Raw Milk Microbiota Modifications as Affected by Chlorine Usage for Cleaning Procedures: The Trentingrana PDO Case

Abstract

Milk microbiota represents a key point in raw milk cheese production and contributes to the development of typical flavor and texture for each type of cheese. The aim of the present study was to evaluate the influence of chlorine products usage for cleaning and sanitizing the milking equipment on (i) raw milk microbiota; (ii) the deriving whey-starter microbiota; and (iii) Trentingrana Protected Designation of Origin (PDO) cheese microbiota and volatilome. Milk samples from three farms affiliated to a Trentingrana PDO cheese factory were collected three times per week during a 6-weeks period in which a sodium hypochlorite detergent (period C) was used and during a subsequent 6-weeks period of non-chlorine detergent usage (period NC). Samples were subjected to microbiological [Standard Plate Count; coliforms; coagulase-positive staphylococci; and lactic acid bacteria (LAB)] and metagenomic analysis (amplification of V3-V4 regions of 16S rRNA gene performed on Illumina MiSeq platform). In addition, cheese volatilome was determined by SPME-GC-MS. In the transition from period C to period NC, higher SPC and LAB counts in milk were recorded. Milk metagenomic analysis showed a peculiar distinctive microbiota composition for the three farms during the whole experimental period. Moreover, differences were highlighted comparing C and NC periods in each farm. A difference in microbial population related to chlorine usage in bulk milk and vat samples was evidenced. Moreover, chlorine utilization at farm level was found to affect the whey-starter population: the usually predominant Lactobacillus helveticus was significantly reduced during NC period, whereas Lactobacillus delbrueckii had the exact opposite trend. Alpha- and beta-diversity revealed a separation between the two treatment periods with a higher presence of L. helveticus , L. delbrueckii , and Streptococcus thermophilus in cheese samples after NC detergent period. Cheese volatilome analysis showed a slight decrease in lipolysis during C period in the inner part of the cheese wheel. Although preliminary, these results suggest a profound influence on milk and cheese microbiota, as well as on raw milk cheese production and quality, due to the use of chlorine. However, further studies will be needed to better understand the complex relationship between chlorine and microbiota along all the cheese production steps.

Microbial association networks in cheese: a meta-analysis

“…and a time series study on Cheddar (Choi et al, 2020) with an unusually high number of samples for each cheese/treatment and, unfortunately, a relatively low diversity due to the use of V4 region as a target and the use of pasteurized milk and defined strains starters, and ST157 (Afshari et al, 2020) which also is related to Cheddar cheese of different ages. Studies on very similar cheeses ST148, Parmigiano Reggiano (Bottari et al, 2020); ST23 Grana Padano (Bassi et al, 2015); Trentingrana (Cremonesi et al, 2020); Grana-like cheese (Alessandria et al, 2016) resulted in networks with very different properties: however, differences in study design may have obscured any similarities between the microbial communities of these cheeses. Some aspects of alpha or beta diversity appear to be related to network properties.…”

“…However, the majority of the associations was detected by three inference methods or more in at least one study and might be therefore relatively robust. Most associations were detected in only a few studies (Bassi et al, 2015; Calasso et al, 2016; Cremonesi et al, 2020; De Filippis et al, 2016; Dolci et al, 2014; Dugat-Bony et al, 2016; Kamimura et al, 2019; Marino et al, 2019; Stellato et al, 2015) (see Supplementary Table 2). Only two mutual exclusion associations are included, although more have been detected by at least 2 methods.…”

Effect of Different Farming Practices on Lactic Acid Bacteria Content in Cow Milk

“…This value, in itself positive, does not ensure a sufficient supply of lactic acid bacteria, with a negative impact on the quality of the cheese [ 7 ]. Recent studies highlighted that the milk microbial community at the farm level is affected by multiple factors that provide, on the whole, a peculiar distinctive microbiota composition for each farm, but further advances are needed to deepen the major drivers providing LAB prevalence and biodiversity in milk microbial population [ 8 ].…”

“…The same conclusion was reached in the study of Tormo et al [ 2 ] in goat dairy farms, where the authors found that certain farming practices such as type of milking parlor and the presence of hay in the bedding area could enable to decrease the contamination of enterococci and promote the development of L. lactis in milk, improving technological and sensorial quality of lactic cheeses. Moreover, Cremonesi et al [ 8 ] evidenced the influence of chlorine products’ usage in milking equipment on raw milk microbiota composition and biodiversity. It is worth considering that different studies [ 2 , 8 , 10 ] focused on the relation among bacterial diversity of milk obtained with phenotypic and genotypic methods and farm management practices, however only Mallet et al [ 1 ] also considered the quantitative characterization of milk microbiota.…”

“…Moreover, Cremonesi et al [ 8 ] evidenced the influence of chlorine products’ usage in milking equipment on raw milk microbiota composition and biodiversity. It is worth considering that different studies [ 2 , 8 , 10 ] focused on the relation among bacterial diversity of milk obtained with phenotypic and genotypic methods and farm management practices, however only Mallet et al [ 1 ] also considered the quantitative characterization of milk microbiota.…”