Development of a model for the alcoholic fermentation of cocoa beans by a Saccharomyces cerevisiae strain.

The aromatic quality of chocolate requires the use of cocoa with high aromatic potential, this being acquired during the fermentation of cocoa beans. Traditional fermentation is still often carried out on a small scale with wild strains of yeasts and acetic bacteria and under poorly controlled conditions leading to cocoa quality ranging from best to worst. This study is the first part of a project aiming to control quality of cocoa to produce high aromatic quality chocolate by using a mixed starter of selected strains of yeast and acetic bacteria and by controlling the conditions of fermentation. To achieve this objective, a mathematical model of the alcoholic fermentation of cocoa beans has been developed. The growth, glucose consumption and ethanol production of Saccharomyces cerevisiae LM strain in synthetic broth were modeled for the most important intrinsic (pH, glucose, ethanol, free nitrogen and oxygen levels) and extrinsic (temperature, oxygen level) fermentation parameters. The model was developed by combining the effects of individual conditions in a multiplicative way using the gamma concept. The model was validated in liquid synthetic medium at two different inoculation levels 104 and 106 CFU/mL with an increase in temperature that recorded during spontaneous fermentations. The model clearly shows that the level of inoculation and the speed of the increase in temperature clearly drive yeast growth, while other factors including pH and ethanol, free nitrogen and oxygen levels have no significant impact on yeast growth.

Modeling mixed fermentation of gowé using selected Lactobacillus plantarum and Pichia kluyveri strains.

This paper describes a mixed fermentation model made by assembling block hosting models for the growth of lactic acid bacteria (Lactobacillus plantarum) and a yeast strain (Pichia kluyveri), metabolic production and the physical-chemical changes which occur during the fermentation of gowé. The growth model for P. kluyveri was developed on a synthetic medium following the gamma concept taking into account the effect of pH, temperature, concentrations in glucose, lactic acid and ethanol. Additional parameters for the previously defined L. plantarum growth model were also determined (glucose and ethanol concentrations). The model was validated in three different gowé processing conditions. Even if the model underestimates LAB growth, it explains what occurs in the product and enables in silico extrapolation to various fermentation conditions. The predicted hydrolysis rates of native and gelatinised starches showed that increasing malt content is not an efficient way to increase the sweetness of gowé in contrast to increasing the level of pre-cooking. The builing-block model developed in this study could be applied to many other fermented foods and particularly to non-alcoholic but acid and sweet cereal based beverages.

Modeling cereal starch hydrolysis during simultaneous saccharification and lactic acid fermentation; case of a sorghum-based fermented beverage, gowé.

Gowé is an acidic beverage obtained after simultaneous saccharification and fermentation (SSF) of sorghum. A previous paper focused on modeling the growth of lactic acid bacteria during gowé processing. This paper focuses on modeling starch amylolysis to build an aggregated SSF model. The activity of α-amylase was modeled as a function of temperature and pH, and the hydrolysis rates of both native and soluble starch were modeled via a Michaelis-Menten equation taking into account the maltose and glucose inhibition constants. The robustness of the parameter estimators was ensured by step by step identification in sets of experiments conducted with different proportions of native and gelatinized starch by modifying the pre-cooking temperature. The aggregated model was validated on experimental data and showed that both the pre-cooking and fermentation parameters, particularly temperature, are significant levers for controlling not only acid and sugar contents but also the expected viscosity of the final product. This generic approach could be used as a tool to optimize the sanitary and sensory quality of fermentation of other starchy products.

Modeling Lactic Fermentation of Gowé Using Lactobacillus Starter Culture.

