Serial fermentation in milk generates functionally diverse community lineages with different degrees of structure stabilization.

Microbial communities offer considerable potential for tackling environmental challenges by improving the functioning of ecosystems. Top-down community engineering is a promising strategy that could be used to obtain communities of desired function. However, the ecological factors that control the balance between community shaping and propagation are not well understood. Dairy backslopping, which consists of using part of the previous production to inoculate a new one, can be used as a model engineering approach to investigate community dynamics during serial propagations. In this study, 26 raw milk samples were serially propagated 6 times each, giving rise to 26 community lineages. Bacterial community structures were analyzed by metabarcoding, and acidification was recorded by pH monitoring. The results revealed that different types of community lineages could be obtained in terms of taxonomic composition and dynamics. Five lineages reached a repeatable community structure in a few propagation steps, with little variation between the final generations, giving rise to stable acidification kinetics. Moreover, these stabilized communities presented a high variability of structure and diverse acidification properties between community lineages. Besides, the other lineages were characterized by different levels of dynamics leading to parallel or divergent trajectories. The functional properties and dynamics of the communities were mainly related to the relative abundance and the taxonomic composition of lactic acid bacteria within the communities. These findings highlight that short-term schemes of serial fermentation can produce communities with a wide range of dynamics and that the balance between community shaping and propagation is intimately linked to community structure. IMPORTANCE: Microbiome applications require approaches for shaping and propagating microbial communities. Shaping allows the selection of communities with desired taxonomic and functional properties, while propagation allows the production of the biomass required to inoculate the engineered communities in the target ecosystem. In top-down community engineering, where communities are obtained from a pool of mixed microorganisms by acting on environmental variables, a major challenge is to master the balance between shaping and propagation. However, the ecological factors that favor high dynamics of community structure and, conversely, those that favor stability during propagation are not well understood. In this work, short-term dairy backslopping was used to investigate the key role of the taxonomic composition and structure of bacterial communities on their dynamics. The results obtained open up interesting prospects for the biotechnological use of microbiomes, particularly in the field of dairy fermentation, to diversify approaches for injecting microbial biodiversity into cheesemaking processes.

Exploring the Effects of Freeze-Dried Sourdoughs with Lactiplantibacillus pentosus 129 and Limosilactobacillus fermentum 139 on the Quality of Long-Fermentation Bread.

Sourdough production is a complex fermentation process. Natural sourdough fermentation without standardization causes great variability in microbial communities and derived products. Starter cultures have emerged as alternatives to natural fermentation processes, which could improve bakery quality and produce bioactive compounds. This study aimed to evaluate the impacts of freeze-drying on the production and viability of sourdoughs with Lactiplantibacillus pentosus 129 (Lp) and Limosilactobacillus fermentum 139 (Lf), as well as their effects on the quality of long-fermentation bread. These strains were selected based on their better performance considering acidification and exopolysaccharide production capacity. Sourdough with Lp and Lf were propagated until the 10th day, when physicochemical and microbiological parameters were determined. The produced sourdoughs were freeze-dried, and bread samples were produced. The freeze-drying process resulted in high survival rates and few impacts on the metabolic activity of Lp and Lf until 60 days of storage. Incorporating Lp and Lf improved the microbiological and physicochemical properties of sourdough and long-fermentation breads. Tested freeze-dried sourdoughs led to reduced bread aging (higher specific volume and decreased starch retrogradation) and increased digestibility. The results show the potential of the freeze-dried sourdoughs produced with Lp and Lf as innovative strategies for standardizing production protocols for the bakery industry, especially for producing long-term fermentation bread.

Real-time PCR-based quantitative microbiome profiling elucidates the microbial dynamic succession in backslopping fermentation of Taiwanese pickled cabbage.

