-independent strains confirmed that a minimum of two amino acid changes were required to confer improved fitness to choline-independent pneumococcal strains when growing in medium lacking any aminoalcohol. Our results suggest complex relationships among the different regions of the TacF teichoic acid repeat unit transporter. 相似文献
7.
Cardaci A Papasergi S Midiri A Mancuso G Domina M Cariccio VL Mandanici F Galbo R Lo Passo C Pernice I Donato P Ricci S Biondo C Teti G Felici F Beninati C 《PloS one》2012,7(5):e36588
There is considerable interest in pneumococcal protein antigens capable of inducing serotype-independent immunoprotection and of improving, thereby, existing vaccines. We report here on the immunogenic properties of a novel surface antigen encoded by ORF spr1875 in the R6 strain genome. An antigenic fragment encoded by spr1875, designated R4, was identified using a Streptococcus pneumoniae phage displayed genomic library after selection with a human convalescent serum. Immunofluorescence analysis with anti-R4 antisera showed that Spr1875 was expressed on the surface of strains belonging to different serotypes. Moreover, the gene was present with little sequence variability in 27 different pneumococcal strains isolated worldwide. A mutant lacking Spr1875 was considerably less virulent than the wild type D39 strain in an intravenous mouse model of infection. Moreover, immunization with the R4 recombinant fragment, but not with the whole Spr1875 protein, induced significant protection against sepsis in mice. Lack of protection after immunization with the whole protein was related to the presence of immunodominant, non-protective epitopes located outside of the R4 fragment. In conclusion, our data indicate that Spr1875 has a role in pneumococcal virulence and is immunogenic. As the R4 fragment conferred immunoprotection from experimental sepsis, selected antigenic fragments of Spr1875 may be useful for the development of a pneumococcal protein-based vaccine. 相似文献
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J Hoskins W E Alborn J Arnold L C Blaszczak S Burgett B S DeHoff S T Estrem L Fritz D J Fu W Fuller C Geringer R Gilmour J S Glass H Khoja A R Kraft R E Lagace D J LeBlanc L N Lee E J Lefkowitz J Lu P Matsushima S M McAhren M McHenney K McLeaster C W Mundy T I Nicas F H Norris M O'Gara R B Peery G T Robertson P Rockey P M Sun M E Winkler Y Yang M Young-Bellido G Zhao C A Zook R H Baltz S R Jaskunas P R Rosteck P L Skatrud J I Glass 《Journal of bacteriology》2001,183(19):5709-5717
Streptococcus pneumoniae is among the most significant causes of bacterial disease in humans. Here we report the 2,038,615-bp genomic sequence of the gram-positive bacterium S. pneumoniae R6. Because the R6 strain is avirulent and, more importantly, because it is readily transformed with DNA from homologous species and many heterologous species, it is the principal platform for investigation of the biology of this important pathogen. It is also used as a primary vehicle for genomics-based development of antibiotics for gram-positive bacteria. In our analysis of the genome, we identified a large number of new uncharacterized genes predicted to encode proteins that either reside on the surface of the cell or are secreted. Among those proteins there may be new targets for vaccine and antibiotic development. 相似文献
10.