A global model of the lactic fermentation step of gowé was developed by assembling blocks hosting models for bacterial growth, lactic acid production, and the drop of pH during fermentation. Commercial strains of Lactobacillus brevis and of Lactobacillus plantarum were used; their growth was modeled using Rosso's primary model and the gamma concept as a secondary model. The optimum values of pH and temperature were 8.3 ± 0.3, 44.6 ± 1.2 °C and 8.3 ± 0.3, 3.2 ± 37.1 °C with µmax values of 1.8 ± 0.2 and 1.4 ± 0.1 for L. brevis and L. plantarum respectively. The minimum inhibitory concentration of undissociated lactic acid was 23.7 mM and 35.6 mM for L. brevis and L. plantarum, respectively. The yield of lactic acid was five times higher for L. plantarum than for L. brevis, with a yield of glucose conversion to lactic acid close to 2.0 for the former and 0.8 for the latter. A model was developed to predict the pH drop during gowé fermentation. The global model was partially validated during manufacturing of gowé. The global model could be a tool to aid in the choice of suitable starters and to determine the conditions for the use of the starter.

Prevalence and fate of Bacillus cereus in African traditional cereal-based foods used as infant foods.

The objective of the present work was to estimate the prevalence of Bacillus cereus group species in traditional cereal-based lactic acid-fermented slurries and nonfermented flours used to prepare infant foods in an African context. High counts on mannitol-egg yolk-polymixin agar medium were determined for the fermented slurries (median, 4.5 × 10(4) CFU/ml of slurry) compared with the nonfermented flours, most of whose counts were lower than 10(-1) CFU/g. Virulence genes were characterized in 60 isolates from 26 traditional cereal-based foods in Ouagadougou (Burkina Faso). Seventy-two and 38 % of isolates were positive for the complete set of genes coding for hemolysin BL and nonhemolytic enterotoxin, respectively, suggesting a high enterotoxigenic potential for these foodborne isolates. No potentially emetic toxin-producing strains were detected. Because of the high counts found for fermented slurries, survival tests with vegetative cells inoculated in fermented slurries were performed, which showed that growth of B. cereus was inhibited. This result suggests that fermentation in traditional production units is presumably not adequately controlled, enabling growth during any unit operations before fermentation, or even during the fermentation step, when the process was poorly controlled. However, adding nisin (0.1 mg/ml) enabled a 5-log reduction in the B. cereus population in 5 h, suggesting that the use of nisin could be a way to upgrade the hygienic quality of this type of food.

Feasibility of coupling dehydration-impregnation by soaking treatment of meat with fermentation by Lactobacillus sakei.

UNLABELLED: This study examined the feasibility of coupling dehydration-impregnation by soaking (DIS) with a subsequent lactic fermentation in the treatment of meat. A series of beef fillets were subjected to 3 different DIS treatments. The resulting DIS-treated fillets had 3 different characteristics in terms of water activity, salt, and fermentable sugars contents. Fillets treated with the DIS with the shortest immersion time (5 h) and the highest salt concentration in the DIS bath (100 g/L) were inoculated with Lactobacillus sakei. A control group was left without inoculation. After 24 h incubation at 25 °C, only inoculated fillets showed signs of lactic fermentation. At 24 h, these fillets had a d-lactic acid content of 68 µmol/g dry basis and a high population of L. sakei revealed by methods of plate count and quantitative PCR. DIS could therefore be compatible with a subsequent fermentation step by L. sakei. PRACTICAL APPLICATION: Traditional meat preservation processes often combine unit operations such as salting, smoking, fermentation, and drying. In tropical countries, high temperatures and high relative humidity, poor infrastructure, and improper slaughterhouse practices explain the need for more drastic processes (more salt, more water loss) for meat preservation. Dehydration-impregnation by soaking (DIS) could be used as a rapid pretreatment of meat, in order to counteract tropical conditions. This study validates a novel approach whereby DIS is coupled with lactic fermentation by surface inoculation with Lactobacillus sakei. With a final drying step this process could be used for the treatment of whole meat pieces.

Toxigenic status of Staphylococcus aureus isolated from bovine raw milk and Minas frescal cheese in Brazil.