BACKGROUND: Microbiota succession determines the flavor and quality of fermented foods. Quantitative PCR-based quantitative microbiome profiling (QMP) has been applied broadly for microbial analysis from absolute abundance perspectives, transforming microbiota ratios into counts by normalizing 16S ribosomal RNA (16S rRNA) gene sequencing data with gene copies quantified by quantitative PCR. However, the application of QMP in fermented foods is still limited. RESULTS: QMP elucidated microbial succession of Taiwanese pickled cabbage. In the spontaneous first-round fermentation (FR), the 16S rRNA gene copies of total bacteria increased from 6.1 to 10 log copies mL-1. The dominant lactic acid bacteria genera were successively Lactococcus, Leuconostoc and Lactiplantibacillus. Despite the decrease in the proportion of Lactococcus during the succession, the absolute abundance of Lactococcus still increased. In the backslopping second-round fermentation (SR), the total bacteria 16S rRNA gene copies increased from 7.6 to 9.9 log copies mL-1. The addition of backslopping starter and vinegar rapidly led to a homogenous microbial community dominated by Lactiplantibacillus. The proportion of Lactiplantibacillus remained consistently around 90% during SR, whereas its absolute abundance exhibited a continuous increase. In SR without vinegar, Leuconostoc consistently dominated the fermentation. CONCLUSION: The present study highlights that compositional analysis would misinterpret microbial dynamics, whereas QMP reflected the real succession profiles and unveiled the essential role of vinegar in promoting Lactiplantibacillus dominance in backslopping fermentation of Taiwanese pickled cabbage. Quantitative microbiome profiling (QMP) was found to be a more promising approach for the detailed observation of microbiome succession in food fermentation compared to compositional analysis. © 2024 Society of Chemical Industry.

Evaluation of Microbial Dynamics of Kombucha Consortia upon Continuous Backslopping in Coffee and Orange Juice.

The kombucha market is diverse, and competitors constantly test new components and flavours to satisfy customers' expectations. Replacing the original brewing base, adding flavours, or using "backslopping" influence the composition of the symbiotic starter culture of bacteria and yeast (SCOBY). Yet, deep characterisation of microbial and chemical changes in kombucha consortia in coffee and orange juice during backslopping has not been implemented. This study aimed to develop new kombucha beverages in less-conventional matrices and characterise their microbiota. We studied the chemical properties and microbial growth dynamics of lactic-acid-bacteria-tailored (LAB-tailored) kombucha culture by 16S rRNA next-generation sequencing in coffee and orange juice during a backslopping process that spanned five cycles, each lasting two to four days. The backslopping changed the culture composition and accelerated the fermentation. This study gives an overview of the pros and cons of backslopping technology for the production of kombucha-based beverages. Based on research conducted using two different media, this work provides valuable information regarding the aspects to consider when using the backslopping method to produce novel kombucha drinks, as well as identifying the main drawbacks that need to be addressed.

Role of lactic acid bacteria and yeasts in sourdough fermentation during breadmaking: Evaluation of postbiotic-like components and health benefits.

Sourdough (SD) fermentation is a traditional biotechnological process used to improve the properties of baked goods. Nowadays, SD fermentation is studied for its potential health effects due to the presence of postbiotic-like components, which refer to a group of inanimate microorganisms and/or their components that confer health benefits on the host. Some postbiotic-like components reported in SD are non-viable microorganisms, short-chain fatty acids, bacteriocins, biosurfactants, secreted proteins/peptides, amino acids, flavonoids, exopolysaccharides, and other molecules. Temperature, pH, fermentation time, and the composition of lactic acid bacteria and yeasts in SD can impact the nutritional and sensory properties of bread and the postbiotic-like effect. Many in vivo studies in humans have associated the consumption of SD bread with higher satiety, lower glycemic responses, increased postprandial concentrations of short-chain fatty acids, and improvement in the symptoms of metabolic or gastrointestinal-related diseases. This review highlights the role of bacteria and yeasts used for SD, the formation of postbiotic-like components affected by SD fermentation and the baking process, and the implications of functional SD bread intake for human health. There are few studies characterizing the stability and properties of postbiotic-like components after the baking process. Therefore, further research is necessary to develop SD bread with postbiotic-related health benefits.

Effects of Extraction Methods on Physicochemical and Structural Properties of Common Vetch Starch.