To analyse the reliability and accuracy of genotype analysis with high-density oligonucleotide microarrays, this method and other experimental approaches were used to analyse genomic DNA of two popular Saccharomyces cerevisiae laboratory strains. S288C was used for systematic sequencing of 'the' S. cerevisiae genome; CEN.PK113-7D is a popular strain for physiological studies and functional genomics. Random amplified polymorphic DNA, electrophoretic karyotyping and microarray analysis all indicated a high level of sequence similarity between the two strains. In the microarray analysis, as few as 288 (4.5%) of the ca. 6300 represented yeast genes were identified that yielded significantly different hybridisation intensities between the two strains. These could be classified as amplified, absent, or with sequence polymorphism in CEN.PK113-7D compared to S288C. A detailed analysis focused on the subset of 25 genes called absent in CEN.PK113-7D. Among these absent genes, 17 were clustered together on five chromosomes, mainly in subtelomeric regions. Thorough analysis of these regions by polymerase chain reaction (PCR) and restriction fragment length polymorphism confirmed the absence of these genes in CEN.PK113-7D. Surprisingly, three of these regions were not smaller in CEN.PK113-7D chromosomes, indicating that they may harbour unidentified and potentially new sequences. In addition, eight genes called absent by the microarrays were scattered over the chromosomes. Using diagnostic PCR most of these genes were actually found to be present in CEN.PK113-7D, but after sequencing were found to differ significantly at the DNA level from S288C, explaining the poor hybridisation to the arrays. Our results indicate that DNA microarrays are a powerful tool for determining genotypic similarity between different yeast strains. However, to obtain meaningful information at the individual gene level, this method should be backed up by additional techniques. 相似文献
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N. Luisa Hiller Azad Ahmed Evan Powell Darren P. Martin Rory Eutsey Josh Earl Benjamin Janto Robert J. Boissy Justin Hogg Karen Barbadora Rangarajan Sampath Shaun Lonergan J. Christopher Post Fen Z. Hu Garth D. Ehrlich 《PLoS pathogens》2010,6(9)
Although there is tremendous interest in understanding the evolutionary roles of horizontal gene transfer (HGT) processes that occur during chronic polyclonal infections, to date there have been few studies that directly address this topic. We have characterized multiple HGT events that most likely occurred during polyclonal infection among nasopharyngeal strains of Streptococcus pneumoniae recovered from a child suffering from chronic upper respiratory and middle-ear infections. Whole genome sequencing and comparative genomics were performed on six isolates collected during symptomatic episodes over a period of seven months. From these comparisons we determined that five of the isolates were genetically highly similar and likely represented a dominant lineage. We analyzed all genic and allelic differences among all six isolates and found that all differences tended to occur within contiguous genomic blocks, suggestive of strain evolution by homologous recombination. From these analyses we identified three strains (two of which were recovered on two different occasions) that appear to have been derived sequentially, one from the next, each by multiple recombination events. We also identified a fourth strain that contains many of the genomic segments that differentiate the three highly related strains from one another, and have hypothesized that this fourth strain may have served as a donor multiple times in the evolution of the dominant strain line. The variations among the parent, daughter, and grand-daughter recombinant strains collectively cover greater than seven percent of the genome and are grouped into 23 chromosomal clusters. While capturing in vivo HGT, these data support the distributed genome hypothesis and suggest that a single competence event in pneumococci can result in the replacement of DNA at multiple non-adjacent loci. 相似文献
13.
Crasta OR Folkerts O Fei Z Mane SP Evans C Martino-Catt S Bricker B Yu G Du L Sobral BW 《PloS one》2008,3(5):e2193
The Brucella abortus strain S19, a spontaneously attenuated strain, has been used as a vaccine strain in vaccination of cattle against brucellosis for six decades. Despite many studies, the physiological and molecular mechanisms causing the attenuation are not known. We have applied pyrosequencing technology together with conventional sequencing to rapidly and comprehensively determine the complete genome sequence of the attenuated Brucella abortus vaccine strain S19. The main goal of this study is to identify candidate virulence genes by systematic comparative analysis of the attenuated strain with the published genome sequences of two virulent and closely related strains of B. abortus, 9-941 and 2308. The two S19 chromosomes are 2,122,487 and 1,161,449 bp in length. A total of 3062 genes were identified and annotated. Pairwise and reciprocal genome comparisons resulted in a total of 263 genes that were non-identical between the S19 genome and any of the two virulent strains. Amongst these, 45 genes were consistently different between the attenuated strain and the two virulent strains but were identical amongst the virulent strains, which included only two of the 236 genes that have been implicated as virulence factors in literature. The functional analyses of the differences have revealed a total of 24 genes that may be associated with the loss of virulence in S19. Of particular relevance are four genes with more than 60 bp consistent difference in S19 compared to both the virulent strains, which, in the virulent strains, encode an outer membrane protein and three proteins involved in erythritol uptake or metabolism. 相似文献
14.
To find potential targets of novel antimicrobial agents, we identified essential genes of Streptococcus pneumoniae using comparative genomics and allelic replacement mutagenesis. We compared the genome of S. pneumoniae R6 with those of Bacillus subtilis, Enterococcus faecalis, Escherichia coli, and Staphylococcus aureus, and selected 693 candidate target genes with > 40% amino acid sequence identity to the corresponding genes in at least two of the other species. The 693 genes were disrupted and 133 were found to be essential for growth. Of these, 32 encoded proteins of unknown function, and we were able to identify orthologues of 22 of these genes by genomic comparisons. The experimental method used in this study is easy to perform, rapid and efficient for identifying essential genes of bacterial pathogens. 相似文献
15.
