Microbiota of Tayohounta, a fermented baobab flavour food of Benin

The present work provides data on the microbial composition of Tayohounta, a product of natural fermentation of baobab seed kernels. Samples were collected from 3 different small scale producers from Benin at the end of the fermentation process. Microorganisms were enumerated and identified using phenotypic and molecular approaches. Tayohounta was also investigated using culture independent techniques, direct DNA extraction, polymerase chain reaction - denaturing gradient gel electrophoresis (PCR-DGGE) and cloning. Isolated microorganisms were tested for their functionality in baobab seed kernels fermentation. Total viable counts were around 9 log cfu/g representing mainly Bacillus spp., whereas lactic acid bacteria (LAB) (8 log cfu/g), yeasts and moulds represent a smaller part of the total flora in all Tayohounta samples. Sequencing of clones of polymerase chain reaction (PCR) products of bacterial DNA directly extracted from Tayohounta revealed large differences between the products made by different producers. In all products, Bacillus licheniformis, B. pumilus, B. subtilis, B. thermoamylovorans and Lactobacillus fermentum were present. Other microorganisms (B. thuringiensis, Brevibacterium borstelensis, Enterococcus casseliflavus, E. durans, Lb. agilis, Pediococcus pentosaceus, Streptococcus equinus and Weissella confusa) were present occasionally. In experimental pure culture fermentations, B. subtilis showed little effect on pH, but degraded protein and caused a typical pungent smell typical of Tayohounta

Functional implications of the microbial community structure of undefined mesophilic starter cultures

This review describes the recent advances made in the studies of the microbial community of complex and undefined cheese starter cultures. We report on work related to the composition of the cultures at the level of genetic lineages, on the presence and activity of bacteriophages and on the population dynamics during cheese making and during starter culture propagation. Furthermore, the link between starter composition and starter functionality will be discussed. Finally, recent advances in predictive metabolic modelling of the multi-strain cultures will be discussed in the context of microbe-microbe interactions

Contribution of Eat1 and Other Alcohol Acyltransferases to Ester Production in Saccharomyces cerevisiae

Esters are essential for the flavor and aroma of fermented products, and are mainly produced by alcohol acyl transferases (AATs). A recently discovered AAT family named Eat (Ethanol acetyltransferase) contributes to ethyl acetate synthesis in yeast. However, its effect on the synthesis of other esters is unknown. In this study, the role of the Eat family in ester synthesis was compared to that of other Saccharomyces cerevisiae AATs (Atf1p, Atf2p, Eht1p, and Eeb1p) in silico and in vivo. A genomic study in a collection of industrial S. cerevisiae strains showed that variation of the primary sequence of the AATs did not correlate with ester production. Fifteen members of the EAT family from nine yeast species were overexpressed in S. cerevisiae CEN.PK2-1D and were able to increase the production of acetate and propanoate esters. The role of Eat1p was then studied in more detail in S. cerevisiae CEN.PK2-1D by deleting EAT1 in various combinations with other known S. cerevisiae AATs. Between 6 and 11 esters were produced under three cultivation conditions. Contrary to our expectations, a strain where all known AATs were disrupted could still produce, e.g., ethyl acetate and isoamyl acetate. This study has expanded our understanding of ester synthesis in yeast but also showed that some unknown ester-producing mechanisms still exist

Microbiological aspects of processing and storage of edible insects

Growing pressure on the worlds’ livestock production sector and enduring protein undernourishment, persuade the search for alternative protein sources. Insects are widely consumed in many parts of the world and are evaluated as food or supplement. Nevertheless, little attention has been given to the food safety and shelf-life of food insects. An exploratory evaluation of the microbiological content of fresh, processed and stored edible insects was carried out, with focus on farmed mealworm larvae (Tenebrio molitor) and house crickets (Acheta domesticus). A short heating step was sufficient to eliminate Enterobacteriaceae, however some sporeforming bacteria will survive in cooked insects. Simple preservation methods such as drying/acidifying without use of a refrigerator were tested and considered promising. Lactic fermentation of composite flour/water mixtures containing 10, or 20% powdered roasted mealworm larvae resulted in successful acidification and was demonstrated effective in safeguarding shelf-life and safety by the control of Enterobacteria and bacterial spores

