Herve cheese is a Belgian soft cheese with a washed rind, and is made from raw or pasteurized milk. The specific microbiota of this cheese has never previously been fully explored and the use of raw or pasteurized milk in addition to starters is assumed to affect the microbiota of the rind and the heart. The aim of the study was to analyze the bacterial microbiota of Herve cheese using classical microbiology and a metagenomic approach based on 16S ribosomal DNA pyrosequencing. Using classical microbiology, the total counts of bacteria were comparable for the 11 samples of tested raw and pasteurized milk cheeses, reaching almost 8 log cfu/g. Using the metagenomic approach, 207 different phylotypes were identified. The rind of both the raw and pasteurized milk cheeses was found to be highly diversified. However, 96.3 and 97.9% of the total microbiota of the raw milk and pasteurized cheese rind, respectively, were composed of species present in both types of cheese, such as Corynebacterium casei, Psychrobacter spp., Lactococcus lactis ssp. cremoris, Staphylococcus equorum, Vagococcus salmoninarum, and other species present at levels below 5%. Brevibacterium linens were present at low levels (0.5 and 1.6%, respectively) on the rind of both the raw and the pasteurized milk cheeses, even though this bacterium had been inoculated during the manufacturing process. Interestingly, Psychroflexus casei, also described as giving a red smear to Raclette-type cheese, was identified in small proportions in the composition of the rind of both the raw and pasteurized milk cheeses (0.17 and 0.5%, respectively). In the heart of the cheeses, the common species of bacteria reached more than 99%. The main species identified were Lactococcus lactis ssp. cremoris, Psychrobacter spp., and Staphylococcus equorum ssp. equorum. Interestingly, 93 phylotypes were present only in the raw milk cheeses and 29 only in the pasteurized milk cheeses, showing the high diversity of the microbiota. Corynebacterium casei and Enterococcus faecalis were more prevalent in the raw milk cheeses, whereas Psychrobacter celer was present in the pasteurized milk cheeses. However, this specific microbiota represented a low proportion of the cheese microbiota. This study demonstrated that Herve cheese microbiota is rich and that pasteurized milk cheeses are microbiologically very close to raw milk cheeses, probably due to the similar manufacturing process. The characterization of the microbiota of this particular protected designation of origin cheese was useful in enabling us to gain a better knowledge of the bacteria responsible for the character of this cheese.

Milk kefir is produced by fermenting milk in the presence of kefir grains. This beverage has several benefits for human health. The aim of this experiment was to analyze 5 kefir grains (and their products) using a targeted metagenetic approach. Of the 5 kefir grains analyzed, 1 was purchased in a supermarket, 2 were provided by the Ministry of Agriculture (Namur, Belgium), and 2 were provided by individuals. The metagenetic approach targeted the V1-V3 fragment of the 16S ribosomal (r)DNA for the grains and the resulting beverages at 2 levels of grain incorporation (5 and 10%) to identify the bacterial species population. In contrast, the 26S rDNA pyrosequencing was performed only on kefir grains with the aim of assessing the yeast populations. In parallel, pH measurements were performed on the kefir obtained from the kefir grains using 2 incorporation rates. Regarding the bacterial population, 16S pyrosequencing revealed the presence of 20 main bacterial species, with a dominance of the following: Lactobacillus kefiranofaciens, Lactococcus lactis ssp. cremoris, Gluconobacter frateurii, Lactobacillus kefiri, Acetobacter orientalis, and Acetobacter lovaniensis. An important difference was noticed between the kefir samples: kefir grain purchased from a supermarket (sample E) harbored a much higher proportion of several operational taxonomic units of Lactococcus lactis and Leuconostoc mesenteroides. This sample of grain was macroscopically different from the others in terms of size, apparent cohesion of the grains, structure, and texture, probably associated with a lower level of Lactobacillus kefiranofaciens. The kefir (at an incorporation rate of 5%) produced from this sample of grain was characterized by a lower pH value (4.5) than the others. The other 4 samples of kefir (5%) had pH values above 5. Comparing the kefir grain and the kefir, an increase in the population of Gluconobacter in grain sample B was observed. This was also the case for Acetobacter orientalis in sample D. In relation to 26S pyrosequencing, our study revealed the presence of 3 main yeast species: Naumovozyma spp., Kluyveromyces marxianus, and Kazachastania khefir. For Naumovozyma, further studies are needed to assess the isolation of new species. In conclusion, this study has proved that it is possible to establish the patterns of bacterial and yeast composition of kefir and kefir grain. This was only achieved with the use of high-throughput sequencing techniques. Key words: kefir, microbiota, metagenetics, 16S rRNA sequencing, 26S rRNA sequencing Short CommunicationOriginally from the Caucasus Mountains (Otles and Cagindi, 2003), kefir is widely consumed in Eastern Europe but now encountered all over the world. The word "kefir" means "good feeling" in Turkish, due to the cooling nature of this beverage. The kefir beverage is produced by mixing milk, water, or fruit juice with kefir grains, which have the appearance of small cauliflowers (Lopitz-Otsoa et al., 2006). These grains are a symbiotic combination of bacteria (mainly lactob...

