Modelling Biolog profiles' evolution for yeast growth monitoring in alcoholic fermentation.

AIMS: A research was undertaken to explore the possibility to express with suitable mathematical models Biolog metabolic curves obtained for oenological yeasts and to use such models for monitoring yeast growth in alcoholic fermentation. METHODS AND RESULTS: Experimental curves of metabolic activity in Biolog YT microplates, obtained in a previous work for various oenological yeast strains in pure cultures and mixed populations, at various cell concentrations, have been modelled with Gompertz's, Gompertz's modified and Lindstrom's mathematical equations. Lindstrom's model proved to be the most suitable to fit the curves of the oenological yeasts under study, providing the highest correlation coefficients between experimental and calculated data. The model made it possible to recognize, in mixed yeast populations, the presence of active dry yeasts used for guided fermentations. Model's constant parameters were used for a numerical characterization of yeast curves. CONCLUSIONS: The application of the model to the experimental data resulted to be suitable for an early prediction of the successive evolution of yeast growth. SIGNIFICANCE AND IMPACT OF THE STUDY: The results obtained indicate the possibility to develop protocols for monitoring yeast presence during alcoholic fermentation, with an early assessment of the correct evolution of their growth, especially when active dry yeasts are employed.

Possible use of Biolog methodology for monitoring yeast presence in alcoholic fermentation for wine-making.

AIMS: A research was undertaken to explore the possibility to use Biolog system of microbial metabolic characterization for the monitoring of yeast population evolution during alcoholic fermentation for wine production. METHODS AND RESULTS: An application of Biolog system was employed for the characterization of yeasts of oenological interest, in pure cultures and mixed consortia, in various cell concentrations. The system's capacity to discriminate among different cell concentrations of the same yeast strain was ascertained, along with the capacity to discriminate between mixed and pure populations. CONCLUSIONS: The tested application of Biolog system resulted suitable for a quick recognition (24 h) of the presence of starter cultures within mixed populations of autochthonous yeasts. Such discrimination was confirmed with the one resulting from molecular techniques. SIGNIFICANCE AND IMPACT OF THE STUDY: The study suggests the possibility to employ Biolog system for an early monitoring of yeast evolution in modern wine-making fermentations, where specialized yeasts are more and more frequently used as starters and their ability to overcome autochthonous yeast populations is crucial.

Maturing dynamics of surface microflora in Fontina PDO cheese studied by culture-dependent and -independent methods.

AIMS: To study the evolution of rind microbial communities in Fontina PDO cheese. METHODS AND RESULTS: Four batches were examined for their surface microflora during ripening, carried out in two different maturing caves, at Ollomont and Pré-Saint-Didier, Aosta Valley region, Northwest of Italy. Culture-dependent methodologies were combined with culture-independent analysis (PCR-DGGE). Yeasts were found to increase from 10(3) to 10(6) CFU cm(-2) in 28 days, with consequent rise of surface pH, which allowed the growth of salt-tolerant bacteria, in particular coryneforms which reached 10(9) CFU cm(-2) at the end of 3 months. Coagulase-negative cocci and lactic acid bacteria reached 10(7) CFU cm(-2) in the same period. Debaryomyces hansenii and Candida sake were the species more constantly present throughout the whole maturing process. As early as after 1 day since manufacture, Lactococcus lactis subsp. lactis and Streptococcus thermophilus were detected on cheese rinds. Arthrobacter nicotianae, Brevibacterium casei and Corynebacterium glutamicum were found after 7-28 days. CONCLUSIONS: According to cluster analysis of DGGE profiles, the maturing environment seemed to influence the dynamics of microbial groups on Fontina surfaces. SIGNIFICANCE AND IMPACT OF THE STUDY: These results represent a first picture of micro-organisms colonizing Fontina PDO rinds. Further studies are in progress to better understand the origin of this surface microflora and to formulate surface starters.

Effects of substratum on the diversity and stability of ammonia-oxidizing communities in a constructed wetland used for wastewater treatment.