A group of 291 Staphylococcus aureus isolates from mastitic cow's milk (n = 125), bulk tank milk (n = 96), and Minas frescal cheese (n = 70) were screened for staphylococcal enterotoxin (SE) genes (sea, seb, sec, sed, see, seg, seh, sei, selj, and sell) and for the tst-1 gene encoding staphylococcal toxic shock syndrome toxin 1 by PCR assay. A total of 109 (37.5%) of the isolates were positive for at least one of these 11 genes, and 23 distinct genotypes of toxin genes were observed. Of the S. aureus isolates bearing SE genes, 17 (13.6%) were from mastitic cow's milk, 41 (41.7%) were from bulk tank milk, and 51 (72.9%) were from Minas frescal cheese. The occurrence of exclusively more recently described SE genes (seg through sell) was considerably higher (87 of 109 PCR-positive strains) than that of classical SE genes (sea through see, 15 strains). The SE genes most commonly detected were seg and sei; they were found alone or in different combinations with other toxin genes, but in 60.8% of the cases they were codetected. No strain possessed see. The tst-1 gene was found in eight isolates but none from mastitic cow's milk. Macrorestriction analysis of chromosomal DNA from 89 S. aureus isolates positive for SE gene(s) was conducted with the enzyme SmaI. Fifty-five distinct pulsed-field gel electrophoresis patterns were found, demonstrating a lack of predominance of any specific clone. A second enzyme, Apa I, used for some isolates was less discriminating than Sma I. The high genotype diversity of potential toxigenic S. aureus strains found in this study, especially from Minas frescal cheese, suggests various sources of contamination. Efforts from the entire production chain are required to improve consumer safety.

Physiological change in camel milk composition (Camelus dromedarius) 1. Effect of lactation stage.

The change in the composition of camel milk in four dromedaries was studied by including the common measured parameters: protein, total fat, lactose, main minerals (calcium, phosphorus, and iron), and vitamin C. The fat matter varied from 4.34% to 7.81% with a slight decrease all along the lactation and a minimal value at the 14th week corresponding to the lactation peak. Those variations were less important for protein content (from 2.58% to 3.64%), but the minimal value was observed at the 14th week also. The lactose varied slightly around its mean of 3.46%. The vitamin C concentration varied from 48 to 256 mg/l with a tendency of increasing all along the lactation. Calcium and phosphorus concentrations were quite parallel and their ratio Ca/P was constant. The minimal values (1.43 g/l for calcium and 1.16 g/l for phosphorus) were observed at the beginning of the lactation. The iron concentrations varied around the mean of 1.73 mg/l.

Determination of fruit origin by using 26S rDNA fingerprinting of yeast communities by PCR-DGGE: preliminary application to Physalis fruits from Egypt.

The determination of geographical origin is a demand of the traceability system of import-export food products. One hypothesis for tracing the source of a product is by global analysis of the microbial communities of the food and statistical linkage of this analysis to the geographical origin of the food. For this purpose, a molecular technique employing 26S rDNA profiles generated by PCR-DGGE was used to detect the variation in yeast community structures of three species of Physalis fruit (Physalis ixocarpa Brat, Physalis pubescens L, Physalis pruinosa L) from four Egyptian regions (Qalyoubia, Minufiya, Beheira and Alexandria Governments). When the 26S rDNA profiles were analysed by multivariate analysis, distinct microbial communities were detected. The band profiles of Physalis yeasts from different Governments were specific for each location and could be used as a bar code to discriminate the origin of the fruits. This method is a new traceability tool which provides fruit products with a unique biological bar code and makes it possible to trace back the fruits to their original location.

Study on the microflora and biochemistry of cocoa fermentation in the Dominican Republic.

