Establishment Of Real-Time Fluorescence Quantitative PCR For Rapid Detecting Brucella And Assemblage Of The Test Kit

Brucella are gram-negative bacteria that are pathogenic for humans and a varietyof livestock animals and wildlife. The genus Brucella has six recognized species onthe basis of host specificity : B.abortus , B.melitensis, B.suis, B.canis, B.ovis,B.neotomae. The six species have been subdivided into 19 biovar based on culture andbiochemical characteristics. The greatest economic impact result from bovinebrucellosis caused by B.abortus. Infection decreases reproductive efficiency mainly byabortion. B.abortus causes nearly all of the cattle abortions that result frombrucellosis . B.melitensis and B,suis infect cattle and apread within herds but rarelycause abortus. but the both cause brucellosis in humans.The diagnosis of brucellosis is currently based on conventional serological andmicrobiological tests. It is well know that serological methods are not always sensitiveor specific .Moreover, they have repeatedly been reported to cross-react withantigens other than those from brucella spp. Microbiological isolation andidentification are the most reliable methods of diagnosing of brucellosis. Howeverthese procedures are not always successful, are cumbersome,and represent a greatrisk of infection for laboratory technicians.Over the past decade,there have been major advancements in all aspects ofmolecular diagnostic with regard to human and animal brucellosis.PCR-based tests areproving to be faster and more sensitive then traditional method. This studyt designgenus-specific primerand and Taqman probe based on BCSP31 gene encoding31Kdantigenic peripasmic protein real time fluorescence quantitative PCR reaction systemand reaction condition. To establish FQ-PCR method for detecting all the Brucellaspecies and biovars. Additionally,this study developed three species-spcific real-timefluorescence quantitative PCR-based assays for rapidly, specificity, sensitivity,detecting Brucella abortus, Brucella melitensis, Brucella suis.In all of above assaycell as low as 1-5 copies it can be detected using Taqman probe.So real-time fluorescence quantitative PCR-based assays has been shown to be amore sensitive technique than bacterial culture and specific than conventionalserological tests for the examination of Brucella.