eryC-5’ nuclease PCR: differentiating wildBrucella strains from vaccine strain S19

Abstract

Brucellosis is a zoonotic disease caused by the bacteria of the genus Brucella that produce infections leading to abortion and infertility, and recurrent fevers in humans. The disease is endemic in many areas of the world. Thus, we designed a TaqMan-based 5′nuclease-real-time PCR for molecular diagnosis by targeting the 702 bp deleted sequence of eryC gene from B. abortus S19 that allowed the specific quantitative detection of Brucella wild strains but not the vaccine strain B. abortus S19. The eryC gene which encodes the enzyme d-erythrulose-1-phosphate dehydrogenase that plays an important role in the ery thritol metabolism. This carbohydrate promotes the growth of some strains and is present in the placenta. The assay proved to be 100 per cent specific, as determined with Brucella isolates and reference Brucella strains, and highly sensitive with excellent linearity and PCR efficiency. When implemented on blood samples, the real time PCR assay detected higher proportion (80%) of positive samples than conventional bcsp31 PCR (70%) and i-ELISA (65%).