Abstract
BackgroundThe contamination of raw milk cheeses (St-Marcellin and Brie) from two plants in France was studied at several steps of production (raw milk, after addition of rennet - St-Marcellin - or after second maturation - Brie -, after removal from the mold and during ripening) using bifidobacteria as indicators of fecal contamination.ResultsBifidobacterium semi-quantitative counts were compared using PCR-RFLP and real-time PCR. B. pseudolongum were detected in 77% (PCR-RFLP; 1.75 to 2.29 log cfu ml-1) and 68% (real-time PCR; 2.19 to 2.73 log cfu ml-1) of St-Marcellin samples and in 87% (PCR-RFLP; 1.17 to 2.40 log cfu ml-1) of Brie cheeses samples. Mean counts of B. pseudolongum remained stable along both processes. Two other populations of bifidobacteria were detected during the ripening stage of St-Marcellin, respectively in 61% and 18% of the samples (PCR-RFLP). The presence of these populations explains the increase in total bifidobacteria observed during ripening. Further characterization of these populations is currently under process. Forty-eight percents (St-Marcellin) and 70% (Brie) of the samples were B. pseudolongum positive/E. coli negative while only 10% (St-Marcellin) and 3% (Brie) were B. pseudolongum negative/E. coli positive.ConclusionsThe increase of total bifidobacteria during ripening in Marcellin's process does not allow their use as fecal indicator. The presence of B. pseudolongum along the processes defined a contamination from animal origin since this species is predominant in cow dung and has never been isolated in human feces. B. pseudolongum was more sensitive as an indicator than E. coli along the two different cheese processes. B. pseudolongum should be used as fecal indicator rather than E. coli to assess the quality of raw milk and raw milk cheeses.
Highlights
The contamination of raw milk cheeses (St-Marcellin and Brie) from two plants in France was studied at several steps of production using bifidobacteria as indicators of fecal contamination
A PCR method based on the hsp60 gene, already sequenced in most Bifidobacterium species [16,17] was developed for a rapid detection of bifidobacteria in a raw milk cheese process
Validation of the PCR methods on pure strains The B. pseudolongum probe based on hsp60 gene was validated on 55 pure Bifidobacterium strains belonging to 13 different species (Table 1)
Summary
The contamination of raw milk cheeses (St-Marcellin and Brie) from two plants in France was studied at several steps of production (raw milk, after addition of rennet - St-Marcellin - or after second maturation - Brie -, after removal from the mold and during ripening) using bifidobacteria as indicators of fecal contamination. The genus Bifidobacterium represents one of the most important bacterial group in human and animal feces [1,2,3,4,5] This organism has stringent nutrient requirements and grows poorly outside of the animal gut, making this bacterial group a potentially useful indicator of fecal pollution as previously described [6]. An advantage in using bifidobacteria instead of other fecal contamination indicators is the host specificity, human or animal, of some groups of Bifidobacterium species [3] contrary to coliforms, which are ubiquitous [7]. A higher level of bifidobacteria was detected comparing to the level of E. coli suggesting that bifidobacteria could be a more convenient indicator This method did not allow the identification of the bifidobacteria species
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