Comparative Phenotypic and Molecular Genetic Profiling of Wild Lactococcus lactis subsp. lactis Strains of the L. lactis subsp. lactis and L. lactis subsp. cremoris Genotypes, Isolated from Starter-Free Cheeses Made of Raw Milk

Abstract

Twenty Lactococcus lactis strains with an L. lactis subsp. lactis phenotype isolated from five traditional cheeses made of raw milk with no added starters belonging to the L. lactis subsp. lactis and L. lactis subsp. cremoris genotypes (lactis and cremoris genotypes, respectively; 10 strains each) were subjected to a series of phenotypic and genetic typing methods, with the aims of determining their phylogenetic relationships and suitability as starters. Pulsed-field gel electrophoresis (PFGE) analysis of intact genomes digested with SalI and SmaI proved that all strains were different except for three isolates of the cremoris genotype, which showed identical PFGE profiles. Multilocus sequence typing (MLST) analysis using internal sequences of seven loci (namely, atpA, rpoA, pheS, pepN, bcaT, pepX, and 16S rRNA gene) revealed considerable intergenotype nucleotide polymorphism, although deduced amino acid changes were scarce. Analysis of the MLST data for the present strains and others from other dairy and nondairy sources showed that all of them clustered into the cremoris or lactis genotype group, by using both independent and combined gene sequences. These two groups of strains also showed distinctive carbohydrate fermentation and enzyme activity profiles, with the strains in the cremoris group showing broader profiles. However, the profiles of resistance/susceptibility to 16 antibiotics were very similar, showing no atypical resistance, except for tetracycline resistance in three identical cremoris genotype isolates. The numbers and concentrations of volatile compounds produced in milk by the strains belonging to these two groups were clearly different, with the cremoris genotype strains producing higher concentrations of more branched-chain, derived compounds. Together, the present results support the idea that the lactis and cremoris genotypes of phenotypic Lactococcus lactis subsp. lactis actually represent true subspecies. Some strains of the two subspecies in this study appear to be good starter candidates.Lactococcus lactis is a lactic acid bacterium (LAB) commonly dominant in milk and fermented dairy products. Not surprisingly, carefully selected strains of L. lactis are majority components of starter cultures for dairy fermentations (38). Worldwide, over 100 million metric tons of milk is transformed annually into dairy products using L. lactis starters, reflecting the industrial and thus economic importance of this organism (38). The growth of L. lactis in milk is associated with the rapid production of lactic acid, which provides flavor, assists in curd formation, prevents the growth of pathogenic and spoilage bacteria, and creates optimal biochemical conditions for ripening. Via their proteolytic and amino acid conversion pathways, lactococci further contribute to the final texture (moisture, softness) and flavor of dairy products (47). These functions determine the sensory quality, safety, and shelf life of fermented dairy products. L. lactis includes three subspecies (L. lactis subsp. cremoris, L...

Lactococcus lactis strains from raw ewe’s milk samples from the PDO Ossau-Iraty cheese area: levels, genotypic and technological diversity

“…cremoris showing a L. lactis subsp. lactis genotype have been reported (Fernández et al 2011). Phenotypic differences between genotypically similar strains of L. lactis have also been found, probably linked to differences in enzymatic expression (de la Plaza et al 2006).…”

“…These studies have highlighted the wide diversity which exists among wild strains, much greater than among commercial or reference strains (Sánchez et al 2000). Strain diversity can produce a wide range of genotypic profiles Fernández et al 2011) which are not always consistent with phenotypic profiles at subspecies or strain levels (de la Plaza et al 2006;Fernández et al 2011). This diversity can lead to a wide range of technological abilities of individual strains including flavor formation, acidifying activity, production of antimicrobial compounds, and phagic resistance.…”

“…lactis and L. lactis subsp. cremoris is based on a set of phenotypic characteristics (ability to grow at 40°C, in 4% NaCl and at pH 9.2, ability to ferment maltose and capacity to deaminate arginine) and is of great technological interest, but this distinction may be erroneous as it may vary among strains and hence is very strain dependent (Fernández et al 2011). As a consequence, various DNA-based techniques such as random amplified polymorphic DNA, pulse-field gel electrophoresis, repetitive sequence-based PCR (Rep-PCR), amplified fragment length polymorphism, multilocus sequence analysis, and restriction fragment length polymorphism have been used to distinguish L. lactis at subspecies and strain levels (Taïbi et al 2010).…”

Lactic acid bacteria activating innate immunity improve survival in bacterial infection model of silkworm

“…Figure 3). Api Zym test was also used for the identification of the strain 11/19-B1 and compared to previous data (18) from yogurt on ARS milk agar and CaCO 3 -MRS agar, Gram staining, and 16S rDNA sequencing were conducted.…”

Lactose and β-Glucosides Metabolism and Its Regulation in Lactococcus lactis: A Review

“…It is well known from sugar fermentation characteristics that L. lactis strains of different origin can utilize a variety of -glucosides (e.g., Aleksandrzak-Piekarczyk et al, 2011;Bardowski et al, 1995;Fernández et al, 2011;Siezen et al, 2008). The metabolic potential for catabolism of these sugars can be chromosomally encoded by more than one genetic system, as was shown for L. lactis IL1403.…”