Microbial diversity and dynamics of microbial communities during black-slop soaking of soybeans as determined by PCR-DGGE and molecular cloning

Tempe is a traditional fermented food in Indonesia. The manufacture process is quite complex, which comprises two stages, preparatory soaking of soybeans and fungal solid state fermentation. Daily addition of previous soak water (back-slopping) during the soybean soaking step is considered to be crucial in the manufacture of high quality tempe. The microbial diversity and dynamics of the microbial communities evolving during back-slop soaking of soybeans for tempe making was investigated by culture-independent PCR–DGGE and molecular cloning. Both DNA and total RNA were isolated and included in this study, to obtain a view on the succession of total and viable bacteria in the complex microbiota. DGGE profiles indicated that Enterobacter sp., Enterococcus sp., Pseudomonas putida, Leuconostoc fallax, Pediococcus pentosaceus, and Weissella cibaria, were the predominant bacteria. Their occurrence shifted dramatically during the back-slop soaking procedure. This study combined with previous culture-dependent studies could gain a better understanding of the complex microbiota of traditional fermented food and give useful information for its quality control

Enhancing vitamin B₁₂ in lupin tempeh by in situ fortification

Tempeh is a traditional, fungal fermented Indonesian product, usually made from soybeans. Tempeh is known to contain vitamin B12 which is essential for a healthy human diet. Therefore, tempeh is of particular interest for vegan diets since B12 is normally found only in animal derived products. The vitamin B12 in tempeh is associated with the presence of opportunistic pathogens like Klebsiella pneumoniae. Levels of B12 in tempeh do not sustain the recommended daily intake though. In addition, the use of a food-grade bacterium instead of K. pneumoniae is preferred. Lupin can serve as alternative substrate for soybeans due to its similar protein content, resulting in ‘lupin tempeh’. In this study, Propionibacterium freudenreichii, a food-grade, vitamin B12 producing bacterium, was used in co-culture with Rhizopus oryzae to produce B12-enriched lupin tempeh. A significant increase of vitamin B12 content (up to 0.97 μg/100 g) was achieved by fermenting lupin using a mixed starter of R. oryzae and P. freudenreichii. Other parameters, such as texture and volatile organic compounds, were not affected by the bacterial co-inoculation. Therefore, these results are promising for in situ vitamin B12 fortification of lupin tempeh making it a sustainable protein source for a healthy human diet.</p

Fecal Bacterial Communities in Insectivorous Bats from the Netherlands and Their Role as a Possible Vector for Foodborne Diseases

Bats are commonly regarded as vectors for viruses, but little is known about bacterial communities in bats and the possible role of bats in the transmission cycle of foodborne diseases. To gain more insight, microbial communities in fecal samples from 37 insectivorous bats of different species from the Netherlands were investigated by polymerase chain reaction and denaturant gradient gel electrophoresis (PCR-DGGE). Subsequently, 10 samples from the following bat species: common pipistrelle (Pipistrellus pipistrellus; n = 3), Daubenton's bat (Myotis daubentonii; n = 3), serotine bat (Eptesicus serotinus; n = 1), whiskered bat (Myotis mystacinus; n = 1), Geoffroy's bat (Myotis emarginatus; n = 1) and Natterer's bat (Myotis nattereri; n = 1) were selected and used in bacterial 16S rDNA cloning and sequencing. The fecal microbiota in bats was found to be diverse with predominant bacterial genera Carnobacterium, Serratia, Pseudomonas, Enterococcus and Yersinia. The presence of opportunistic pathogens Citrobacter freundii, Escherichia coli, Enterococcus faecalis, Serratia fonticola and Rahnella aquatilis was also recorded. Based on cloning results, we found no proof that bats in the Netherlands are a major vector for the transmission of bacterial zoonotic diseases, although previous findings in literature reported isolation of foodborne pathogens from bats.</p

