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Conventional PCR Test for the Detection of Actinobacillus pleuropneumoniae


Código: ADI031-0. Categorías: , , .
   
Descripción

1. Purpose of the test
This qualitative PCR(a) test enables the detection of A. pleuropneumoniae and the typing of its
constituent strains into 4 groups. It may be used for the identification of colonies isolated on solid
media, or for the detection of A. pleuropneumoniae on a plate with a high level of contamination. A.
pleuropneumoniae can also be detected directly from tonsils.
2. Actinobacillus pleuropneumoniae
Actinobacillus pleuropneumoniae is the etiological agent of pig haemorrhagic pleuropneumonia, or
actinobacillosis. This disease is the cause of economic losses in a large number of industrial farms.
Actinobacillus pleuropneumoniae is a pleomorphic Gram negative coccobacillus from the
Pasteurellaceae family (Borr, 1991).
NAD requirements subdivide the A. pleuropneumoniae species into 2 distinct biovars: biovar 1 whose
constituent strains are NAD-dependent and biovar 2, whose constituent strains are NAD-independent.
Serotyping of A. pleuropneumoniae strains is based upon the organism’s capsular polysaccharide
antigens (Mittal et al., 1983). In this manner, 14 serotypes have been distinguished, serotypes 1 and 5
being subdivided into 1a, 1b and 5a, 5b respectively (Jolie et al., 1994 – Nielsen, 1986).
A. pleuropneumoniae is frequently isolated from the nasal cavities, from tonsils and from lungs.
It is now known that other bacteria belonging to the Pasteurellaceae family are also present in the
upper respiratory tract of pigs: Haemophilus parasuis (causative agent of Glasser’s disease),
Actinobacillus minor, Actinobacillus porcinus, Actinobacillus indolicus and Actinobacillus taxon C
(Moller et al., 1996).
All of these species share a certain number of biochemical properties with A. pleuropneumoniae, hence
rendering their identification somewhat difficult.
3. Test performance
This PCR test was assessed against a panel of 238 strains belonging to 32 bacterial species isolated from
pigs. Test sensitivity is of 100 % for biovar 1 strains and specificity is of 99 %.
4. Description and purpose of the test
This test is based on enzymatic gene amplification or PCR technique. It uses specific Actinobacillus
pleuropneumoniae primers. Lysis solutions enclosed or QIAamp® DNA mini kit (Qiagen, Hilden,
Germany) if samples contain inhibitors can be used for DNA extraction. Amplified products are
visualised on a 2% agarose gel.
A control DNA, referred as “internal control”, is present in each reaction in order to validate each
negative result.

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