Cecilia Alejandra Fontana

Direct molecular approach to monitoring bacterial colonization on vacuum-packaged beef.

ABSTRACT Denaturing gradient gel electrophoresis allowed us to monitor total bacterial communities and to establish a pattern of succession between species in vacuum-packaged beef stored at 2 and 8°C for 9 weeks and 14 days. Species-specific PCR was used to confirm the presence of Lactobacillus sakei and Lactobacillus curvatus. Multiplex PCRs using 16S rRNA-specific primers allowed differentiation between Leuconostoc species. These methods provided the desired information about microbial diversity by detecting the main microorganisms capable of colonizing this ecological niche.

Surface microbiota analysis of Taleggio, Gorgonzola, Casera, Scimudin and Formaggio di Fossa Italian cheeses.

The composition of the bacterial consortia of the smear Italian cheeses and their role on quality and safety is still poorly understood. The objective of this study was to identify and characterize the bacterial communities present on the surface of five traditional Italian cheeses, Casera Valtellina, Scimudin, Formaggio di Fossa, Gorgonzola and Taleggio. DGGE analysis performed using total DNA obtained from cheese surfaces enabled us to identify the dominant bacterial populations. Bands showing different intensity and identified as Staphylococcus, Micrococcus, Psychrobacter, Enterococcus and Brevibacterium species were detected on the surface of cheeses. The cluster analysis showed that Gorgonzola, Taleggio and Formaggio di Fossa cheeses present high similarity in their surface bacterial composition while major differences in the DGGE profiles were observed in Scimudin and Casera. The molecular taxonomical identification among the Gram positive isolates, reveals the presence of the following bacterial genera: Staphylococcus, Micrococcus, Macrococcus, Enterococcus, Lactobacillus, Carnobacterium, Leuconostoc, Brevibacterium, Corynebacterium, Brochothrix, Bacillus. The combination of culture dependent and independent techniques allowed us to obtain information about the bacterial species covering the surface of five different traditional Italian cheeses.

Biodiversity and technological-functional potential of lactic acid bacteria isolated from spontaneously fermented quinoa sourdoughs

To analyse lactic acid bacteria (LAB) diversity and technological‐functional and safety properties of strains present during spontaneous fermented quinoa sourdoughs.

Draft Genome Sequence of Clostridium tyrobutyricum Strain UC7086, Isolated from Grana Padano Cheese with Late-Blowing Defect

ABSTRACT Clostridium tyrobutyricum is considered the main agent of late-blowing defect in the production of hard cheese. Here, we described the draft genome sequences and annotation of C. tyrobutyricum strain UC7086, which was isolated from Grana Padano cheese with blowing defect, and C. tyrobutyricum DSM 2637 type strain in a comparative study.

Population structure and safety aspects of Enterococcus strains isolated from artisanal dry fermented sausages produced in Argentina

Enterococci population from Argentinean artisanal dry fermented sausage was identified and their safety aspects were evaluated. Species‐specific PCR was used to distinguish between Enterococcus faecium (56%) and Enterococcus faecalis (17%). Other isolates (27%) were identified as Enterococcus durans, Enterococcus casseliflavus and Enterococcus mundtii by using 16S RNA gene sequence. RAPD analyses showed different biotypes for Ent. faecium and Ent. faecalis species. Low incidence of antibiotic resistance and high virulence traits in Ent. casseliflavus and Ent. faecalis were found; the majority of the Ent. faecium strains were shown to be free of virulence factors. The absence of virulence/resistance traits and the anti‐Listeria activity of Ent. faecium isolates may be exploited to enhance natural preservation thereby guaranteeing organoleptic/safety characteristics of artisanal fermented sausages.

Characteristics and Applications of Microbial Starters in Meat Fermentations

Fermentation and drying are among the most ancient food preservation techniques used by man. Developed through the years, these processes prolonged the storage time of meats (and meat products) and brought favorable changes to their organoleptic properties, with respect to the original substrate. The microbiology of fermented sausages is varied and complex. The type of microflora that develops is often closely related to the ripening technique utilized. Several investigations established two groups of microorganisms as being the main organisms responsible for the transformations involved during fermentation and ripening of sausages. The spontaneous fermentation of dry sausages involves the participation of the Lactic Acid Bacteria (LAB), Gram-positive Catalase-positive Cocci (GCC+, mostly Staphylococcus and Kocuria species), and, less importantly, yeasts and moulds. Most of the commercially available meat starter cultures contain mixtures of LAB and GCC+. These bacterial groups are responsible for the basic microbial reactions that occur simultaneously during fermentation; the decrease of pH values via glycolysis by LAB, the reduction of nitrate, and the development of aroma by GCC+. Research on the use of starter cultures in meat products began in the USA in the 1940s, inoculating the raw material for fermented sausages using lactobacilli, with the aim to govern and accelerate the fermentation. In the late fifties, the Niinivaara (1955) helped to launch this idea in Europe developing mixed cultures of the Micrococcus sp. and Pediococcus cerevisiae. According to the definitions of Hammes, Bantleon, and Min (1990), meat starter cultures are preparations that contain active or dormant microorganisms that develop the desired metabolic activity in the meat. They are, by definition, used to drive the fermentation process in the desired direction, reducing the variability in the quality of the product, limiting the growth of spoilage bacteria by accelerating

Strategies for Pathogen Biocontrol Using Lactic Acid Bacteria and Their Metabolites: A Focus on Meat Ecosystems and Industrial Environments

