Functional and technological characterization of lactic acid bacteria isolated from Turkish dry-fermented sausage (sucuk).

In this study, 10 lactic acid bacteria were isolated from Turkish fermented sausage (sucuk) and identified as 5 Lactobacillus plantarum, 1 Pediococcus acidilactici, 1 Weissella hellenica, 1 Lactobacillus pentosus, and 2 Lactobacillus sakei. PCR screening of genes encoding plantaricin A and pediocin showed the presence of plantaricin A gene in 9 and pediocin gene in 3 of strains. All isolates showed antibacterial and antifungal effect on most of the tested microorganisms. gad gene, encoding glutamic acid decarboxylase enzyme, was detected in all isolates except Weisella hellenica KS-24. Eight of isolates were determined as gamma-amino butyric acid (GABA) producer in the presence of 53 mM mono sodium glutamate (MSG) by HPLC and TLC analysis. DPPH scavenging activity was observed for all isolates. Additionally, isolates were able to produce exopolysaccharide in the presence of sucrose. The best exopolysaccharide (EPS) production was achieved with L. plantarum KS-11 and L. pentosus KS-27. As a result, this study characterized some techno-functional properties of LAB isolates from sucuk. It was concluded that the isolates studied have the potential to be used in obtaining functional products in meat industry, as well as strain selection may be effective in providing the desired properties in the product.

Molecular tools for the analysis of the microbiota involved in malolactic fermentation: from microbial diversity to selection of lactic acid bacteria of enological interest.

Winemaking is a complex process involving two successive fermentations: alcoholic fermentation, by yeasts, and malolactic fermentation (MLF), by lactic acid bacteria (LAB). During MLF, LAB can contribute positively to wine flavor through decarboxylation of malic acid with acidity reduction and other numerous enzymatic reactions. However, some microorganisms can have a negative impact on the quality of the wine through processes such as biogenic amine production. For these reasons, monitoring the bacterial community profiles during MLF can predict and control the quality of the final product. In addition, the selection of LAB from a wine-producing area is necessary for the formulation of native malolactic starter cultures well adapted to local winemaking practices and able to enhance the regional wine typicality. In this sense, molecular biology techniques are fundamental tools to decipher the native microbiome involved in MLF and to select bacterial strains with potential to function as starter cultures, given their enological and technological characteristics. In this context, this work reviews the different molecular tools (both culture-dependent and -independent) that can be applied to the study of MLF, either in bacterial isolates or in the microbial community of wine, and of its dynamics during the process.

In vitro and genetic screening of probiotic properties of lactic acid bacteria isolated from naturally fermented cow-milk and yak-milk products of Sikkim, India.

A total of 272 isolates of lactic acid bacteria (LAB) were isolated from 22 samples of naturally fermented milk products of Sikkim in India viz. dahi, soft-variety chhurpi, hard-variety chhurpi, mohi and philu, out of which, 68 LAB isolates were randomly grouped on the basis of phenotypic characteristics, and were identified by 16S rRNA gene sequence analysis. Leuconostoc mesenteroides was the most dominant genus, followed by Leuc. mesenteroides subsp. jonggajibkimchii, Lactococcus lactis subsp. cremoris, Lc. lactis, Lc. lactis subsp. hordniae, Lc. lactis subsp. tructae, Enterococcus faecalis, E. italicus and E. pseudoavium. LAB strains were tested for probiotics attributes by in vitro and genetic screening, based on marker genes. LAB strains showed tolerance to pH 3.0, bile salt, resistance to lysozyme and ß-galactosidase activity. Enterococcus faecalis YS4-11 and YS4-14 and Lactococcus lactis subsp. cremoris SC3 showed more than 85% of hydrophobicity. Genes clp L and tdc encoding for low pH tolerance, agu A and Ir1516 encoding for bile tolerance, LBA1446 gene encoding for BSH activity, map A, apf, mub 1 and msa encoding for mucosal binding property were detected. Gene mesY for bacteriocin production was detected only in Leuconostoc spp. Based on the in vitro and genetic screening of probiotic attributes, Leuc. mesenteroides; Leuc. mesenteroides subsp. jonggajibkimchii and Lc. lactis subsp. cremoris were tentatively selected for possible probiotic candidates.

