Detection of lactococcal 936-species bacteriophages in whey by magnetic capture hybridization PCR targeting a variable region of receptor-binding protein genes

Abstract

To develop PCR assays able to distinguish between groups within lactococcal 936-species bacteriophages, as defined by their different receptor-binding protein (RBP) genes. DNA sequences of RBP genes from 17 lactococcal bacteriophages of the 936-species were compared, and six phage groups were identified. For each phage group a specific primer pair targeting a variable region of the RBP genes was designed. In nine of 20 whey samples, from dairies with recorded phage problems, between one and six phage groups were identified by conventional PCR. The sensitivity and specificity of the method was improved by magnetic capture hybridization (MCH)-PCR using a capture probe targeting an 80-bp highly conserved region just upstream from the RBP gene in all the investigated phages. The MCH-PCR was performed on 100 microl whey samples and the detection limit of the assay was 10(2)-10(3) PFU ml(-1) as opposed to the detection limit of 10(4) PFU ml(-1) for conventional PCR performed on 1-microl whey samples. In this study, PCR assays have been developed to detect six different types of RBP genes in lactococcal 936-species bacteriophages. The PCR assays have practical applications at cheese plants for detection of 936-species phages with different RBP and thereby potentially with different host ranges. This knowledge will make it possible to improve starter culture rotation systems in the dairy industry.