Antifungal activity of lactic acid bacteria isolated from milk against Penicillium commune, P. nordicum, and P. verrucosum.

Penicillium spp. is considered a major spoilage fungus of cheeses. The use of lactic acid bacteria (LAB) with antifungal activity is an interesting possibility of biopreservation. In this study, the isolation and characterization of anti-Penicillium LAB from milk was carried out. Ninety-three milk samples were analysed and a total of 57 strains of LAB active against P. nordicum were isolated, mainly from goat and cow milk. Thirty-four isolates with strong activity were selected and identified, Lacticaseibacillus casei (11), L. paracasei (9) and L. rhamnosus (5) being the dominant species. The antifungal spectrum of these 34 LAB against strains of P. commune and P. verrucosum was investigated. L. casei, L. paracasei and L. rhamnosus were the most active and P. nordicum was the most susceptible fungus. Two isolates (L. casei Lc-51/3 and L. paracasei Lp-25/1) with high antifungal activity showed a moderate to high reduction on the growth of Penicillium nordicum and, in a lesser extent, of P. commune, and also a reducing effect on the ochratoxin A and cyclopiazonic acid production. In addition, these isolates demonstrated activity against several food pahogens. These findings indicate their suitability for the development of protective adjunct starters against spoilage and toxigenic microorganisms in cheese processing.

Behaviour of Non-O157 STEC and Atypical EPEC during the Manufacturing and Ripening of Raw Milk Cheese.

This study was carried out to assess the survival of Shiga toxin-producing E. coli (STEC) and atypical enteropathogenic Escherichia coli (aEPEC) during the traditional manufacturing and ripening of Spanish hard cheese from raw cow's milk. Milk samples were spiked with up to 3.1-3.5 log cfu/mL of one strain of STEC (O140:H32 serotype) and one of aEPEC (serotype O25:H2). The first steps of cheesemaking allow for a STEC and aEPEC increase of more than 1 log cfu/mL (up to 4.74 log cfu/g and 4.55 log cfu/g, respectively). After cheese pressing, a steady reduction of both populations was observed, with the STEC strain being more sensitive. The studied pathogenic E. coli populations decreased by 1.32 log cfu/g in STEC and 0.59 log cfu/g in aEPEC in cheese ripened during a minimum period of 60 d. Therefore, a moderate contamination by these diarrhoeagenic E. coli pathotypes, in particular, with aEPEC, on cheese manufactured from raw milk may not be totally controlled through the cheesemaking process and during a maturation of 90 d. These findings remark the importance of improvement in bacteriological quality of raw milk and cross-contamination prevention with diarrhoeagenic E. coli in the dairy industry.

X-ray diffraction has limited applicability in investigation of milk tampering.

The aim of this work was to use X-ray diffraction to identify substances used for adulteration of raw milk and to determine if crystallographic analysis can detect extraneous substances in milk. Two unknown substances were sent anonymously by employers linked to the dairy chain, who claimed that they were added directly in milk prior to water addition by truck drivers. The samples were analyzed by X-ray diffraction and submitted to physicochemical analysis. The first substance was identified by X-ray diffraction as sodium citrate, complying with its physicochemical attributes, such as the powerful ability to decrease the freezing point. The second substance was identified by X-ray diffraction as sucrose and this result was also in agreement with its ability to increase the density, decrease the freezing point and finally, to be positive for sucrose in the resorcinol qualitative test. To evaluate if X-ray diffraction can detect extraneous substances already mixed in milk, fresh raw milk samples tampered with urea, sodium hydroxide, sodium citrate and sucrose were freeze dried and analyzed by X-ray diffraction, with no detection of any extraneous substances at any percentage. This is the first report of attempted diagnosis of extraneous substances in milk by X-ray diffraction. However, this technique can be useful only when applied to identify substances used for adulteration prior to its dilution in milk, since the amorphous nature of milk seems to be a limitation for the accurate detection of extraneous substances.

Polyphasic identification of Penicillium spp. isolated from Spanish semi-hard ripened cheeses.

Fifteen samples of semi-hard ripened cheeses, both spoiled (10) and unspoiled (5), and obtained from cheese factories located in Northwest of Spain, were analysed by a dilution plating technique and direct sampling. A total of 32 isolates were identified at species level by a polyphasic approach (phenotypic characterization, partial extrolite analysis and molecular identification). Most isolates (65.6%) belonged to the species P. commune; other species found were P. solitum, P. chrysogenum, P. nordicum, P. expansum and P. cvjetkovicii. All of the P. commune isolates were able to produce cyclopiazonic acid, while the P. nordicum and the P. expansum isolates were producers of ochratoxin A and patulin respectively. Despite this, the role of P. commune as beneficial fungi in cheese ripening should be investigated. Molecular identification based on BenA sequence analysis was able to identify the majority of isolates. The three mycotoxins investigated can be considered key for identification. The polyphasic approach seems to be a very valuable tool for identification of isolates of this complex genus.