PCR detection of Mycobacterium avium subspecies paratuberculosis (MAP), introduction of a multitarget approach

Mycobacterium avium subspecies paratuberculosis (MAP) causes a wasting, and progressive infectious chronic enteritis principally in ruminants. MAP has brought extensive economic losses in livestock industries across the world. MAP is broadly known as a pathogen potentially involved in Crohn’s disease (CD). PCR-based molecular methods have recently achieved a global acceptance in detection of MAP. Some of these are now at par with conventional biochemical/pathological methods routinely used in OIE reference laboratories.In order to improve current methods and extend genomic markers suitable in routine laboratory tracing of MAP in Iran, we have selected the previously-reported genomic fragments of 16S rRNA، F57, ISMAV2، HspX and Locus255. New protocols and primer pairs were designed to enable amplification of these MAP specific fragments through individual single-round PCRs. Feasibly of an all-in-one system for amplification of the loci was then successfully, examined against a panel of Reference strains (MAP III&V and MAP 316F). We assume application of a multi-target detection system of MAP is best suited for diagnostics as uncertainties might apply to specificity of certain single target regions. Furthermore, single-round PCRs are more reliable than nested PCRs due to less contamination risk.

Keywords:

MAP , crohn’s disease , 16srRNA , ISMAV2 , HspX , Locus255