The globalization of trade and lifestyle ensure that the factors responsible for the emergence of diseases are more present than ever. Despite biotechnology advancements, meat-based foods are still under scrutiny because of the presence of pathogens, which causes a loss of consumer confidence and consequently a fall in demand. In this context, Lactic Acid Bacteria (LAB) as GRAS organisms offer an alternative for developing pathogen-free foods, particularly avoiding Listeria monocytogenes, with minimal processing and fewer additives while maintaining the foods’ sensorial characteristics. The use of LAB strains, enabling us to produce antimicrobial peptides (bacteriocins) in addition to lactic acid, with an impact on quality and safety during fermentation, processing, and/or storage of meat and ready-to-eat (RTE) meat products, constitutes a promising tool. A number of bacteriocin-based strategies including the use of bioprotective cultures, purified and/or semi-purified bacteriocins as well as their inclusion in varied packaging materials under different storage conditions, have been investigated. The application of bacteriocins as part of hurdle technology using non-thermal technologies was explored for the preservation of RTE meat products. Likewise, considering that food contamination with L. monocytogenes is a consequence of the post-processing manipulation of RTE foods, the role of bacteriocinogenic LAB in the control of biofilms formed on industrial surfaces is also discussed.

Microbial ecology involved in the ripening of naturally fermented llama meat sausages. A focus on lactobacilli diversity

Llama represents for the Andean regions a valid alternative to bovine and pork meat and thanks to the high proteins and low fat content; it can constitute a good product for the novel food market. In this study, culture-dependent and independent methods were applied to investigate the microbial ecology of naturally fermented llama sausages produced in Northwest Argentina. Two different production technologies of llama sausage were investigated: a pilot-plant scale (P) and an artisanal one (A). Results obtained by High-Throughput Sequencing (HTS) of 16S rRNA amplicons showed that the production technologies influenced the development of microbial communities with a different composition throughout the entire fermentation process. Both sequencing and microbiological counts demonstrated that Lactic Acid Bacteria (LAB) contributed largely to the dominant microbiota. When a total of 230 isolates were approached by RAPD-PCR, presumptive LAB strains from P production exhibited an initial variability in RAPD fingerprints switching to a single profile at the final of ripening, while A production revealed a more heterogeneous RAPD pattern during the whole fermentation process. The constant presence of Lactobacillus sakei along the fermentation in both productions was revealed by HTS and confirmed by species-specific PCR from isolated strains. The technological characterization of Lb. sakei isolates evidenced their ability to grow at 15°C, pH4.5 and 5% NaCl (95%). Most strains hydrolyzed myofibrillar and sarcoplasmic proteins. Bacteriocins encoding genes and antimicrobial resistance were found in 35% and 42.5% of the strains, respectively. An appropriate choice of a combination of autochthonous strains in a starter formulation is fundamental to improve and standardize llama sausages safety and quality.

Comparative proteomic analyses for elucidating metabolic changes during EPS production under different fermentation temperatures by Lactobacillus plantarum Q823.

Exopolysaccharide (EPS)-producing bacteria are of growing interest in industrial processes, mainly concerning food. Lactic acid bacteria are widely appreciated for their GRAS (generally recognized as safe) status and their ascertained or putative probiotic features. Detailed investigation on what happens at metabolic level during EPS production is scarce in the literature. The facultative heterofermenter Lactobacillus plantarum Q823 was studied in order to compare growth and EPS production at 30°C and 37°C. A higher growth rate was observed at 37°C, whereas, a significantly higher (tenfold increase) EPS amount was produced at 30°C. To understand the molecular mechanisms leading to the different EPS production in the two conditions, a comparative proteomic experiment was performed. The results of the in-gel proteomics revealed that: i) at 37°C a higher abundance of proteins involved in carbon catabolism and nucleic acid biosynthesis together with a significant amount of stress proteins was observed; ii) at 30°C the production of an atypical manganese-containing non-heme catalase (pseudocatalase) was increased, in agreement with previous data reporting that growth-rates of catalase negative Lactobacillus plantarum strains were greater than that of catalase positive strains. Taken together, all these findings provide further insights about the metabolic pathways stimulated during EPS production, and the mechanism that triggers EPS biosynthesis.

Biodiversity and technological potential of lactic acid bacteria isolated from spontaneously fermented amaranth sourdough.

Spontaneous fermented sourdoughs prepared from amaranth flour were investigated for the presence of autochthonous lactic acid bacteria (LAB) predominating microbiota. The doughs were fermented with daily backslopping on a laboratory scale at 30°C for 10 days. LAB counts ranged from 2·60 to 8·54 log CFU g−1 with a pH declined from 6·2 to 3·8 throughout fermentation. The combined use of randomly amplified polymorphic DNA (RAPD)‐PCR analysis and sequence analysis of 16S rRNA was applied for LAB intraspecies differentiation and taxonomic identification, respectively. Enterococcus, Pediococcus and Lactobacillus species were present in amaranth sourdoughs (AS). After the first refreshment step, Lactobacillus plantarum dominated AS until the end of fermentation. In coincidence, when DGGE analysis was performed, the occurrence of a progressive change in bacterial communities allowed the selection of Lact. plantarum as a dominant species. Moreover, technological, functional and safety characteristics of representative RAPD‐biotypes were investigated. Lact. plantarum CRL1898 was selected as a potential candidate for gluten‐free amaranth sourdough starter.