Rapid detection of Brachyspira hyodysenteriae and Lawsonia intracellularis in swine faecal and mucosal specimens by multiplex PCR

Multiplex polymerase chain reaction (M-PCR) was developed to determine its usefulness as a rapid and specific method for routine diagnosis of Brachyspira hyodysenteriae and Lawsonia intracellularis infections in pigs. The assay was based on simultaneous amplification of the tlyA-encoded haemolysin gene of B. hyodysenteriae and 16S rRNA sequences of L. intracellularis. The results of this study demonstrated that the M-PCR method is a useful tool for rapid diagnosis of the etiological agents only of acute forms of swine dysentery (SD) and proliferative enteropathy (PE). Screening of pooled diagnostic specimens from chronic form of PE was possible by using only our modified one tube nested PCR where detection limit was 1.1x10(4) bacterial cells per 0.1 g of faeces. No positive results were obtained when M-PCR for the identification of L. intracellularis from chronic forms of this disease was used. On the other hand, detection of B. hyodysenteriae in faeces from acute and chronic cases was both sensitive and specific by either single or M-PCR techniques. The sensitivity of the single and M-PCRs for the detection of B. hyodysenteriae in 10-fold dilutions of bacterial culture was estimated on the level of 10(-4). It was revealed that only two dilutions were countable: 10(-6) and 10(-7) corresponding to 203 and 14 CFU/ml, respectively.