Slanetz bartley agar

Subsamples for enumeration of faecal coliforms (FC), intestinal enterococci (IE) and Vibrio spp. were filtered onto sterile cellulose nitrate membranes (0.22 μm pore size, 47 mm diameter, GE Healthcare Life Sciences, Little Chalfont, UK), through a hand-pump, placed in Chromogenic Coliform Agar (Biokar diagnostics, Fr), Slanetz-Bartley Agar (Oxoid, Waltham, MA, USA) and (Vibrio Chrome agar medium, Fr) plates, respectively, and incubated at 44.5 °C for 24 h (FC) or 48 h (IE), or 37 °C for 24 h (Vibrio spp.). Aerobic mesophilic microorganisms (AMM) were determined according to ISO 4833: 2003, using the incorporation technique, by pipetting 1 mL of sample onto each plate with the addition of mFc-agar Yeast Extract Agar (Biokar diagnostics, Fr), and incubation at 37 °C for 24 h. For FC, E. coli and IE, a volume of 100 mL was used, while for Vibrio spp., it was 300 mL. Typical colonies were counted, and the result was expressed as colony forming units (CFU/100 mL).

For the microbiological analysis, serial dilutions in sodium citrate (2% w/v) were prepared starting from 10 g of each cheese The following microorganisms were investigated: aerobic mesophilic bacteria (AMB) on Plate Count Agar (PCA; Oxoid, Milan, Italy) at 30 °C for 2 days; mesophilic lactobacilli on MRS agar (Oxoid, Basingstoke, UK), acidified to pH 5.4 with acetic acid, at 30 °C for 2 days in anaerobic conditions using the Gas-Pack anaerobic system (AnaeroGen; Oxoid, Basingstoke, UK); lactococci on M17 (Oxoid, Basingstoke, UK), containing 1% (w/v) lactose (Fluka Chimica, Milan, Italy), at 30 °C for 2 days in anaerobic conditions; enterococci on Slanetz-Bartley agar (Oxoid, Basingstoke, UK) at 37 °C for 48 h; Enterobacteriaceae on Violet Red Bile Glucose Agar (VRBGA; Oxoid, Basingstoke, UK) at 37 °C for 24 h. Cell counts were performed in duplicate.

Feces (10 g) were mixed with 90 mL of a tryptone salt broth (Oxoid). Suspensions were transferred to sterile stomacher bags and homogenized for 2 min in a stomacher blender. Serial dilutions were made in tryptone salt broth. Dilutions were inoculated in Slanetz-Bartley Agar (Oxoid) (37 °C, 24–48 h). Up to four red-colored colonies were selected to collect a variety of enterococcus strains. For further identification, catalase, growth in Bile Esculin Agar (Oxoid), and trypticase soy broth + 6.5% NaCl tests were done. If there was any doubt about the identification, the API STREP (BioMérieux, France) was used.