Three extraction methods: water extraction, lactic acid bacteria fermentation, and back-slopping fermentation were applied to extract a new type of legume starch, common vetch starch. Our results showed that the lactic acid bacteria fermented starch had the highest amylose content (35.69%), followed by the back-slopping fermented starch (32.34%), and the water-extracted starch (30.25%). Furthermore, erosion surface, lower molecular weight, smaller particle size, larger specific surface area, and a higher proportion of B1 chain were observed in the fermented starch, especially in the back-slopping fermented starch. All the extracted starches showed a type C structure, but a type CB structure was observed in the back-slopping fermented starch. In addition, the relative crystallinity of the lactic acid bacteria fermented starch (34.16%) and the back-slopping fermented starch (39.43%) was significantly higher than that of the water-extracted starch (30.22%). Moreover, the swelling power, solubility, pasting, and thermal properties of the fermented starches were also improved. In conclusion, the fermentation extraction method, especially back-slopping fermentation, could improve the quality of the extracted common vetch starch when compared with the traditional water extraction method.

Backslopping Time, Rinsing of the Grains During Backslopping, and Incubation Temperature Influence the Water Kefir Fermentation Process.

For eight backslopping steps, eight series of water kefir fermentation processes differing in backslopping time and rinsing of the grains during each backslopping step and eight series of fermentation processes differing in incubation temperature and backslopping time were followed. Short backslopping times resulted in high relative abundances of Liquorilactobacillus nagelii and Saccharomyces cerevisiae, intermediate backslopping times in high relative abundances of Leuconostoc pseudomesenteroides, and long backslopping times in high relative abundances of Oenococcus sicerae and Dekkera bruxellensis. When the grains were rinsed during each backslopping step, the relative abundances of Lentilactobacillus hilgardii and Leuc. pseudomesenteroides increased and those of D. bruxellensis and Liql. nagelii decreased. Furthermore, rinsing of the grains during each backslopping step resulted in a slightly higher water kefir grain growth and lower metabolite concentrations. The relative abundances of Liquorilactobacillus mali were highest at 17°C, those of Leuc. pseudomesenteroides at 21 and 25°C, and those of Liql. nagelii at 29°C. With a kinetic modeling approach, the impact of the temperature and rinsing of the grains during the backslopping step on the volumetric production rates of the metabolites was determined.

Brine salt concentration reduction and inoculation with autochthonous consortia: Impact on Protected Designation of Origin Nyons black table olive fermentations.

Nyons table olives, named after the French city where they are processed, are naturally fermented black table olives. Their specificity relies on the use of the "Tanche" olive variety harvested at full maturity and their slow spontaneous fermentation in 10% salt brine driven by yeast populations. This study aimed at investigating the benefit of inoculating autochthonous consortia to produce Nyons table olives by fermentation in 10% salt brine and in reduced salt conditions (8%). Two strategies were evaluated: inoculation with a defined autochthonous consortium and inoculation by spent brine backslopping. To define the consortium, yeasts were selected among 48 autochthonous isolates and key features included high halotolerance, low pectinolytic and proteolytic activities, however none had ß-glucosidase activities. The consortium included eight yeast strains with distinct technological properties belonging to five dominant species, i.e. Citeromyces nyonsensis, Pichia membranifaciens, Wickerhamomyces anomalus, Zygotorulaspora mrakii and Candida atlantica. Fermentation trials were conducted over a year and compared by evaluating microbial community shifts (16S and ITS metagenetics) and volatile profiles (GC-MS). Regarding fermentations with the defined consortium, four out of five species implanted in early stages while one, Pichia membranifaciens, persisted and largely dominated by the end of the fermentation. Altogether, inoculation with the defined consortium did not disrupt microbial shifts compared to traditional fermentations although minor differences were observed in volatile profiles. The backslopping method yielded the highest impact on microbial populations and olive volatile profiles, with higher ester abundances at the end of fermentation. Finally, reduced salt in brine gave very promising results as no deleterious effects on microbial communities, volatile dynamics but also safety criteria of the olives were observed compared to traditional fermented olives.

Dynamic interactions of lactic acid bacteria in Korean sourdough during back-slopping process.

