Gianluigi Mauriello

Isolation and technological properties of coagulase negative staphylococci from fermented sausages of Southern Italy

The aims of this study were to characterize the population of Micrococcaceae in different types of fermented sausages of Southern Italy and to determine the technological properties of Staphylococcus strains in order to evaluate the suitability of selected strains as starter cultures in the processing of dry fermented pork sausages. Ninety-six strains were studied to evaluate nitrate reductase, proteolytic, lipolytic and antioxidant activities as well as growth ability at different temperatures, pHs and NaCl concentrations. All the strains were classified as Staphylococcus except for one isolate assigned to Kocuria spp. The species most often isolated were S. saprophyticus, S. xylosus and S. equorum, although they were not equally distributed within the different sausages. Other species isolated were, in descending order of abundance, S. succinus, S. warneri, S. lentus, S. vitulus, S. pasteuri, S. epidermidis, and S. haemolyticus. In general, the S. xylosus strains exhibited the best technological properties that would make them eligible as good starter cultures for fermented meat products. However, strains belonging to other species also showed good technological properties. Finally, all strains grew at 10, 15 and 20 °C, in the presence of 10% and 15% of NaCl and at pH 5.0 and 5.5. The results showed that it is possible to formulate a broad variety of staphylococcal starter cultures, adaptable to different technological conditions and sausage manufacture practices.

Selection of Lactobacillus strains from fermented sausages for their potential use as probiotics.

A rapid screening method was used to isolate potentially probiotic Lactobacillus strains from fermented sausages after enrichment in MRS broth at pH 2.5 followed by bile salt stressing (1% bile salts w/v). One hundred and fifty acid- and bile-resistant strains were selected, avoiding preliminary and time-consuming isolation steps. Strains were further characterized for survival at pH 2.5 for 3 h in phosphate-buffered saline and for growth in the presence of 0.3% bile salts with and without pre-exposure at low pH. Twenty-eight strains showed a survival >80% at pH 2.5 for 3 h; moreover, most of the strains were able to grow in the presence of 0.3% bile salts. Low pH and bile resistance was shown to be dependent on both the species, identified by phenotypic and molecular methods, and the strain tested. This is the first report on the direct selection of potentially probiotic lactobacilli from dry fermented sausages. Technologically interesting strains may be used in the future as probiotic starter cultures for novel fermented sausage manufacture.

Monitoring of Microbial Metabolites and Bacterial Diversity in Beef Stored under Different Packaging Conditions

ABSTRACT Beef chops were stored at 4°C under different conditions: in air (A), modified-atmosphere packaging (MAP), vacuum packaging (V), or bacteriocin-activated antimicrobial packaging (AV). After 0 to 45 days of storage, analyses were performed to determine loads of spoilage microorganisms, microbial metabolites (by solid-phase microextraction [SPME]-gas chromatography [GC]-mass spectrometry [MS] and proton nuclear magnetic resonance [1H NMR]), and microbial diversity (by PCR–denaturing gradient gel electrophoresis [DGGE] and pyrosequencing). The microbiological shelf life of meat increased with increasing selectivity of storage conditions. Culture-independent analysis by pyrosequencing of DNA extracted directly from meat showed that Brochothrix thermosphacta dominated during the early stages of storage in A and MAP, while Pseudomonas spp. took over during further storage in A. Many different bacteria, several of which are usually associated with soil rather than meat, were identified in V and AV; however, lactic acid bacteria (LAB) dominated during the late phases of storage, and Carnobacterium divergens was the most frequent microorganism in AV. Among the volatile metabolites, butanoic acid was associated with the growth of LAB under V and AV storage conditions, while acetoin was related to the other spoilage microbial groups and storage conditions. 1H NMR analysis showed that storage in air was associated with decreases in lactate, glycogen, IMP, and ADP levels and with selective increases in levels of 3-methylindole, betaine, creatine, and other amino acids. The meat microbiota is significantly affected by storage conditions, and its changes during storage determine complex shifts in the metabolites produced, with a potential impact on meat quality.

Antimicrobial activity of a nisin-activated plastic film for food packaging

Aims:  To determine the effectiveness of a packaging film coated with nisin to inhibit Micrococcus luteus ATCC 10240 in tryptone soya broth (TSB) and the microbiota of raw milk during storage. A further aim was to examine the release of nisin from the activated film.