Mechanistic Insight into Yeast Bloom in a Lactic Acid Bacteria Relaying-Community in the Start of Sourdough Microbiota Evolution.

The spontaneous microbiota of wheat sourdough, often comprising one yeast species and several lactic acid bacteria (LAB) species, evolves over repeated fermentation cycles, which bakers call backslopping. The final product quality largely depends on the microbiota functions, but these fluctuate sometimes during the initial months of fermentation cycles due to microbiota evolution in which three phases of LAB relay occur. In this study, the understanding of yeast-LAB interactions in the start of the evolution of the microbiota was deepened by exploring the timing and trigger interactions when sourdough yeast entered a preestablished LAB-relaying community. Monitoring of 32 cycles of evolution of 6 batches of spontaneous microbiota in wheat sourdoughs revealed that sourdough yeasts affected the LAB community when the 2nd- or 3rd-relaying types of LAB genera emerged. In in vitro pairwise cocultures, all 12 LAB strains containing the 3 LAB-relaying types arrested the growth of a Saccharomyces cerevisiae strain, a frequently found species in sourdoughs, to various extents by sugar-related interactions. These findings suggest competition due to different affinities of each LAB and a S. cerevisiae strain for each sugar. In particular, maltose was the driver of S. cerevisiae growth in all pairwise cocultures. The functional prediction of sugar metabolism in sourdough LAB communities showed a positive correlation between maltose degradation and the yeast population. Our results suggest that maltose-related interactions are key factors that enable yeasts to enter and then settle in the LAB-relaying community during the initial part of evolution of spontaneous sourdough microbiota. IMPORTANCE Unpredictable evolution of spontaneous sourdough microbiota sometimes prevents bakers from making special-quality products because the unstable microbiota causes the product quality to fluctuate. Elucidation of the evolutionary mechanisms of the sourdough community, comprising yeast and lactic acid bacteria (LAB), is fundamental to control fermentation performance. This study investigated the mechanisms by which sourdough yeasts entered and settled in a bacterial community in which a three-phase relay of LAB occurred. Our results showed that all three layers of LAB restricted the cohabiting yeast population by competing for the sugar sources, particularly maltose. During the initial evolution of spontaneous sourdough microbiota, yeasts tended to grow synchronously with the progression of the lactic acid bacterial relay, which was predictably associated with changes in the maltose degradation functions in the bacterial community. Further study of ≥3 species' interactions while considering yeast diversity can uncover additional interaction mechanisms driving the initial evolution of sourdough microbiota.

Technological, functional and safety properties of lactobacilli isolates from soft wheat sourdough and their potential use as antimould cultures.

Bakery products are a common medium for fungal growth due to their high-water activity and nutrients availability. The application of lactic acid bacteria (LAB) isolated from wheat bran or other cereals has shown great potential in controlling the growth of spoilage fungi, guarantee quality and prolong the shelf life of bakery products. This study outlines the antifungal, technological, functional and safety properties of autochthonous LAB microbiota isolated from type 0 soft wheat sourdough fermentation. Antifungal activity of 77 LAB belonging to Lactiplantibacillus plantarum and Lacticaseibacillus casei species isolated from spontaneous sourdough fermentation was tested in vitro against 16 spoilage fungi. Our findings demonstrated that the antifungal activity, enzymatic and safety properties of LAB isolates vary strain-dependently. Four LAB isolates (Lp. plantarum A16, A25, B11, and B15) showed the best traits, in particular strong antifungal activity and good capabilities to produce exopolysaccharides from different carbon sources in vitro. Care should be taken when using Lp. plantarum A310 and B18 and Lc. casei A23, as starter cultures, since these isolates exhibited a multiple antibiotic-resistance. Here we showed the promising potential of different LAB isolates as bio-preservative agents and to provide new insights regarding their prospective use as starter cultures to guarantee safety and palatability.