AIMS: This study aimed to clarify the cause of quality reduction in Korean sourdough after successive back-slopping. METHODS AND RESULTS: We investigated the dynamic changes in lactic acid bacteria during the back-slopping process using genetic fingerprinting techniques. During the initial propagation phases, the dominant lactic acid bacteria were Fructilactobacillus sanfranciscensis (<5 log CFU per g sourdough), Latilactobacillus curvatus (9·5 log CFU per g sourdough) and Levilactobacillus brevis (6·5 log CFU per g sourdough). However, after the 11th propagation, F. sanfranciscensis became more prominent (>9·0 log CFU per g sourdough), whereas L. curvatus and L. brevis rapidly decreased. Monitoring these bacteria in the co-culture system revealed that acid-tolerant F. sanfranciscensis rapidly utilized maltose (1·65 g l-1  h-1 ) and produced large amounts of lactic acid, whereas L. brevis and L. curvatus consumed maltose slowly and L. curvatus was poorly tolerant to lactic acid. CONCLUSION: The results indicate that competition exists between the lactic acid bacteria in sourdough during the back-slopping process, and microbial succession by acid-tolerant species results in quality reduction of sourdough. SIGNIFICANCE AND IMPACT OF THE STUDY: This study uncovered the cause of microbial changes during the propagation of Korean sourdough and proposed a strategy to develop starters to produce high-quality bakery products.

Linking Pélardon artisanal goat cheese microbial communities to aroma compounds during cheese-making and ripening.

Pélardon is an artisanal French raw goat's milk cheese, produced using natural whey as a backslop. The aim of this study was to identify key microbial players involved in the acidification and aroma production of this Protected Designation of Origin cheese. Microbial diversity of samples, collected from the raw milk to 3-month cheese ripening, was determined by culture-dependent (MALDI-TOF analysis of 2877 isolates) and -independent (ITS2 and 16S metabarcoding) approaches and linked to changes in biochemical profiles (volatile compounds and acids). In parallel, potential dominant autochthonous microorganism reservoirs were also investigated by sampling the cheese-factory environment. Complex and increasing microbial diversity was observed by both approaches during ripening although major discrepancies were observed regarding Lactococcus lactis and Lacticaseibacillus paracasei fate. By correlating microbial shifts to biochemical changes, Lactococcus lactis was identified as the main acidifying bacterium, while L. mesenteroides and Geotrichum candidum were prevalent and associated with amino acids catabolism after the acidification step. The three species were dominant in the whey (backslop). In contrast, L. paracasei, Enterococcus faecalis, Penicillium commune and Scopulariopsis brevicaulis, which dominated during ripening, likely originated from the cheese-making environment. All these four species were positively correlated to major volatile compounds responsible for the goaty and earthy Pélardon cheese aroma. Overall, this work highlighted the power of MALDI-TOF and molecular techniques combined with volatilome analyses to dynamically follow and identify microbial communities during cheese-making and successively identify the key-players involved in aroma production and contributing to the typicity of Pélardon cheese.

Diversity and dynamics of sourdough lactic acid bacteriota created by a slow food fermentation system.

Sourdough is a naturally fermented dough that is used worldwide to produce a variety of baked foods. Various lactic acid bacteria (LAB), which can determine the quality of sourdough baked foods by producing metabolites, have been found in the sourdough ecosystem. However, spontaneous fermentation of sourdough leads to unpredictable growth of various micro-organisms, which result in unstable product quality. From an ecological perspective, many researchers have recently studied sourdough LAB diversity, particularly the elucidation of LAB community interactions and the dynamic mechanisms during the fermentation process, in response to requests for the control and design of a desired sourdough microbial community. This article reviews recent advances in the study of sourdough LAB diversity and its dynamics in association with unique characteristics of the fermentation system; it also discusses future perspectives for better understanding of the complex sourdough microbial ecosystem, which can be attained efficiently by both in vitro and in situ experimental approaches.

Divergent role of abiotic factors in shaping microbial community assembly of paocai brine during aging process.

Microbial communities in fermented food are shaped by a myriad of abiotic factors. The respective roles of abiotic factors in shaping the dynamic bacterial community of paocai during aging remain unclear. In the present study, 100 paocai samples (pH: 2.95-5.23; NaCl content: 0.13-15.41%; total acid: 6.61-18.33 mg/mL; total sugars: 7.96-487.90 µg/mL; total viable count: 3.55-8.99 LogCFU/mL; aging time: 5 day-15 year) were analyzed through high-throughput sequencing and the results revealed five dominant bacterial genera across different samples, including Lactobacillus, Pediococcus, Leuconostoc, Lactococcus and unclassified genera. Both NaCl and total acid (TA) were the major factors regulating bacterial community divergence in paocai. Based on these results, the microbial communities were reconstructed by manipulating the TA and NaCl contents in vitro to validate the effectiveness of these factors in shaping microbial communities during paocai fermentation. Results showed that roles of abiotic factors differentiated during fermentation. At the early stage, salt was the first abiotic filter, mainly working through promoting the abundance of Lactococcus and Leuconostoc. As the TA content increased, the selective role of salt weakened while acid became the dominant at the later stage, as evidenced by the increased abundance of Lactobacillus and Pediococcus following the increase of TA content.

