Lactococcus lactis Diversity in Undefined Mixed Dairy Starter Cultures as Revealed by Comparative Genome Analyses and Targeted Amplicon Sequencing of epsD

Abstract

Undefined mesophilic mixed (DL) starter cultures are used in the production of continental cheeses and contain unknown strain mixtures of and leuconostocs. The choice of starter culture affects the taste, aroma, and quality of the final product. To gain insight into the diversity of strains in starter cultures, we whole-genome sequenced 95 isolates from three different starter cultures. Pan-genomic analyses, which included 30 publically available complete genomes, grouped the strains into 21 subsp and 28 subsp. lineages. Only one of the 95 isolates grouped with previously sequenced strains, and the three starter cultures showed no overlap in lineage distributions. The culture diversity was assessed by targeted amplicon sequencing using , a core gene, and, present in 93 of the 95 starter culture isolates but absent in most of the reference strains. This enabled an unprecedented discrimination of starter culture and revealed substantial differences between the three starter cultures and compositional shifts during the cultivation of cultures in milk. In contemporary cheese production, standardized frozen seed stock starter cultures are used to ensure production stability, reproducibility, and quality control of the product. The dairy industry experiences significant disruptions of cheese production due to phage attacks, and one commonly used countermeasure to phage attack is to employ a starter rotation strategy, in which two or more starters with minimal overlap in phage sensitivity are used alternately. A culture-independent analysis of the lactococcal diversity in complex undefined starter cultures revealed large differences between the three starter cultures and temporal shifts in lactococcal composition during the production of bulk starters. A better understanding of the lactococcal diversity in starter cultures will enable the development of more robust starter cultures and assist in maintaining the efficiency and stability of the production process by ensuring the presence of key bacteria that are important to the characteristics of the product.

Microbial Community Structure and Diversity of Shrimp Paste at Different Fermentation Stages

“…In addition, various antibacterial substances such as organic acids, diacetyl, hydrogen peroxide, and bacteriocins produced by Lactococcus may prevent the growth of food spoilage bacteria and extend the shelf life of shrimp paste. Lactococcus may thus improve the functional attributes and shelf life of shrimp paste (43–45).…”

Unprecedented Diversity of Lactococcal Group 936 Bacteriophages Revealed by Amplicon Sequencing of the Portal Protein Gene

“…Core genes from these 204 genomes were searched for variable regions suitable for diversity analyses by targeted amplicon sequencing using the same approach as described previously [9].…”

“…In a previous work we were able to identify up to 31 lactococcal bacterial lineages (OTUs) in a mixed starter culture [9]. The much higher richness of phage OTUs clearly indicates that many of the strains were infected by more than one phage, possibly at the same time, allowing for genetic recombination between different 936 phages [26].…”

“…Phages can be quantified by plaque testing, but this is laborious and for an undefined mixed culture such testing would require the isolation of pure host strains for all phages. Such strain isolation itself is likely to be difficult as not all strains are isolated by plating (or culturable) on standard laboratory media [6,9].…”

Microbial characterization of an artisanal production of Robiola di Roccaverano cheese

“…While the depth of sampling within each sample was limited, only two species were detected in the While the microbial composition of NSC used in the production of Robiola di Roccaverano was not previously investigated, several studies have been conducted on undefined starter culture applied in cheese-manufacturing process by means of metagenomic approach (Erkus et al, 2013;Frantzen et al, 2018). For example, the microbial community of undefined Gouda cheese starter was investigated and the metagenome revealed that in this starter culture L. lactis was dominant followed by a small community of L. mesenteroides (Erkus et al, 2013).…”