Real time detection of enterococci in dairy foods using spore germination based bioassay

摘要

Real time detection of Enterococci in dairy foods using spore germination based bioassay The present invention relates to a spore germination based assay for the detection of microorganism Enterococci in a milk sample. The said assay comprises interaction of marker enzyme and non specific substrate resulting into the production of a germinant stimulus, which in turn indicates the presence or absence of target microorganism in a milk sample. Particularly, the invention enables the rapid detection of Enterococci which is based on targeting β-D-glucosidase as marker enzyme and its specific action on non specific substrate i.e. esculin / or cellobiose / or CMC resulting in germinant stimulus for spores produced by specific strain of B. megaterium. The assay is highly rapid and sensitive as the said assay enables the detection of target microorganism i.e. Enterococci in 8 ±2 hrs including sample enrichment in SAEBM (without esculin & ferric ammonium citrate) up to 4.5 ± 0.5 hrs and is capable of detecting the sample comprising as low as 5.5 ± 0.4 log cells. SAEBM has its application as selective media & micro-technique for detection and enumeration of Enterococci in milk sample. The assay time of micro-technique is ≤ 12 hrs with sensitivity up to 2-3 log count /ml. The invention also provides a sodium azide and esculin based culture media (SAEBM) which enables the selective enrichment of the target microorganism within 5-12 hrs.