The influence of backslopping on lactic acid bacteria diversity in tarhana fermentation.

Tarhana is produced at batch systems in which the microbiota has changed accordingly to the microbial load from ingredients. In order to stabilize the microbiota, the effects of backslopping carried out under different temperature regimes (25 and 30 °C), pH (3.70 and 4.00) and inoculation rates (5, 10 and 15%) on lactic acid bacteria (LAB) diversity were determined in tarhana dough. LAB and Total Aerobic Mesophilic Bacteria (TAMB) numbers increased in all tarhana dough samples subjected to backslopping. Temperature and pH significantly affected the microbiological diversity of tarhana whereas the different inoculation rates did not. Tarhana dough showed complex tarhana microbiota following backslopping at pH 4.00 independently on the temperature applied. When backslopping was carried out at pH 3.70 and 25 °C, tarhana microbiota stabilized and became steady after several cycles. The LAB species found in all dough samples after the final backslopping were Lactobacillus plantarum, Lactobacillus alimentarius and Lactobacillus brevis which were able to carry out the fermentation in all conditions tested. In order to obtain a stable presence of LAB populations at industrial level for tarhana production, this work showed that backslopping is recommended at pH 3.70 and 25 °C with any inoculation ratios.

Microbial Diversity Associated with Gwell, a Traditional French Mesophilic Fermented Milk Inoculated with a Natural Starter.

Gwell is a traditional mesophilic fermented milk from the Brittany region of France. The fermentation process is based on a back-slopping method. The starter is made from a portion of the previous Gwell production, so that Gwell is both the starter and final product for consumption. In a participatory research framework involving 13 producers, Gwell was characterized from both the sensory and microbial points of view and was defined by its tangy taste and smooth and dense texture. The microbial community of typical Gwell samples was studied using both culture-dependent and culture-independent approaches. Lactococcus lactis was systematically identified in Gwell, being represented by both subspecies cremoris and lactis biovar diacetylactis which were always associated. Geotrichum candidum was also found in all the samples. The microbial composition was confirmed by 16S and ITS2 metabarcoding analysis. We were able to reconstruct the history of Gwell exchanges between producers, and thus obtained the genealogy of the samples we analyzed. The samples clustered in two groups which were also differentiated by their microbial composition, and notably by the presence or absence of yeasts identified as Kazachstania servazii and Streptococcus species.

Influence of the production technology on kefir characteristics: Evaluation of microbiological aspects and profiling of phosphopeptides by LC-ESI-QTOF-MS/MS.

The influence of production technology, namely, temperature, pH and 2-step fermentation (back-slopping approach), on the microbiological characteristics and on the phosphopeptide profile of kefir obtained with kefir grains was investigated. The growth of yeasts, lactic acid bacteria (LAB) and acetic acetic bacteria (AAB) in both grains and kefir was affected by the incubation temperature and by the use of back-slopping. In particular, at 25 °C the microbiota of kefir grains was mainly composed by LAB and yeasts, while at 18 °C yeasts represented the dominant group in kefir. Back-slopping at 25 °C determined a significant increase of AAB. A comprehensive characterization of potentially bioactive peptides, including caseino-phosphopeptides (CPPs), was performed, for the first time, in kefir obtained with kefir grains, using preliminary enrichment on hydroxyapatite followed by dephosphorylation and analysis by Liquid Chromatography-ElectroSpray Ionization-Quadrupole-Time of Flight-tandem mass spectrometry (LC-ESI-QTOF-MS/MS). As a result, seventy-three phosphopeptides, mostly arising from caseins (79% ß-casein, 8% αs1-casein and 9% αs2-casein) and all including from three to five serine residues in their sequences, were identified. Seventy-one of them showed the typical motif "SerP-SerP-SerP-Glu-Glu", which is crucial for the ability of caseins to bind to minerals. Several peptides were observed, for the first time, from the 1-40 region of ß-casein. As for the effect of production technology, phosphopeptide profiles of kefirs obtained at 25 °C and 18 °C were very similar, whereas kefir produced under acidic conditions showed a predominance of smaller peptides, suggesting a higher level of proteolysis. Conversely, kefir obtained through back-slopping at 25 °C contained longer peptides, thus indicating a lower proteolytic activity and a poor reproducibility in the kefir phosphopeptide profile occurring when grains are reused.