Cocoa fermentation was monitored at the IDIAF (Instituto Dominicano de Investigaciones Agropecuarias y Forestales) "Mata Larga" experimental station, in San Francisco de Macoris, Dominican Republic. The maximum average fermentation temperature reached 51 degrees C after 48 h and the pH reached 4.5 after 144 h of fermentation. A significant decrease in glucose, fructose and citric acid was seen in the pulp over the first 48 h. There was a delay of 24 h between maximum microbial growth and maximum concentrations of the respective metabolites, which occurred after 48 h for ethanol and after 72 h for acetic acid. A maximum concentration in lactic acid was found after around 120 h of fermentation. The aerobic mesophilic flora increased from 6.1x10(6) to a maximum of 4.2x10(7) CFU g(-1) of dry matter after 48 h of fermentation. Yeasts displayed maximum development after 24 h (6.1x10(7) CFU g(-1) of dry matter), whilst for lactic and acetic acid bacteria it occurred after 48 h (7.3x10(7) and 1.5x10(8) CFU g(-1) of dry matter respectively). The yeasts isolated belonged to the genera Hanseniaspora and Candida, the lactic acid bacteria to the genus Lactobacillus, and the acetic acid bacteria to the genus Acetobacter. The differences compared to other fermentation trials concerned the micropopulation from a qualitative point of view.

Effect of the lactoperoxidase system against three major causal agents of disease in mangoes.

The antibacterial activity of the lactoperoxidase system (LPS) on the growth of Xanthomonas campestris, the causal agent of bacterial black spot in mangoes, Botryodiplodia theobromae, the causal agent of stem-end rot disease in mangoes, and Colletotrichum gloeosporioides, the causal agent of anthracnose disease in mangoes, was determined during culture at 30 degrees C and at several pH values (4.5, 5.5, and 6.5). When the results of using the LPS were compared with those from control cultures without the LPS reagents, the growth of the three microorganisms was totally inhibited in all of the conditions tested. Viability tests enumerating cultivable cells of X. campestris showed that the LPS had a bactericidal effect, whatever the pH value. This effect is faster at pH 5.5, corroborating the results reported in the literature (optimal pH for the LPS efficiency). Further, we proved that hydrogen peroxide alone had little inhibition effect on the growth of the microorganisms studied. This compound is essentially used to convert thiocyanate into hypothiocyanate during the lactoperoxidase reaction. The potential of the LPS for the postharvest treatment of the fruits for controlling microbial diseases was thus demonstrated. Nevertheless, further studies are needed on fresh fruits before envisaging any application.

Evaluation of lactoperoxidase system treatment to reduce anthracnose, stem-end rot, and bacterial black spot development during storage of mangoes.

The lactoperoxidase system (LPS) was evaluated for the prevention of postharvest diseases caused by Xanthomonas campestris, Botryodiplodia theobromae, and Colletotrichum gloeosporioides in 'Keitt' and 'Kent' mangoes. The LPS treatment significantly reduced the disease development on both cultivars after storage at 12 degrees C for 2 weeks, which was followed by a ripening at 25 degrees C. The LPS treatment did not alter the sensory quality of mango fruits (color, firmness, titrable acidity, and total soluble solids) when compared to untreated fruits. The LPS thus presents good potential alternative to the chemical fungicides traditionally used to improve the shelf life of mangoes.

Use of ATP bioluminescence to determine the bacterial sensitivity threshold to a bacteriocin.

A new ATP bioluminescence-based method was developed to determine the effectiveness of nisin on a sensitive strain of Lactococcus cremoris. The principle of the method is to quantify the release of adenylic-nucleotides (AN) by a sensitive strain under the action of the bacteriocin, with the complex luciferin-luciferase. Nisin-induced leakage of AN included ATP from a sensitive L. cremoris to the external medium immediately after the contact with the bacteria. The growth of L. cremoris was correlated with the extracellular AN content. The extracellular ATP and AN concentration exhibited a linear correlation to the logarithm of the nisin concentration. For the determination of the effectiveness threshold, the concentration of AN was more sensitive and more reliable than the direct quantification of ATP. The effectiveness threshold, corresponding to a 100% inhibition of L. cremoris growth, was obtained for a null concentration of intracellular nucleotides, i.e. for a AN(tot):AN(ext) ratio = 1. For an initial concentration of 1.4 x 10(7) bacteria/mL, the nisin effectiveness threshold is 3.4 +/- 0.01 mg nisin/L. It is possible to detect effectiveness threshold concentration by taking into account the physiological state of the cells.