High throughput screening of technological and biopreservation traits of a large set of wild lactic acid bacteria from Brazilian artisanal cheeses.

This study aimed to evaluate technological (acidification, proteolysis, lipolysis, resistance to low pH, NaCl, and bile salts) and biopreservation (antimicrobial activity against foodborne pathogens) features of 1002 LAB by high throughput screening (HTS) methods. The LAB was isolated from 11 types of Brazilian artisanal cheeses (BAC) marketed in the main 5 producing regions. Remarkable intra-species variability in acidification rates have been found, which was most pronounced between isolates from Mina's artisanal cheeses, Caipira and Coalho cheeses. Lacticaseibacillus paracasei and Levilactobacillus brevis showed the fastest acidification rate; however, all isolates showed slower acidification rates than a lactococcal control strain (4.3 × lower). When testing inhibitory effects, > 75% of LAB isolates could inhibit the growth of Staphylococcus aureus ATCC 19095 and Listeria monocytogenes ATCC 7644. Two of these isolates, identified as Lactiplantibacillus plantarum and Lentilactobacillus buchneri, the sterile and neutral supernatants alone, were sufficient to inhibit L. monocytogenes growth. Principal component analysis (PCA) allowed the identification of functional groups based on proteolytic and lipolytic activity, osmotic stress resistance, and inhibition of L. monocytogenes. The type of cheese the isolates were recovered from influenced properties such as anti-listerial compounds and lipolytic enzyme production. The use of HTS and multivariate statistics allowed insights into a diverse set of LAB technological and biopreservation properties. These findings allow a profound knowledge of the heterogeneity of a large set of isolates, which can be further used to design starter cultures with varied and combined properties, such as biopreservation and technological features. Besides that, HTS makes it possible to analyze a vast panel of LAB strains, reducing costs and time within laboratory analysis, while avoiding the loss of information once all LAB are tested at the same time (differently from the traditional labor-intensive approach, in which a few numbers of strains is tested per time).

Dairy associations for the targeted control of opportunistic Candida.

Antifungal and antibacterial activities of twenty-six combinations of lactic acid bacteria, propionibacteria, acetic acid bacteria and dairy yeasts inoculated in whey and milk were investigated. Associations including acetic acid bacteria were shown to suppress growth of the opportunistic yeast Candida albicans in well-diffusion assays. The protective effect of milk fermented with the two most promising consortia was confirmed in Caco-2 cell culture infected with C. albicans. Indeed, these fermented milks, after heat-treatment or not, suppressed lactate dehydrogenase release after 48 h while significant increase in LDH release was observed in the positive control (C. albicans alone) and with fermented milk obtained using commercial yogurt starter cultures. The analysis of volatile compounds in the cell-free supernatant using solid phase microextraction (SPME) coupled to gas chromatography-mass spectrometry (GC-MS) showed accumulation of significant amount of acetic acid by the consortium composed of Lactobacillus delbrueckii 5, Lactobacillus gallinarum 1, Lentilactobacillus parabuchneri 3, Lacticaseibacillus paracasei 33-4, Acetobacter syzygii 2 and Kluyveromyces marxianus 19, which corresponded to the zone of partial inhibition of C. albicans growth during well-diffusion assays. Interestingly, another part of anti-Candida activity, yielding small and transparent inhibition zones, was linked with the consortium cell fraction. This study showed a correlation between anti-Candida activity and the presence of acetic acid bacteria in dairy associations as well as a significant effect of two dairy associations against C. albicans in a Caco-2 cell model. These two associations may be promising consortia for developing functional dairy products with antagonistic action against candidiasis agents.

Selection of lactic acid bacteria with promising probiotic aptitudes from fruit and ability to survive in different food matrices.