Fouts DE Tyler HL DeBoy RT Daugherty S Ren Q Badger JH Durkin AS Huot H Shrivastava S Kothari S Dodson RJ Mohamoud Y Khouri H Roesch LF Krogfelt KA Struve C Triplett EW Methé BA 《PLoS genetics》2008,4(7):e1000141
We report here the sequencing and analysis of the genome of the nitrogen-fixing endophyte, Klebsiella pneumoniae 342. Although K. pneumoniae 342 is a member of the enteric bacteria, it serves as a model for studies of endophytic, plant-bacterial associations due to its efficient colonization of plant tissues (including maize and wheat, two of the most important crops in the world), while maintaining a mutualistic relationship that encompasses supplying organic nitrogen to the host plant. Genomic analysis examined K. pneumoniae 342 for the presence of previously identified genes from other bacteria involved in colonization of, or growth in, plants. From this set, approximately one-third were identified in K. pneumoniae 342, suggesting additional factors most likely contribute to its endophytic lifestyle. Comparative genome analyses were used to provide new insights into this question. Results included the identification of metabolic pathways and other features devoted to processing plant-derived cellulosic and aromatic compounds, and a robust complement of transport genes (15.4%), one of the highest percentages in bacterial genomes sequenced. Although virulence and antibiotic resistance genes were predicted, experiments conducted using mouse models showed pathogenicity to be attenuated in this strain. Comparative genomic analyses with the presumed human pathogen K. pneumoniae MGH78578 revealed that MGH78578 apparently cannot fix nitrogen, and the distribution of genes essential to surface attachment, secretion, transport, and regulation and signaling varied between each genome, which may indicate critical divergences between the strains that influence their preferred host ranges and lifestyles (endophytic plant associations for K. pneumoniae 342 and presumably human pathogenesis for MGH78578). Little genome information is available concerning endophytic bacteria. The K. pneumoniae 342 genome will drive new research into this less-understood, but important category of bacterial-plant host relationships, which could ultimately enhance growth and nutrition of important agricultural crops and development of plant-derived products and biofuels. 相似文献
16.
DNA/DNA genome microarray analysis together with genome sequencing suggests that the genome of members of the genus Streptomyces would seem to have a common structure including a linear genomic structure, a core of common syntenous Actinomycete genes, the presence of species specific terminal regions and two intermediate group of syntenous genes that seem to be genus specific. We analyzed Streptomyces species using DNA/DNA microarray comparative genome analysis. Only Streptomyces rimosus failed to give a congruent genome pattern for the genes found in Streptomyces coelicolor. We expanded the analysis to include a number of strains related to the type strain of S. rimosus and obtained a similar divergence from the main body of Streptomyces species. These strains showed very close identity to the original strain with no gene deletion or duplication detected. The 16S rRNA sequences of these S. rimosus strains were confirmed as very similar to the S. rimosus sequences available from the Ribosomal Database Project. When the SSU ribosomal RNA phylogeny of S. rimosus is analyzed, the species is positioned at the edge of the Streptomyces clade. We conclude that S. rimosus represents a distinct evolutionary lineage making the species a worthy possibility for genome sequencing. 相似文献
17.
Evaluating gene expression in C57BL/6J and DBA/2J mouse striatum using RNA-Seq and microarrays 总被引:1,自引:0,他引:1
Bottomly D Walter NA Hunter JE Darakjian P Kawane S Buck KJ Searles RP Mooney M McWeeney SK Hitzemann R 《PloS one》2011,6(3):e17820
C57BL/6J (B6) and DBA/2J (D2) are two of the most commonly used inbred mouse strains in neuroscience research. However, the only currently available mouse genome is based entirely on the B6 strain sequence. Subsequently, oligonucleotide microarray probes are based solely on this B6 reference sequence, making their application for gene expression profiling comparisons across mouse strains dubious due to their allelic sequence differences, including single nucleotide polymorphisms (SNPs). The emergence of next-generation sequencing (NGS) and the RNA-Seq application provides a clear alternative to oligonucleotide arrays for detecting differential gene expression without the problems inherent to hybridization-based technologies. Using RNA-Seq, an average of 22 million short sequencing reads were generated per sample for 21 samples (10 B6 and 11 D2), and these reads were aligned to the mouse reference genome, allowing 16,183 Ensembl genes to be queried in striatum for both strains. To determine differential expression, 'digital mRNA counting' is applied based on reads that map to exons. The current study compares RNA-Seq (Illumina GA IIx) with two microarray platforms (Illumina MouseRef-8 v2.0 and Affymetrix MOE 430 2.0) to detect differential striatal gene expression between the B6 and D2 inbred mouse strains. We show that by using stringent data processing requirements differential expression as determined by RNA-Seq is concordant with both the Affymetrix and Illumina platforms in more instances than it is concordant with only a single platform, and that instances of discordance with respect to direction of fold change were rare. Finally, we show that additional information is gained from RNA-Seq compared to hybridization-based techniques as RNA-Seq detects more genes than either microarray platform. The majority of genes differentially expressed in RNA-Seq were only detected as present in RNA-Seq, which is important for studies with smaller effect sizes where the sensitivity of hybridization-based techniques could bias interpretation. 相似文献
18.