Microbial succession during ripening of Naples-type salami, a southern Italian fermented sausage

Studies were carried out on the microbiological and physico-chemical changes which occurred during the ripening of five batches of Naples-type salami, manufactured without starter cultures. Salami were sampled internally and externally, and the following microbial groups were studied: lactic acid bacteria, Micrococcaceae and yeasts. The results obtained indicated that lactobacilli constituted the predominant flora, both on the surface and in the interior of the pieces throughout the ripening period. Micrococcaceae and yeasts were also found in considerable number in both locations. Characterisation of 191 lactic isolates indicated that the salami microflora was dominated by homofermentative lactobacilli; approximately 63% of them could be identified as Lactobacillus sake; 40% showing the traits of a racemase negative variant of this species, once referred to Lactobacillus bavaricus. Yeast population mainly comprised Debaryomyces strains. All the colonies grown on mannitol salt and Kranep agar were catalase-positive cocci; novobiocin-resistant staphylococci were the only Micrococcaceae found. The API Staph identification system did not prove to be reliable: 82% of the isolates remained unidentified. To achieve improved characterisation, cluster analysis was subsequently performed on this group, corroborating the existence of a fairly homogeneous group representing an intermediate variety between Staphylococcus xylosus and Staphylococcus saprophyticus that was isolated during the whole ripening process.

Development of polythene films for food packaging activated with an antilisterial bacteriocin from Lactobacillus curvatus 32Y

Aims:  The aims of this work were to (i) use a bacteriocin produced by Lactobacillus curvatus 32Y active against Listeria monocytogenes to activate polythene films by different methods, (ii) implement a large‐scale process for antilisterial polythene films production and (iii) verify the efficacy of the developed films in inhibiting the growth of L. monocytogenes during the storage of meat products.

PCR-DGGE fingerprints of microbial succession during a manufacture of traditional water buffalo mozzarella cheese

Aims:  To monitor the process and the starter effectiveness recording a series of fingerprints of the microbial diversity occurring at different steps of mozzarella cheese manufacture and to investigate the involvement of the natural starter to the achievement of the final product.

Development of spoilage microbiota in beef stored in nisin activated packaging.

The aim of this study was to assess the microbial populations causing the spoilage of chilled beef during storage and to evaluate the effect of the use of an antimicrobial packaging for the meat storage. A nisin activated antimicrobial packaging was developed by using a nisin, HCL and EDTA solution and used for the storage of beef cuts at 1 degrees C. The common spoilage related microbial groups were monitored during the storage of beef in activated and non activated plastic bags by using selective media. The use of the antimicrobial packaging caused an overall significant reduction of viable counts of Gram positive bacteria such as carnobacteria, lactic acid bacteria and Brochotrix thermosphacta whose development was inhibited for at least 11 days of storage compared to the control. Moreover, a 1-3 log cycles reduction of enterobacteria was also registered between 22 and 32 days of storage. The microbiota was assessed at species level by using Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis of 16S rRNA gene of DNA extracted directly from meat and from bulk cells from selective media plates and showed that the species occurring within the targeted microbial groups did not change according to storage conditions. In conclusion, the use of the nisin activated packaging reduced the number of spoilage populations but did not affect the species diversity. Improved antimicrobial packaging is needed, possibly coupled with vacuum storage, to possibly achieve a simultaneous inhibition of more spoilage microbial groups and to preserve the microbiological quality of beef during chilled storage.

Detection and characterization of a bacteriocin, garviecin L1-5, produced by Lactococcus garvieae isolated from raw cow's milk

Aims: The identification of a bacteriocin‐producing lactococcal strain isolated from raw cow’s milk is reported, along with production conditions, physical and chemical properties, and mode of action of the bacteriocin.

Simultaneous detection of Pseudomonas fragi, P. lundensis, and P. putida from meat by use of a multiplex PCR assay targeting the carA gene.

ABSTRACT Species-specific primers and a multiplex PCR assay were developed for the simultaneous identification and differentiation of Pseudomonas fragi, P. lundensis, and P. putida based on the coamplification of different portions of the small subunit of the carbamoyl phosphate synthase gene (carA). The carA multiplex PCR was used to detect the presence of the three Pseudomonas species from beef, chicken, and pork samples and proved to be effective in showing their evolution during the storage of meat.