From 1997 to 1999, the prevalence of Salmonella was assessed at different stages through the pork, poultry, and beef meat production chains. Different dilutions of the initial sample suspension were analyzed to provide a semiquantitative evaluation of Salmonella contamination and to determine the most representative dilution necessary to detect a reduction in prevalence. An average of 300 samples for each type of meat were analyzed. According to Fisher's exact test, the dilution to be used to detect a reduction in prevalence was chosen based on an initial prevalence of 20 to 26%. Based on this introductory study, a new sampling plan representative of the nationwide Belgian meat production process was used from 2000 through to 2003. This study confirmed the consistently high rate and level of contamination of poultry meat: broiler and layer carcasses were the most contaminated samples followed by broiler fillets and poultry meat preparations. A constant and significant decrease in Salmonella prevalence was observed for pork carcasses, trimmings, and minced meat and for beef minced meat. Less than 3% of beef carcasses and trimming samples were positive for Salmonella. The Belgian plan, as utilized from 2000 to 2003, was suitable for monitoring of zoonoses because the sampling plan was representative of nationwide production processes, covered all periods of the year, and was executed by trained samplers and the analyses were carried out by recognized laboratories using an identical analytical method.

A survey was conducted to collect data on Salmonella prevalence, Escherichia coli counts (ECCs), and aerobic bacteria colony counts (ACCs) on pig carcasses after chilling at the 10 largest Belgian pig slaughterhouses during 2000 through 2004. Potential risk factors of contamination associated with production parameters, technical descriptions of the installations, and cleaning and disinfection methods were assessed during investigations in the slaughterhouses. These variables were used first in a univariate analysis and then were extended to a multivariate analysis with a logistic mixed regression model for Salmonella and a linear mixed model for ECCs and ACCs with slaughterhouses as the random effect. The results indicated high variability concerning Salmonella contamination among the 10 slaughterhouses, with prevalence ranging from 2.6 to 34.3% according to the area of origin. The median ECC and median ACC ranged from -0.43 to 1.11 log CFU/cm2 and from 2.37 to 3.65 log CFU/cm2, respectively. The results of the logistic and linear regressions revealed that some working practices such as scalding with steam, second flaming after polishing, and complete cleaning and disinfection of the splitting machine several times a day were beneficial for reducing Salmonella prevalence, ECCs, and ACCs. Changing the carcass hooks just before chilling, using water as the cleaning method, and a higher frequency of disinfection of the lairage seemed to be protective against E. coli in the multivariate mixed linear model. The monitoring of critical points, slaughterhouse equipment, good slaughtering practices, and effective washing and disinfection are the keys to obtaining good microbiological results.

The method presented in this paper should prove useful in assessing the effectiveness of HACCP plans developed in slaughterhouses. Samples were collected by swabbing well-defined areas of pork and beef carcasses with sterile gauze. Between 160 and 420 half-carcasses were swabbed in each of nine pork or beef slaughterhouses. Swabs from five carcasses were placed in the same sterile Stomacher bag, constituting a single composite sample. Standard or validated analytical methods were used to isolate and characterize four foodbome pathogens. Salmonella spp., Listeria monocytogenes, Campylobacter spp., and verocytotoxin-producing E. coli were detected, respectively, in 27, 2, 2, and 14% of the pork samples and 0, 22. 10, and 5% of the beef samples. Of the 10 samples positive for E. coli O157, only one yielded an isolate confirmed to be entcrohemorrhagic. Since Salmonella spp. appear as the main contaminant of pork (27%) and L. monocytogenes as the main contaminant of beef (22%), any slaughterhouse sampling plan should include testing for the former in the case of pork carcasses and for the latter in the case of beef carcasses. One should also test regularly for the presence of E. coli O157 and Campylobacter spp. in pork and beef abattoirs. The method presented here is an easy way to assess the contamination rate of carcasses at the end of the slaughtering process.