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Transcriptomic analysis comparing the spontaneous acapsular strain AL1114 to the parent capsulated strain AL1115

ABSTRACT: P. multocida is the causative agent of a wide range of diseases of animals, including fowl cholera in birds. Fowl cholera isolates of P. multocida generally express a capsular polysaccharide composed of hyaluronic acid. There have been reports of spontaneous capsule loss in P. multocida fowl cholera-causing strains but the mechanism by which this occurs has not been determined. In this study, we identified three independent strains that had spontaneously lost the ability to produce capsular polysaccharide. Quantitative RT-PCR showed that these strains had significantly reduced transcription of the capsule biosynthetic genes, but DNA sequence analysis identified no mutations within the cap biosynthetic locus. However, whole genome sequencing of paired capsulated and acapsular strains identified a single nucleotide polymorphism within fis that was present only in the acapsular strain. Sequencing of fis from two independently derived spontaneous acapsular strains showed that each contained a mutation within fis. Complementation of these strains with an intact copy of fis returned normal capsule expression to all strains. Therefore, expression of a functional Fis protein is absolutely required for normal capsule expression in P. multocida.DNA microarray studies comparing one of the acapsular pairs (AL114 to AL1115) identified approximately 30 genes as down-regulated in the mutant; including pfh_B2 which encodes the filamentous hemagglutinin, a known P. multocida virulence factor and the cross protective surface antigen plpE.

ORGANISM(S): Pasteurella multocida Pasteurella multocida subsp. multocida str. Pm70

PROVIDER: GSE17686 | GEO | 2010/05/24

SECONDARY ACCESSION(S): PRJNA118421

REPOSITORIES: GEO

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Project description:Background. Pneumococcus is a major human pathogen and the polysaccharide capsule is considered its main virulence factor. Nevertheless, strains lacking a capsule, named non-typeable pneumococcus (NT), are maintained in nature and frequently colonise the human nasopharynx. Interest in these strains, not targeted by any of the currently available pneumococcal vaccines, has been rising as they seem to play an important role in the evolution of the species. Currently, there is a paucity of data regarding this group of pneumococci. Also, questions have been raised on whether they are true pneumococci. We aimed to obtain insights in the genetic content of NT and the mechanisms leading to non-typeability and to genetic diversity. Methods. A collection of 52 NT isolates representative of the lineages circulating in Portugal between 1997 and 2007, as determined by pulsed-field gel electrophoresis and multilocus sequence typing, was analysed. The capsular region was sequenced and comparative genomic hybridisation (CGH) using a microarray covering the genome of 10 pneumococcal strains was carried out. The presence of mobile elements was investigated as source of intraclonal variation. Results. NT circulating in Portugal were found to have similar capsular regions, of cps type NCC2, i.e., having aliB-like ORF1 and aliB-like ORF2 genes. The core genome of NT was essentially similar to that of encapsulated strains. Also, competence genes and most virulence genes were present. The few virulence genes absent in all NT were the capsular genes, type-I and type-II pili, choline-binding protein A (cbpA/pspC), and pneumococcal surface protein A (pspA). Intraclonal variation could not be entirely explained by the presence of prophages and other mobile elements. Conclusions. NT circulating in Portugal are a homogeneous group belonging to cps type NCC2. Our observations support the theory that they are bona-fide pneumococcal isolates that do not express the capsule but are otherwise essentially similar to encapsulated pneumococci. Thus we propose that NT should be routinely identified and reported in surveillance studies.

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Transcriptomic analysis comparing the spontaneous acapsular strain AL1114 to the parent capsulated strain AL1115

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