Artisanal and farmer bread making practices differently shape fungal species community composition in French sourdoughs

Preserving microbial diversity in food systems is one of the many challenges to be met to achieve food security and quality. Although industrialization led to the selection and spread of specific fermenting microbial strains, there are still ongoing artisanal processes that may allow the conservation of a wider species diversity and genetic diversity. We examined whether the diversity of artisanal practices could lead to an increased level in fungal species diversity for bread making. We used an interdisciplinary participatory research approach including bakers, psycho-sociologists and microbiologists to analyze French bread making practices and describe fungal communities in naturally fermented sourdough of 27 bakers and 12 farmer bakers. Bread making practices were classified in two groups: the farmer-like practice group and the artisanal-like practice group. The well-known bakery yeast, Saccharomyces cerevisiae, was dominant (i.e. with a relative abundance over 50%) in only 24% of sourdoughs while other yeast species, belonging to the Kazachstania genus, were dominant in 54% of sourdoughs. Bread making practices were found to drive the distribution of fungal species across sourdoughs. The most striking bread making practice effect was the occurrence of Kazachstania humilis in sourdoughs made with artisanal-like practices and the occurrence of Kazachstania bulderi in sourdoughs made with farmer-like practices. Phenotypic divergences between sourdough and non-sourdough strains were found for K. humilis but not for K. bulderi. Overall, our results showed that preserving bread making practice diversity allows the preservation of a higher species and phenotypic diversity in microbial communities

Very early acetaldehyde production by industrial Saccharomyces cerevisiae strains: a new intrinsic character

International audienceDuring a general survey of the acetaldehyde-producing properties of commercially available wine yeast strains, we discovered that, although final acetaldehyde production cannot be used as a discriminating factor between yeast strains, initial specific acetaldehyde production rates were of highly interest for classifying yeast strains. This parameter is very closely related to the growth- and fermentation-lag phase durations. We also found that this acetaldehyde early production occurs with very different extent between commercial active dry yeast strains during the rehydration phase and could partially explain the known variable resistance of yeast strains to sulfites. Acetaldehyde production appeared, therefore, as very precocious, strain-dependent, and biomass-independent character. These various findings suggest that this new intrinsic characteristic of industrial fermenting yeast may be likely considered as an early marker of the general fermenting activity of industrial fermenting yeasts. This phenomenon could be particularly important for understanding the ecology of colonization of complex fermentation media by Saccharomyces cerevisiae

Early lysis of Lactobacillus helveticus CNRZ 303 in Swiss cheese is not prophage-related

Lactobacillus helveticus is mainly used as starter in Swiss-type cheeses. Often, lysogenic strains are eliminated because of the risk of early lysis and acidification failure due to phage expression. On the other hand, L. helveticus lysis was shown to positively influence cheese proteolysis during ripening. In order to better assess the relationship between lysis and lysogeny, a prophage-cured derivative of L. helveticus CNRZ 303 was isolated (LH 303-G11) and relysogenised (LH 303-G11R), as demonstrated by hybridisation using the whole phage DNA as probe. The growth, lysis in buffered solutions and lytic activities in zymogram using either Micrococcus luteus or L. helveticus as substrate were identical between the mother strain and its curedderivatives. Only morphological differences were observed by scanning electron microscopy: the cells of the cured derivative were shorter in length. The mother strain and its cured and relysogenised derivatives were assayed in triplicate in experimental Swiss cheeses (scale 1:100). No differences were noted during the cheese making: the three strains exhibited identical kinetics of acidification, leading to similar cheeses at day 1 in terms of gross composition and pH. Phages were detected only in the cheeses made with the mother strain and the relysogenised derivative. The lysis of L. helveticus, estimated by viability decrease and release of the intracellular marker D-lactate deshydrogenase, started early before brining and continued during the coldroom ripening. No obvious differences of lysis extent were observed. These results demonstrated for the first time that, in the case of LH 303, the extensive lysis observed in cheese is mainly due to autolysin activity and not to prophage induction

Mur-LH, the Broad-Spectrum Endolysin of Lactobacillus helveticus Temperate Bacteriophage phi-0303

-0303 is a temperate bacteriophage isolated from Lactobacillus helveticus CNRZ 303 strain after mitomycinC induction. In this work, the gene coding for a lytic protein of this bacteriophage was cloned using a libraryof -0303 in Escherichia coli DH5 . The lytic activity was detected by its expression, using whole cells of thesensitive strain L. helveticus CNRZ 892 as the substrate. The lysin gene was within a 4.1-kb DNA fragment of -0303 containing six open reading frames (ORFs) and two truncated ORFs. No sequence homology with holingenes was found within the cloned fragment. An integrase-encoding gene was also present in the fragment, butit was transcribed in a direction opposite that of the lysin gene. The lysin-encoding lys gene was verified by PCRamplification from the total phage DNA and subcloned. The lys gene is a 1,122-bp sequence encoding a proteinof 373 amino acids (Mur-LH), whose product had a deduced molecular mass of 40,207 Da. Comparisons withsequences in sequence databases showed homology with numerous endolysins of other bacteriophages.Mur-LH was expressed in E. coli BL21, and by renaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresiswith L. helveticus CNRZ 892 as the substrate, the recombinant protein showed an apparent molecularmass of 40 kDa. The N-terminal sequence of the protein confirmed the start codon. Hydrolysis of cell walls ofL. helveticus CNRZ 303 by the endolysin and biochemical analysis of the residues produced demonstrated thatMur-LH has N-acetylmuramidase activity. Last, the endolysin exhibited a broad spectrum of lytic activity, asit was active on different species, mainly thermophilic lactobacilli but also lactococci, pediococci, Bacillussubtilis, Brevibacterium linens, and Enterococcus faecium

Early lysis of Lactobacillus helveticus CNRZ 303 in Swiss cheese is not prophage-related

Lactobacillus helveticus is mainly used as starter in Swiss-type cheeses. Often, lysogenic strains are eliminated because of the risk of early lysis and acidification failure due to phage expression. On the other hand, L. helveticus lysis was shown to positively influence cheese proteolysis during ripening. In order to better assess the relationship between lysis and lysogeny, a prophage-cured derivative of L. helveticus CNRZ 303 was isolated (LH 303-G11) and relysogenised (LH 303-G11R), as demonstrated by hybridisation using the whole phage DNA as probe. The growth, lysis in buffered solutions and lytic activities in zymogram using either Micrococcus luteus or L. helveticus as substrate were identical between the mother strain and its curedderivatives. Only morphological differences were observed by scanning electron microscopy: the cells of the cured derivative were shorter in length. The mother strain and its cured and relysogenised derivatives were assayed in triplicate in experimental Swiss cheeses (scale 1:100). No differences were noted during the cheese making: the three strains exhibited identical kinetics of acidification, leading to similar cheeses at day 1 in terms of gross composition and pH. Phages were detected only in the cheeses made with the mother strain and the relysogenised derivative. The lysis of L. helveticus, estimated by viability decrease and release of the intracellular marker D-lactate deshydrogenase, started early before brining and continued during the coldroom ripening. No obvious differences of lysis extent were observed. These results demonstrated for the first time that, in the case of LH 303, the extensive lysis observed in cheese is mainly due to autolysin activity and not to prophage induction