Detection and Characterization of Streptococcus thermophilus Bacteriophages by Use of the Antireceptor Gene Sequence

Abstract

In the dairy industry, the characterization of Streptococcus thermophilus phage types is very important for the selection and use of efficient starter cultures. The aim of this study was to develop a characterization system useful in phage control programs in dairy plants. A comparative study of phages of different origins was initially performed based on their morphology, DNA restriction profiles, DNA homology, structural proteins, packaging mechanisms, and lifestyles and on the presence of a highly conserved DNA fragment of the replication module. However, these traditional criteria were of limited industrial value, mainly because there appeared to be no correlation between these variables and host ranges. We therefore developed a PCR method to amplify VR2, a variable region of the antireceptor gene, which allowed rapid detection of S. thermophilus phages and classification of these phages. This method has a significant advantage over other grouping criteria since our results suggest that there is a correlation between typing profiles and host ranges. This association could be valuable for the dairy industry by allowing a rational starter rotation system to be established and by helping in the selection of more suitable starter culture resistance mechanisms. The method described here is also a useful tool for phage detection, since specific PCR amplification was possible when phage-contaminated milk was used as a template (detection limit, 10 5 PFU ml ؊1 ).In the dairy environment, lytic phages are the leading cause of fermentation failure during the manufacture of cheese and fermented milk products. Consequently, the success of commercial lactic starter cultures depends primarily on the selection of non-phage-related strains able to withstand viral infections. Streptococcus thermophilus, a gram-positive thermophilic lactic acid bacterium, is a component of dairy starter cultures used for the manufacture of yogurt and other fermented milk products. As observed for other dairy lactic acid bacteria, this streptococcus is often susceptible to phage attack, which can result in slow lactic acid fermentation and the loss of product quality. Besides its use as a starter in the yogurt industry, this species is very important in the production of a large variety of cheeses (26), from which many S. thermophilus phages responsible for fermentation failure have recently been isolated (32). Since a large dairy plant can processes more than 5 ϫ 10 5 liters of milk per day, phage problems can be very costly. The economic implications have led to intensive research into methods to control bacteriophage infections in the industry. The traditional strategies used to minimize the consequences of phage infection include rotation of non-phage-related strains (the principal precautionary measure employed), the use of phageinhibiting media for culture propagation, and aseptic processing conditions (14). However, the effectiveness is sometimes limited by the dynamic evolutionary properties of phages (7,22) and, in the case of S. ther...

Multiplex Fast Real-Time PCR for Quantitative Detection and Identification of cos - and pac -Type Streptococcus thermophilus Bacteriophages

“…(8), but these methods are time-consuming. To speed up the analysis, PCR techniques have been used to detect phages in different kinds of dairy samples (1,4,6,7,10,12). Increasing demand for quantitative, more sensitive, and quicker procedures is prompting the development of real-time quantitative PCR (qPCR) methods.…”

“…The specificity was assessed by testing 27 different S. thermophilus bacteriophages previously isolated in Europe from failed industrial fermentations and characterized in our laboratory (unpublished results), 15 different S. thermophilus phages isolated in America (1,13), and the type phages Sfi11 and Sfi21. Four different bacteriophages infecting Lactobacillus delbrueckii and nine bacteriophages infecting Lactococcus lactis were also tested.…”

Lactic Acid Bacteria Resistance to Bacteriophage and Prevention Techniques to Lower Phage Contamination in Dairy Fermentation

“…However, PCR is a fast method able to confirm the presence of bacteriophages within 30 minutes and can be applied to determine the potential utility and quality of big batches of milk. At the same time, the method could be handy in finding niches of phage accumulation, in order to reduce their impact in dairy fermentations [105][106][107][108].…”

The Characteristics, Detection and Control of Bacteriophage in Fermented Dairy Products

“…Raw or pasteurized milk is used in cheese manufacturing and many phages will resist pasteurization (Madera et al, 2004). Since a large dairy plant can process more than 500 tons of milk per day, a phage problem can lead to great economic loss (Binetti et al, 2005). A potential problem in a fermented batch is likely to occur if lactococcal phages exist in a population level of 10 5 to 10 7 PFU/mL in starter media or milk as this phage titer is able to retard milk acidification (Svenson and Christiansson, 1991).…”

“…The susceptibility of these bacteria to infection by bacteriophages that are omnipresent in the dairy environment can have serious economic consequences for the dairy industry (Binetti et al, 2005).…”