AIM: To study the relationship between the nature of the substratum and the diversity and stability of the ammonia-oxidizing microbial community in a constructed wetland for the treatment of wastewaters. METHODS AND RESULTS: Samples have been taken the year around from sections of the wetland filled with different substrata. When present, the root zones of the helophyte Phragmites australis were also sampled. The diversity of the ammonia-oxidizing community was established by a coupled PCR-DGGE method based on the 16s rRNA gene. Averaged over the seasons, no large differences in community composition were observed between the different substrata, although the section with zeolite always showed the highest frequencies of bands belonging to ammonia-oxidizing bacteria of the beta-subclass of the Proteobacteria. Only sequences related to the Nitrosospira lineage were detected. Averaged again over the seasons, the section with zeolite was also most constant with respect to the potential ammonia-oxidizing activity. CONCLUSIONS: Although the ammonia-oxidizing communities did not differ significantly between the different sections of the constructed wetland, the characteristics of zeolite were most appropriate to accommodate a stable and active community of ammonia-oxidizing bacteria. The presence of the helophyte had no effect on the diversity and stability of the ammonia-oxidizing community. SIGNIFICANCE AND IMPACT OF THE STUDY: It has been shown that substrata used in constructed wetlands made no distinction between ammonia-oxidizing strains in relation to attachment. However, zeolite had the best performance with respect to activity over the seasons.

Use of biolog methodology for optimizing the degradation of hydrocarbons by bacterial consortia.

Biolog methodology was used for the preliminary screening of different cultural conditions in order to detect the best combination/s of factors influencing the metabolic performance of bacterial consortia active in the degradation of hydrocarbons. Two microbial consortia were tested for their activity on 2 hydrocarbons (nonadecane and eicosane) in presence of the following cultural coadjuvants: vegetal oil, beta-cyclodextrine, sodium acetate, mineral solution. Tests were conducted in Biolog MT plates, where only the redox indicator of microbial growth (tetrazolium violet) and no carbon sources are provided. The microwells were filled with various combinations of hydrocarbons, microbial inoculum and coadjuvants. Blanks were prepared with the same combinations but without hydrocarbons. The results obtained show the suitability of the methodology developed to identify the most active consortium and the conditions for its best degradation performance. The efficacy of Biolog methodology (Biolog Inc., USA) for the characterization of microbial communities on the basis of the metabolic profiles obtained on specific carbon sources in the microwells of Elisa-type plates, is widely acknowledged (Garland, 1997; Pietikäinen et al., 2000; Dauber and Wolters, 2000). Due to its aptitude to simultaneously evaluate multiple microbial responses and directly organize the results, it can be adapted to meet specific study purposes (Gamo and Shji, 1999). In the present research Biolog methodology was fitted for the preliminary screening of different cultural conditions, in order to detect the best combination/s of factors influencing the metabolic performance of bacterial consortia active in the degradation of aliphatic hydrocarbons, in view of their utilization for the bioremediation of polluted sites.

Identification of spore-forming strains involved in biodegradation of acifluorfen.

We isolated and identified four spore-forming bacteria from activated sludges and soil, three of which were able to degrade acifluorfen. Biochemical characteristics, DNA base composition and DNA-DNA homology indicated that the degrading strains belonged to the species Bacillus thuringiensis, Clostridium perfringens and Clostridium sphenoides. The fourth strain, identified as C. sphenoides and showing the same characteristics of the corresponding degrading strain, was unable to metabolize acifluorfen. Thus, the plasmid content of these strains was analysed to study the possible correlation between the presence of extrachromosomal elements and the ability to degrade this herbicide.

Cometabolic degradation of acifluorfen by a mixed microbial culture.

Laboratory experiments were conducted to study the degradation of acifluorfen 5-[2-chloro-4-(trifluoromethyl)-phenoxyl]-2-nitrobenzoic acid by a mixed microbial population. Concentrations of acifluorfen up to 100 mg/l had no inhibitory effect on the growth of microbial culture. The microorganisms degraded acifluorfen through a cometabolic process in presence of 2-nitrobenzoate. The degradation rate of acifluorfen, determined by liquid chromatography analysis in batch cultures incubated under oxygen-limited conditions were compared. The degradation was slower under oxygen than oxygen-limited conditions. Aminoacifluorfen was produced in both conditions.

Microbial metabolism of fluazifop-butyl.

