Construction ofa DNA ProbeandDetection ofActinobacillus pleuropneumoniae byUsingPolymerase ChainReaction

A 1.5-kb Actinobacillus pleuropneumoniae 4074DNA fragmentfrom a genomic librarywas found to hybridize toDNA fromthe12A. pleuropneumoniae serotypesby Southernhybridization. No crosshybridization was detected withDNAs fromhemolytic members ofthefamilyPasteurellaceae. From the nucleotide sequenceoftheputative genomicprobe,threeprimers were synthesized foruse inpolymerasechain reactions (PCRs), with31strains tested byusing purified andcrudeDNA targets.PCR amplification products of610and985bpwere observed innucleic acids extracted fromthe12knownserotypesanda biotype 2strain. Template DNAsfromother gram-negative andgram-positive bacteria, some ofthemfoundinthenormalflora ofswine andtheupper respiratory tract, were notamplified byPCR.Theonlyexception was an amplification ofa similar 610-or 985-bp sequence inActinobacillus lignieresii, a species thatisclosely relatedtoA. pleuropneumoniae butthathasneverbeenisolated fromswine. Amplification ofspecific A.pleuropneumoniae sequencesbyPCRdirectly fromclinical specimens may findapplications intheidentification ofasymptomatic carriers aswell asinefforts toeradicate porcine pleuropneumonia. Porcine pleuropneumonia, a major contagious respiratory disease inpigs, isdistributed worldwide andcauses severe economic losses totheswineindustry (30). Thedisease is characterized byfibrinous pleuritis, withhemorrhagic and necrotic lesions inthelungs(19); clinical symptomsand pathologic lesions foundatthetimeofnecropsyleadtoa clear diagnosis. Thecausal agentofporcine pleuropneumonia,Actinobacillus pleuropneumoniae, isa gram-negative