Metataxonomic Profiling of Native and Starter Microbiota During Ripening of Gouda Cheese Made With Listeria monocytogenes-Contaminated Unpasteurized Milk

Abstract

Unpasteurized milk is used to produce aged artisanal cheeses, which presents a safety concern due to possible contamination with foodborne pathogens, especially Listeria monocytogenes. The objective of this study was to examine the composition of the bacterial community in unpasteurized milk used to prepare Gouda cheese artificially contaminated with L. monocytogenes (~1 log CFU/ml) and assess the community dynamics and their potential interaction with L. monocytogenes during a 90-day ripening period using targeted 16S rRNA sequencing. The diversity of bacterial taxa in three batches of unpasteurized milk was not significantly different, and the microbiomes were dominated by species of Lactococcus, Streptomyces, Staphylococcus, and Pseudomonas. The highest relative abundances were observed for Pseudomonas fluorescens (31.84–78.80%) and unidentified operational taxonomic units (OTUs) of Pseudomonas (7.56–45.27%). After manufacture, both with and without L. monocytogenes-contaminated unpasteurized milk, Gouda cheese was dominated by starter culture bacteria (including Lactococcus lactis subsp. cremoris, lactis, lactis bv. diacetylactis, and Streptococcus thermophilus), in addition to unassigned members in the taxa L. lactis and Streptococcus. During ripening there was an overall decrease in L. lactis abundance and an increase in the number of taxa with relative abundances >0.1%. After 90-day ripening, a total of 82 and 81 taxa were identified in the Gouda cheese with and without L. monocytogenes, respectively. Of the identified taxa after ripening, 31 (Gouda cheese with L. monocytogenes) and 56 (Gouda cheese without L. monocytogenes) taxa had relative abundances >0.1%; 31 were shared between the two types of Gouda cheese, and 25 were unique to the Gouda cheese without added L. monocytogenes. No unique taxa were identified in the Gouda cheese with the added L. monocytogenes. This study provides information on the dynamics of the bacterial community in Gouda cheese during ripening, both with and without the addition of L. monocytogenes.

A Snapshot, Using a Multi-Omic Approach, of the Metabolic Cross-Talk and the Dynamics of the Resident Microbiota in Ripening Cheese Inoculated with Listeria innocua

“…Through the changes in their own metabolism and the subsequent release of antimicrobial molecules, the lactic acid bacteria impact the growth of pathogens [ 70 ]. The dynamics of variation of the resident microbiota in control and L. innocua -inoculated cheese observed in our study are consistent with the findings of other research involving raw milk in the production of Gouda cheese [ 71 ]. The authors claimed that in the first phase of cheese production, the microbial populations were dominated by the starter microorganisms, whose main technological action was carried out with the lowering of the milk pH; during the maturation phase, the counts of these bacteria slightly decreased, and the bacteria were involved in other important enzymatic reactions such as lipolysis, proteolysis, and production of flavor compounds.…”

“…The authors claimed that in the first phase of cheese production, the microbial populations were dominated by the starter microorganisms, whose main technological action was carried out with the lowering of the milk pH; during the maturation phase, the counts of these bacteria slightly decreased, and the bacteria were involved in other important enzymatic reactions such as lipolysis, proteolysis, and production of flavor compounds. Furthermore, the authors stated that the inoculum of Listeria monocytogenes had no inhibitory effect on the relative abundance of starter populations, nor vice versa [ 71 ].…”

Fermented Foods, Health and the Gut Microbiome

“…While the majority of cheeses are produced from cow’s milk, cheese from sheep, goats, buffalo, camels, and even donkeys is manufactured [ 43 ]. Genera common to all raw milk include Lactococcus , Lactobacillus , Leuconostoc , Streptococcus , and Enterococcus , although certain varieties are more strongly associated with one type or another, which impacts the microbiome of a final product [ 44 , 45 , 46 ]. For example, while the predominant genera in cow’s milk include Pseudomonas , Bacillus , Lactococcus , and Acinetobacter , the genera most common to raw camel’s milk include Enterococcus , Lactococcus , and Pediococcus [ 47 , 48 ].…”

The Use of Unique, Environmental Lactic Acid Bacteria Strains in the Traditional Production of Organic Cheeses from Unpasteurized Cow’s Milk

“…The quality of the cheeses also showed the differences between the treatments, which were probably due to the addition of the different starter cultures. According to Salazar et al [ 26 ], the addition of a known starter culture to unpasteurized milk makes it dominant in the developing product. In addition, Vázquez-Velázquez et al [ 27 ] found that the use of a starter culture helped keep the total pathogenic microbes in pasteurized cheese below standard maximum values.…”