Detection methods for milk pathogenic bacteria by loop-mediated isothermal amplification

Abstract

Milk is a common food, which is consumed all over the world. It is an important source of calcium. Meanwhile, it provides abundant protein, minerals and vitamins. However, pathogenic bacteria which exist in milk not only causes nutrition loss, but also produces toxins which may cause diarrhea, food poisoning, and even death. In order to control the microbial level of raw milk and eliminate the contamination of materials, this assay applied loop-mediated isothermal amplification to explore a new way to detect enterotoxigenic Escherichia coli (ETEC) in raw milk. The best reaction condition in detecting ETEC from raw milk was confirmed to be: 0.016 μM each of forward outer primer (primer F3) and backward outer primer (primer B3), 0.128 μM each of forward inner primer (primer FIP) and backward inner primer (primer BIP), 0.45 μM deoxy-ribonucleoside triphosphate (dNTPs), 2IU Bst DNA polymerase large fragment and template DNA were incubated at 63°C for 60 min. LAMP was proved to be specific, rapid and sensitive in detecting pathogenic bacteria which exist in milk.