Zhang Q Melcher U Zhou L Najar FZ Roe BA Fletcher J 《Applied and environmental microbiology》2005,71(12):7716-7723
Cucurbit yellow vine disease (CYVD) is caused by disease-associated Serratia marcescens strains that have phenotypes significantly different from those of nonphytopathogenic strains. To identify the genetic differences responsible for pathogenicity-related phenotypes, we used a suppressive subtractive hybridization (SSH) strategy. S. marcescens strain Z01-A, isolated from CYVD-affected zucchini, was used as the tester, whereas rice endophytic S. marcescens strain R02-A (IRBG 502) was used as the driver. SSH revealed 48 sequences, ranging from 200 to 700 bp, that were present in Z01-A but absent in R02-A. Sequence analysis showed that a large proportion of these sequences resembled genes involved in synthesis of surface structures. By construction of a fosmid library, followed by colony hybridization, selection, and DNA sequencing, a phage gene cluster and a genome island containing a fimbrial-gene cluster were identified. Arrayed dot hybridization showed that the conservation of subtracted sequences among CYVD pathogenic and nonpathogenic S. marcescens strains varied. Thirty-four sequences were present only in pathogenic strains. Primers were designed based on one Z01-A-specific sequence, A79, and used in a multiplex PCR to discriminate between S. marcescens strains causing CYVD and those from other ecological niches. 相似文献
19.
Carlos H Luna‐Flores Robin W Palfreyman Jens O Krömer Lars K Nielsen Esteban Marcellin 《Biotechnology journal》2017,12(2)
Traditionally derived from fossil fuels, biological production of propionic acid has recently gained interest. Propionibacterium species produce propionic acid as their main fermentation product. Production of other organic acids reduces propionic acid yield and productivity, pointing to by‐products gene‐knockout strategies as a logical solution to increase yield. However, removing by‐product formation has seen limited success due to our inability to genetically engineer the best producing strains (i.e. Propionibacterium acidipropionici). To overcome this limitation, random mutagenesis continues to be the best path towards improving strains for biological propionic acid production. Recent advances in next generation sequencing opened new avenues to understand improved strains. In this work, we use genome shuffling on two wild type strains to generate a better propionic acid producing strain. Using next generation sequencing, we mapped the genomic changes leading to the improved phenotype. The best strain produced 25% more propionic acid than the wild type strain. Sequencing of the strains showed that genomic changes were restricted to single point mutations and gene duplications in well‐conserved regions in the genomes. Such results confirm the involvement of gene conversion in genome shuffling as opposed to long genomic insertions. 相似文献
20.
Key to Pseudomonas aeruginosa 's ability to thrive in a diversity of niches is the presence of numerous genomic islands that confer adaptive traits upon individual strains. We reasoned that P. aeruginosa strains capable of surviving in the harsh environments of multiple hosts would therefore represent rich sources of genomic islands. To this end, we identified a strain, PSE9, that was virulent in both animals and plants. Subtractive hybridization was used to compare the genome of PSE9 with the less virulent strain PAO1. Nine genomic islands were identified in PSE9 that were absent in PAO1; seven of these had not been described previously. One of these seven islands, designated P. aeruginosa genomic island (PAGI)-5, has already been shown to carry numerous interesting ORFs, including several required for virulence in mammals. Here we describe the remaining six genomic islands, PAGI-6, -7, -8, -9, -10, and -11, which include a prophage element and two Rhs elements. 相似文献