Performance of non-conventional yeasts in co-culture with brewers’ yeast for steering ethanol and aroma production

Increasing interest in new beer types has stimulated the search for approaches to extend the metabolic variation of brewers’ yeast. Therefore, we tested two approaches using non-conventional yeast to create a beer with lower ethanol content and a complex aroma bouquet. First, the mono-culture performance was monitored of 49 wild yeast isolates of Saccharomyces cerevisiae (16 strains), Cyberlindnera fabianii (9 strains) and Pichia kudriavzevii (24 strains). Interestingly, both C. fabianii and P. kudriavzevii isolates produced relatively more esters compared with S. cerevisiae isolates, despite their limited fermentation capacity. Next, one representative strain of each species (Sc131, Cf65 and Pk129) was applied as co-culture with brewers’ yeast (ratio 1:1). Co-cultures with Cf65 and Pk129 resulted in a beer with lower alcohol content (3.5, 3.8 compared with 4.2% v/v) and relatively more esters. At higher inoculum ratios of Cf65 over brewers’ yeast, growth inhibition of brewers’ yeast was observed, most likely caused by competition for oxygen between brewers’ yeast and Cf65 resulting in a reduced level of ethanol and altered aroma profiles. With this study, we demonstrate the feasibility of using non-conventional yeast species in co-cultivation with traditional brewers’ yeast to tailor aroma profiles as well as the final ethanol content of beer.</p

Performance of non-conventional yeasts in co-culture with brewers’ yeast for steering ethanol and aroma production

Increasing interest in new beer types has stimulated the search for approaches to extend the metabolic variation of brewers’ yeast. Therefore, we tested two approaches using non-conventional yeast to create a beer with lower ethanol content and a complex aroma bouquet. First, the mono-culture performance was monitored of 49 wild yeast isolates of Saccharomyces cerevisiae (16 strains), Cyberlindnera fabianii (9 strains) and Pichia kudriavzevii (24 strains). Interestingly, both C. fabianii and P. kudriavzevii isolates produced relatively more esters compared with S. cerevisiae isolates, despite their limited fermentation capacity. Next, one representative strain of each species (Sc131, Cf65 and Pk129) was applied as co-culture with brewers’ yeast (ratio 1:1). Co-cultures with Cf65 and Pk129 resulted in a beer with lower alcohol content (3.5, 3.8 compared with 4.2% v/v) and relatively more esters. At higher inoculum ratios of Cf65 over brewers’ yeast, growth inhibition of brewers’ yeast was observed, most likely caused by competition for oxygen between brewers’ yeast and Cf65 resulting in a reduced level of ethanol and altered aroma profiles. With this study, we demonstrate the feasibility of using non-conventional yeast species in co-cultivation with traditional brewers’ yeast to tailor aroma profiles as well as the final ethanol content of beer.</p

Bacterial concentration and diversity in fresh tropical shrimps (Penaeus notialis) and the surrounding brackish waters and sediment

This study aimed at determining bacterial concentration and diversity in fresh tropical shrimps (. Penaeus notialis) and their surrounding brackish waters and sediment. Freshly caught shrimp, water and sediment samples were collected in Lakes Nokoue and Aheme in Benin (West Africa) during two periods with different water salinity and temperature. We used complementary culture-dependent and culture-independent methods for microbiota analysis. During both sampling periods, total mesophilic aerobic counts in shrimp samples ranged between 4.4 and 5.9 log CFU/g and were significantly higher than in water or sediment samples. In contrast, bacterial diversity was higher in sediment or water than in shrimps. The dominant phyla were Firmicutes and Proteobacteria in shrimps, Firmicutes, Proteobacteria, and Actinobacteria in water, and Proteobacteria and Chloroflexi in sediment. At species level, distinct bacterial communities were associated with sediment, water and shrimps sampled at the same site the same day. The study suggests that the bacterial community of tropical brackish water shrimps cannot be predicted from the microbiota of their aquatic environment. Thus, monitoring of microbiological quality of aquatic environments might not reflect shrimp microbiological quality