A microbial mixed culture able to grow on fluazifop-butyl and fluazifop was isolated. Fluazifop degradation by this microbial population was studied either when the herbicide was applied as the sole carbon source or in the presence of a second carbon source (sodium acetate or sodium propionate). The degradation rate was enhanced by sodium propionate. The degradation was found to be stereoselective. The S-enantiomer of fluazifop was degraded at a much higher rate than the R-enantiomer. Fluazifop disappearance was accompanied by formation of three metabolites which were identified by UV, IR, MS and NMR analyses. The metabolites were shown to be: 4-(5-trifluoromethyl-2-pyridyl)oxyphenol, 5-trifluoromethyl-2- hydroxypyridine and 2-(5-trifluoro-methyl pyridyl)hydroxy acetate.

Content of alpha S1-casein and coagulation properties in goat milk.

Samples of goat milk with low and high alpha S1-casein content collected from animals of Alpine and Saanen breeds in the same stage of lactation were compared for coagulation properties (coagulation time, rate of curd formation, curd firmness) and chemical composition (total solids, ash, total protein, total casein, whey protein, fat, Ca, P, pH). Milk with low alpha S1-casein had a faster coagulation time, whereas milk with high levels produced the firmer curd associated with a better chemical composition. Within high alpha S1-casein milk, comparison between breeds showed milk from the Alpine breed had significantly better coagulation properties than that from the Saanen breed. Milk composition accounted for 27% of the variation in coagulation time, 21% of variation in cured formation rate, and 54% of variation in curd firmness.

[Control of the fermentation activity during ensilage of Lolium italicum A. Br. and Medicago sativa L]. / Controllo dell'attivita' fermentativa durante l'insilamento di Lolium italicum A. Br. e Medicago sativa L.

Samples with different dry matter contents of Lolium italicum A. Br., Medicago sativa L., have been ensiled in hermetically sealed containers, in order to study the evolution of microflora and its activity. The principal microbial groups (coliforms, proteolytics , lactic acid bacteria, clostridia, yeasts) have been detected and enumerated, in anaerobic atmosphere, after ensiling periods of 3, 6, 10, 13, 20, 100 days. At the same time, the samples were chemically analyzed for the detection of: pH, moisture, ashes, total and ammoniacal nitrogen, total reducing sugars, lactic acid, volatile fatty acids, short chain alcohols. The relations found between chemical and microbiological data, show that a strong lactic fermentation is not enough for the inhibition of silage spoiling microorganisms, such as coliforms, yeasts, and (less) proteolytics . This phenomenon may be related to the "quality" of the lactic acid microflora, more than to the characteristics of the environment.

[Evolution of microflora during the conservation of forage in microsilos]. / Evoluzione della microflora durante la conservazione di foraggere in microsili.

Fresh and wilted samples of Lolium italicum A. Br., Dactylis glomerata L., Medicago sativa L., have been ensiled in laboratory microsilos for the study of microflora evolution. The principal microbial groups (coliforms, proteolytic bacteria, clostridia, lactic acid bacteria, yeasts) have been detected and enumerated, in anaerobic atmosphere, after 8, 15, 22, 29, 36, 180 days of ensiling. Although the number of lactic acid bacteria resulted to the high in all the samples studied, they failed to perform a quick pH decrease and silage spoiling bacteria were not inhibited. This suggests the possibility of a cohabitation between useful and noxious bacteria, not affected by the chemical composition of ensiled plants.

[Effect of biological activation on the evolution of microflora during composting of urban waste]. / Effetto dell'attivazione biologica sull'evoluzione della microflora durante il compostaggio di rifiuti urbani.

In two solid urban wastes piles, one treated with 7.5% of good bovine manure, the other used as a test, the evolution of the microflora has been studied, during the first period of biodegradation and at the end of the maturation process, in order to control if bioactivation can positively affect the biodegradation development. Actinomytcetes and fungi have been examined, and the following functional microbial groups: aerobic nitrogen fixing bacteria, ammonia bacteria, proteolytics, nitrosants, nitricants, aerobic cellulolytics, amilolytics, pectinolytics. During the process, no significant quantitative difference between the microflora of the two piles was found, except for aerobic cellulolytic, which were strongly superior in number in the end product from the bioactivated pile.