Temporal and production-related determinants of microbial counts in pingo, the natural starter culture of minas artisanal cheese

Minas Artisanal Cheese (MAC) is among the most widely consumed and recognized artisanal cheeses in Brazil, produced from raw milk, rennet, salt, and a natural starter culture known locally as pingo. Pingo is an endogenous starter culture generated by backslopping, which carries into MAC cheese both technologically beneficial microorganisms and spoilage contaminants. Studies addressing the microbial diversity of MAC have accumulated in recent years, yet few have examined the composition of the microorganisms present in pingo or the factors that may influence microbial loads in this endogenous culture. Here we analysed 200 pingo samples from 50 cheesemaking units distributed across all MAC producing microregions, collected over four six-monthly campaigns. Lactic acid bacteria (LAB), total coliform, Staphylococcus aureus, and yeast/mould counts were estimated, and pH and NaCl concentration were determined. Linear mixed-effects models showed that LAB counts remained similar across sampling campaigns, although counts on MRS agar were associated with production scale and pingo volume. Total coliform counts in pingo were approximately tenfold higher during the wet season, whereas yeasts/moulds counts were associated with both season and sampling campaign. The effect of season on S. aureus depended on the year: counts did not differ between seasons in 2024 but were higher in the dry season in 2025. Smaller cheesemaking units additionally showed lower S. aureus counts in pingo. Milking infrastructure, pH, and NaCl concentration showed no association with the counts of any microbial group investigated. Our results revealed that the wet season is a period of elevated risk for the occurrence of potentially spoilage-causing microorganisms, irrespective of the scale of the cheesemaking unit and of the robustness of the installed milk production infrastructure.