Lactic acid bacteria (LAB) are among the most prevalent microorganisms forming the autochthonous microbiota of fruit. This study aimed to select LAB isolates with probiotic aptitudes from apple, banana, grape, and orange through evaluation of in vitro safety, technological, and functional-related properties. The ability of most promising selected isolates to survive in commercial apple and orange juice, meat stew, vegetable puree, and UHT milk during 28 days of refrigeration storage was evaluated. Ninety-three isolates identified preliminarily as LAB were recovered from fruit and 66 of these isolates passed safety tests. Most of these isolates were pre-identified as belonging to Lactobacillus or Enterococcus genus based on MALDI-ToF MS profiling. These 66 isolates were categorized into three homogeneous groups based on a preliminary cluster analysis run with data from experiments to measure technological characteristics. Nine LAB isolates were selected as the most promising for probiotic use based on a principal component analysis run with data from experiments to measure probiotic-related properties. Four of these isolates were sensitive to different antibiotics and identified (16S-rRNA gene sequencing) as Lactobacillus brevis (recently reclassified as L. brevis) or Lactobacillus spp. These 4 selected isolates had high viable counts and high percentages of physiologically active cells in apple and orange juice, beef stew, vegetable puree, and UHT milk during refrigeration storage. The results showed that apple, banana, orange, and grape are potential sources of LAB with aptitudes to be exploited for a possible probiotic use and distinguished abilities to survive in different food matrices.

Design of an autochthonous starter culture using strains isolated from traditional Matsoni.

Artisanal products support the conservation of the indigenous biodiversity of food microbiomes, although they do not always comply to quality and hygienic requirements for the dairy industry. This study describes the development of an autochthonous starter culture to produce Matsoni, a traditional Georgian fermented milk. To this end, strains of lactic acid bacteria isolated from artisanal Matsoni samples were used to design a starter formulation reproducing the dominant microbial diversity, also preserving quality characteristics and ensuring the safety of the product. As a result, strains that represent the acidifying portion of the starter (Lactobacillus delbrueckii subsp. lactis, L. delbrueckii subsp. bulgaricus and Streptococcus thermophilus) were combined in different ratios and strain combinations, together with cultures of Lactobacillus rhamnosus that were chosen for their potential beneficial traits. The strain association acting better in milk cultures at laboratory scale was selected as starter culture for the production of Matsoni in pilot-scale industrial trials.

Isolation, identification and utilization of lactic acid bacteria from silage in a warm and humid climate area.

The study aimed to isolate and identify lactic acid bacteria (LAB) from silages and their application to improve the fermentation quality of alfalfa. Forty-nine LAB strains were isolated from silages, and two strains were screened for growth and acid production rates. Then two strains were selected for Physiological and morphological tests and 16S rRNA sequencing. They were Gram-positive and Catalase-negative and were able to grow at pH 3.5 and at 45 °C, were unable to grow different NaCl concentrations as 3.0% and 6.5%. Strain BDy3-10 was identified as Lactobacillus rhamnosus, while TSy1-3 was identified as L. buchneri. The selected strains were evaluated on fermentation of alfalfa silage. The highest crude protein content occurred in the BDy3-10 treatment group. The contents of neutral detergent fiber and acid detergent fiber in the TSy1-3 treatment were significantly lower than other treatment (P < 0.05). Compared to the control treatment, inoculation treatments deceased pH during ensiling (P < 0.001) and provided the most increased lactic acid content after ensiling for 10 days (P < 0.001). The acetic acid contents of all the inoculation groups were significantly increased (P < 0.001) during ensiling, and were lower than that of control group (P < 0.001). So, the TSy1-3 treatment most effectively improved the fermentation quality of alfalfa silage in warm and humid climate area.

Technological characterization of indigenous lactic acid bacteria from Moroccan camel milk for their potential use as starter or adjunct culture.

