Identification of Actinobacillus pleuropneumoniae serotypes 1, 7, and 12 by multiplex PCR based on genes involved in encapsulation

Øystein Angen, Stine Graakjær Jessing, Peter Ahrens, T. Inzana

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    Abstract

    Based on differences in the capsular polysaccharides, 15 serotypes have until now been described for Actinobacillus pleuropneumoniae, the etiological agent of swine pleuropneumonia. Identification of the causative serotype is important both as a virulence marker and for epidemiological purposes. However, a number of isolates show cross-reaction between serotypes and this has urged the development of quick, serotype specific DNA-based methods necessary. Serotype specific tests have until now been described for the serotypes 2, 5, and 6 (Jessing et al, 2003) and serotypes 1, 2 and 8 (Schuchert et al., 2004). In the present work, serotype specific DNA regions involved in the biosynthesis of the capsular polysaccharides (cps region) were used to develop a multiplex PCR test for simultaneous species identification and serotyping of A. pleuropneumoniae serotypes 1, 7, and 12. Primers specific for serotypes 1, 7, and 12 were combined with an already existing species-specific PCR test based on the omlA gene. The PCR test was evaluated with the serotype reference strains of A. pleuropneumoniae as well as 165 Danish field isolates. For all strains investigated, a complete correspondence was found between results obtained with the multiplex PCR test and results from the traditional serotyping methods. The species specificity of the assay was evaluated using a collection of 93 strains representing 29 different species within the family Pasteurellaceae, as well as species normally found in the respiratory tract of swine. Also 45 field isolates of the phylogenetically closely related species A. lignieresii were tested. All these isolates tested negative for the cps-genes, except 5 isolates of A. lignieresii, which gave an amplicon of the same size as serotype 7 but without the species-specific omlAamplicon. Combined with an earlier published multiplex PCR test for serotypes 2, 5, and 6, we are now able to allocate approx. 99 % of Danish isolates of A. pleuropneumoniae to a serotype by PCR. Genetical determination of the serotype based on the cps-genes represents a convenient and specific method for serotyping A. pleuropneumoniae in diagnostic laboratories.
    Original languageEnglish
    Title of host publicationPasteurellaceae 2005
    Number of pages99
    VolumeB8
    Place of PublicationUnited States of America
    PublisherASM International
    Publication date2005
    Pages34-34
    ISBN (Print)1-55581-370-4
    Publication statusPublished - 2005
    EventASM Conference on Pasteurellaceae - Kohala Coast, HI, United States
    Duration: 23 Oct 200526 Oct 2005
    http://conferences.asm.org/index.php/2012-02-09-21-04-52/2012-02-09-21-03-56/9-past-conferences/69-asm-conference-on-pasteurellaceae

    Conference

    ConferenceASM Conference on Pasteurellaceae
    CountryUnited States
    CityKohala Coast, HI
    Period23/10/200526/10/2005
    Internet address

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