Glycerol production by Oenococcus oeni during sequential and simultaneous cultures with wine yeast strains.

Growth and fermentation patterns of Saccharomyces cerevisiae, Kloeckera apiculata, and Oenococcus oeni strains cultured in grape juice medium were studied. In pure, sequential and simultaneous cultures, the strains reached the stationary growth phase between 2 and 3 days. Pure and mixed K. apiculata and S. cerevisiae cultures used mainly glucose, producing ethanol, organic acids, and 4.0 and 0.1 mM glycerol, respectively. In sequential cultures, O. oeni achieved about 1 log unit at 3 days using mainly fructose and L-malic acid. Highest sugars consumption was detected in K. apiculata supernatants, lactic acid being the major end-product. 8.0 mM glycerol was found in 6-day culture supernatants. In simultaneous cultures, total sugars and L-malic acid were used at 3 days and 98% of ethanol and glycerol were detected. This study represents the first report of the population dynamics and metabolic behavior of yeasts and O. oeni in sequential and simultaneous cultures and contributes to the selection of indigenous strains to design starter cultures for winemaking, also considering the inclusion of K. apiculata. The sequential inoculation of yeasts and O. oeni would enhance glycerol production, which confers desirable organoleptic characteristics to wines, while organic acids levels would not affect their sensory profile.

Characterization of a bacteriocin produced by Lactococcus lactis subsp. lactis CRL 1584 isolated from a Lithobates catesbeianus hatchery.

Lactococcus lactis CRL 1584 isolated from a Lithobates catesbeianus hatchery inhibits the growth of Citrobacter freundii (a bullfrog pathogen) and Listeria monocytogenes by a synergistic effect between lactic acid, hydrogen peroxide and a bacteriocin-like molecule. The chemical characterization of the bacteriocin in cell-free supernatants indicates that it has a proteinaceous nature. Hexadecane and ethyl acetate did not modify the bacteriocin activity, while 10 and 20 % (v/v) chloroform decreased the activity by 29 and 43 %, respectively. The antimicrobial peptide was heat stable since 85 % of residual activity was detected when neutralized supernatants were heated at 80 °C for 30 min. Moreover, no bacteriocin inactivation was observed when supernatants were kept at -20 °C for 3 months. The synthesis of the bacteriocin was associated with bacterial growth, highest production (2,100 AU/ml) being detected at the end of the exponential growth phase. At pH ranges of 5-6.5 and 5.0-5.5 the inhibitory molecule was stable when stored for 2 days at 4 and 25 °C, respectively. Moreover, it had a bactericidal effect on L. monocytogenes and the ultrastructural studies of pathogenic cells revealed clumping of the cytoplasmic material, increased periplasmic space and cell wall modifications. The deduced amino acid sequence of the bacteriocin was identical to nisin Z and the genetic determinants for its production are harbored in the chromosome. These results, described for the first time in L. lactis from a bullfrog hatchery, will increase knowledge of the bacteriocin under study with a view to its potential inclusion in probiotics for raniculture or biopreservatives.

Cultivable microbiota of Lithobates catesbeianus and advances in the selection of lactic acid bacteria as biological control agents in raniculture.

The cultivable microbiota of skin and cloaca of captive Lithobates catesbeianus includes microorganisms generally accepted as beneficial and potentially pathogenic bacteria. In order to select a group of potentially probiotic bacteria, 136 isolates were evaluated for their surface properties and production of antagonistic metabolites. Then, 11 lactic acid bacteria (LAB) strains were selected and identified as Lactobacillus plantarum, Lb. brevis, Pediococcus pentosaceus, Lactococcus lactis, L. garvieae and Enterococcus gallinarum. Studies of compatibility indicate that all the strains could be included in a multi-strain probiotic, with the exception of Ent. gallinarum CRL 1826 which inhibited LAB species through a bacteriocin-like metabolite. These results contribute to the design of a probiotic product to improve the sanitary status of bullfrogs in intensive culture systems, to avoid the use of antibiotics and thus to reduce production costs. It could also be an alternative to prevent infectious diseases during the ex situ breeding of amphibian species under threat of extinction.