Lactococcus lactis SK1virus Taxonomy and Development of High-Throughput qPCR Methods to Evaluate SK1virus Development in Dairies Using Undefined Starter Cultures
Research output: Book/Report › Ph.D. thesis › Research
Bacteriophages attacking the Lactic acid bacteria (LAB), used as starter cultures in the dairy industry, produce many problems due to increased processing time, low quality or even failure of fermentation, as well as huge economic burden worldwide. In the first part of this study, we dealt with taxonomy of SK1virus in genomic level. We collected 60 fully sequenced phages (11 phages were from public database; the rest were from our own collection) from the lactococcal Sk1virus genus (former known as the 936 phage group or 936 species), and analyzed the effect of using >95% ANI for species differentiation based on whole genome sequence and based on core genome sequence. The results showed that, if a simple >95 % ANI was used, a 35 new species of SK1virus would be generated almost, whenever a new phage genome was sequenced giving a very unstable taxonomy. Most of core genes from SK1virus phages were located in the conserved late-expressed region and a small part was located in the early- and middle- expressed regions. Using a >95% ANI for the core genes for designation of species still results in an unmanageable number of species. So both methods generated on average a new species almost for every second new sequence. This will mean generating an useless taxonomy below genus level, and we suggest that no species is assigned to the SK1virus genus based on whole genome or whole core genome using >95 % ANIb or that the 936 group should be assigned to the 936species. In the second part, we further investigated the abundance in the diversity of L. lactis and Leuconostoc phages in whey samples in a Danish dairy using named TK5 starter culture over 12 years. L. lactis phage from 936 species and P335 species were both appeared frequently in the whey samples. Based on receptor binding protein analysis, most of the 936 subgroups were predicted to attack the strains of L. lactis subsp. cremoris. The control sample (without any whey sample added) was dominated by 936 phage species, suggesting the possibility of evolution among these phages at any point during the 12-year period. Furthermore, the emergences of new phages were found over time indicating that they probably have developed from already existing phages or from lysogenic strains. In the last part, we finally present a high-throughput qPCR system to detect different groups of lactococcal bacteriophages in 936 species by their RBP. We compared the DNA sequences of RBP genes from a large number of recently sequenced phages belonging to the 936-species and correlated sequences with host-range. The host-range related groups were identified, and the groups a pair of qPCR specific primers were designed targeting the region of RBP genes.
|Publisher||Department of Food Science, Faculty of Science, University of Copenhagen|
|Publication status||Published - 2018|