Diversity of lactic acid bacteria in dadih produced by either back-slopping or spontaneous fermentation from two different regions of West Sumatra, Indonesia.

AIM: Dadih samples from two different origins (Kamang and Gadut in West Sumatra) manufactured with different methods (back-slopping or spontaneous fermentation) were evaluated for the diversity of lactic acid bacteria (LAB). MATERIALS AND METHODS: Four dadih samples manufactured with two different fermentation methods were obtained from Kamang and Gadut regions. Both genotypic and phenotypic characteristic (16S rRNA partial gene sequence analysis and carbohydrate fermentation profile) were used to analyze the diversity of dadih LAB population. RESULTS: This study showed that LAB count in back-slopping fermented dadih was one log cycle higher than spontaneous fermented dadih. LAB isolates from the two regions were divided into three genera, namely Lactococcus, Lactobacillus, and Pediococcus. Sequencing results showed that 41.6% (five isolates) were identified as Lactococcus lactis ssp. lactis, 25% (three isolates) were identified as Lactobacillus plantarum ssp. plantarum, 16.6% (two isolates) were identified as L. lactis ssp. cremoris, and 8.3% (one isolate each) were identified as Pediococcus pentosaceus and Lactobacillus pentosus. CONCLUSION: Five species were determined in back-slopping fermented dadih, i.e., L. lactis ssp. lactis, L. lactis ssp. cremoris, L. plantarum ssp. plantarum, L. pentosus, and P. pentosaceus. On the other hand, spontaneous fermented dadih only contained three different species, namely L. lactis ssp. lactis, L. lactis ssp. cremoris, and L. plantarum ssp. plantarum. This research showed that back-slopping fermentation offers greater abundance and diversity compared to spontaneous fermentation in dadih.

Microbiota composition of dadih - a traditional fermented buffalo milk of West Sumatra.

Dadih is an Indonesian traditional spontaneously fermented buffalo milk, produced in West-Sumatra, which is nutritious and has health benefits. The mechanism of action behind the health benefits is largely unknown, but several probiotic strains have been isolated from dadih, which may contribute to its health properties. To identify the composition of its microbiota, two artisanal dadih samples (n = 8) were collected from four producers. The raw buffalo milk used for fermentation was either pasteurized (n = 4) or not (n = 4), and back-slopping was used as a starter-culture (n = 5) or not (n = 3). DNA was extracted from each sample in duplicate and the microbiota composition was determined by 16S-rRNA-gene amplicon-sequencing of the V3-V4 region. PCoA analysis showed clear separation of the samples by producer, but no separation due to pasteurization or use of back-slopping. Lactococcus (52-83%) predominated in all samples, followed by Klebsiella (5-26%), and Lactobacillaceae, Bifidobacterium (particularly high (c. 18%) in the non-pasteurized, back-slopped product from Palupuh), Streptococcus and Leuconostoc. Back-slopping practice correlated significantly with higher abundance of Lactobacillaceae, Pediococcus, species of the order Burkholderiales, and Serratia, but with lower abundance of several other Enterobacteriaceae (including Klebsiella), Streptococcaceae, Staphylococcus and Brachybacterium. Pasteurization was not significantly correlated with the presence of certain members of the final microbiota. Taken together, fermentation results differ significantly from producer to producer and back-slopping practice would be advisable. SIGNIFICANCE AND IMPACT OF THE STUDY: Using state-of-the-art methods we determined the microbiota composition of dadih, an artisanal, traditional fermented buffalo milk of West Sumatra with health benefits. We show that the artisanal practice leaves room for standardization and optimization with respect to the presence of potential pathogenic species in the final product. The Dadih Initiative in Indonesia aims to expand production of this health promoting product, and the findings help to determine important steps for potential food safety issues and good-manufacturing-practices to obtain a safe, nutritious and healthy traditional yoghurt-like functional food.

Effects of selected lactobacilli on the functional properties and stability of gluten-free sourdough bread.

The aim of this investigation was to determine the influence of seven different Lactobacillus spp. (Lb.) strains compared with a commercial starter culture (CS) on the functional properties of gluten-free (GF) sourdough-breads. The sourdough stability of selected strains was also evaluated upon back-slopping. Results showed that the bread properties were greatly affected by the Lb. strains. Millet breads achieved lower specific volumes (1.80-2.19 cm3/g), higher crumb firmness (19.01-42.19 N) and lower relative elasticities (21.5-43.4%) than buckwheat breads. Compared with the CS, Lactobacillus pentosus and Lb. hammesii positively influenced the crumb firmness of buckwheat and millet breads, respectively, while Lb. paralimentarius enhanced this property in both breads. Only one of the two Lactobacillus sanfranciscencis strains was able to improve all functional properties in both GF breads. Back-slopping of the sourdoughs revealed stable properties in case of buckwheat, while maturity of the millet sourdough could not be reached. These observations were supported by the microbial count, metabolite production and carbohydrate consumption. Mature sourdough significantly improved the crumb firmness and porosity of the GF breads. These results highlighted the importance of selecting the appropriate lactic acid bacteria strains, to maximize the quality of GF bread.

Direct Application of Rep-PCR on Type I Sourdough Matrix to Monitor the Dominance and Persistence of a Lactobacillus plantarum Starter Throughout Back-Slopping.

This study describes the optimization and application of repetitive element-PCR (rep-PCR) technique directly on microbial DNA extracted from type I sourdoughs for fast monitoring of a Lb. plantarum starter strain (P1FMC) throughout daily back-slopping. The challenge was to follow and study the performance of a starter culture directly in sourdoughs without cultivation on selective media. The extraction of good quality microbial DNA suitable for amplification from a complex matrix such as dough was the first target. In addition, the objective to obtain a clear rep-PCR profile referable to a specific starter strain among a microbial community was pursued. Co-inoculum trials, in flour matrix, with Lb. plantarum P1FMC and L. lactis LC71 strains and, subsequently, type I sourdough back-slopping trials were performed. The rep-PCR amplification profiles obtained were clearly referable to that of Lb. plantarum P1FMC starter in both co-inoculum trials (also when it was present with one order of magnitude less with respect to L. lactis LC71) and back-slopping trials where it dominated the fermentation process with loads of 108 cfu g-1 and prevailed on the autochthonous microbiota. Thus, the approach proposed in this paper could be considered a methodological advancement, based on a culture-independent one-step rep-PCR, suitable for fast monitoring of starter performance.

Microbial Ecology and Process Technology of Sourdough Fermentation.

From a microbiological perspective, sourdough is to be considered as a specific and stressful ecosystem, harboring yeasts and lactic acid bacteria (LAB), that is used for the production of baked goods. With respect to the metabolic impact of the sourdough microbiota, acidification (LAB), flavor formation (LAB and yeasts), and leavening (yeasts and heterofermentative LAB species) are most noticeable. Three distinct types of sourdough fermentation processes can be discerned based on the inocula applied, namely backslopped ones (type 1), those initiated with starter cultures (type 2), and those initiated with a starter culture followed by backslopping (type 3). A sourdough-characteristic LAB species is Lactobacillus sanfranciscensis. A sourdough-characteristic yeast species is Candida humilis. Although it has been suggested that the microbiota of a specific sourdough may be influenced by its geographical origin, region specificity often seems to be an artefact resulting from interpretation of the research data, as those are dependent on sampling, isolation, and identification procedures. It is however clear that sourdough-adapted microorganisms are able to withstand stress conditions encountered during their growth. Based on the technological setup, type 0 (predoughs), type I (artisan bakery firm sourdoughs), type II (industrial liquid sourdoughs), and type III sourdoughs (industrial dried sourdoughs) can be distinguished. The production of all sourdoughs, independent of their classification, depends on several intrinsic and extrinsic factors. Both the flour (type, quality status, etc.) and the process parameters (fermentation temperature, pH and pH evolution, dough yield, water activity, oxygen tension, backslopping procedure and fermentation duration, etc.) determine the dynamics and outcome of (backslopped) sourdough fermentation processes.