This study aimed to isolate lactic acid bacteria (LABs) of technological interest from Moroccan camel milk and select starter or adjunct culture for dairy product manufacturing. The phenotypic and biochemical identification of 47 isolates revealed the existence of ten Lactococcus lactis, eleven Lactobacillus plantarum, three Lactobacillus brevis, two Lactobacillus paracasei, eleven Enterococcus spp., seven Lactococcus spp. and two Lactobacillus spp. Our strains showed a fast acidifying ability (ΔpH ranged between 0.69 ± 0.01 and 1.22 ± 0.05 after 6 h), high proteolytic and autolytic activities (1.93 ± 0.02 to 9.9 ± 0.022 mM glycine and 15.21 ± 2.21% to 83.24 ± 1% respectively), and an important lipolytic and free radical scavenging capacity. Furthermore, they were able to use citrate, to produce exopolysaccharide, and they exhibited antibacterial activity against Gram-negative and Gram-positive pathogenic bacteria and had no hemolytic activity. This study has shown that Moroccan camel milk represents a rich biotope of interesting LABs for dairy products industry.

Microbial profiles of Greek PDO cheeses assessed with amplicon metabarcoding.

Greece is a country possessing many cheese products granted with a PDO (Protected Designation of Origin) certificate, with high exporting activities. In this study, we analyzed six popular cheese PDO products purchased from different industries to assess their microbial communities using amplicon metabarcoding analysis. To this end, using Next Generation Sequencing technology, we sequenced the 16S rRNA gene and the ITS spacer for prokaryotes and fungi, respectively. Alpha diversity indices revealed higher bacterial species richness for some cheeses (Kopanisti, Batzos) and poor for others (Feta, Galotiri). Kopanisti, together with Kalathaki and Anevato, also presented increased species diversity concerning fungal populations. Results showed that lactic acid bacteria (LAB) prevailed the bacterial populations in all samples (Lactococcus, Lactobacillus, Streptococcus, Leuconostoc), whereas for fungi, members of the Saccharomycetaceae, Dipodascaceae and Debaryomycetaceae families prevailed the fungal populations. Several other genera were identified that make up each product's microbiome leading to the creation of the unique organoleptic attributes of Greek PDO cheeses. However, the identified species could not be directly linked to certain cheese types, assuming that starter and adjunct cultures, combined with the raw material used during production greatly impact the microbial communities in cheeses. Our data, produced for the first time for six Greek PDO cheeses, can be exploited in the process of creating a core microbial signature within each cheese type, supporting the Greek brand name and valorizing cheese products.

The lactic acid bacteria and yeast community of home-made sauerkraut from three provinces in Southwest China.

The aim of this study was to investigate the lactic acid bacteria (LAB) and yeast community from home-made sauerkraut collected from Southwest China through culture-dependent and culture-independent technology. Forty-eight samples of home-made sauerkraut were collected from households at three different locations in Southwest China. The pH, total acidity and salt contents among these fermented vegetables were 3.69 ± 0.42, 0.86 ± 0.43 g/100 ml, and 3.86 ± 2.55 g/100 ml, respectively. The number of lactic acid bacteria (LAB) and yeasts were 7.25 ± 1.05 log10 colony-forming units (CFU)/ml and 3.74 ± 1.01 log CFU/ml, respectively. A total of 182 LAB and 81 yeast isolates were identified. The dominant isolates were Lactobacillus plantarum, L. brevis, Pediococcus ethanolidurans, Pichia membranifaciens, P. fermentans and Kazachstania bulderi. Denaturing gradient gel electrophoresis (DGGE) showed that L. plantarum, uncultured Lactobacillus sp, P. ethanolidurans, and K. exigua were the predominant microflora. Our studies demonstrated that the DGGE technique combined with a culture-dependent method is very effective for studying the LAB and yeast community in Chinese traditional fermentation vegetables. The results will give us an understanding of LAB and yeast community of Chinese sauerkraut and improve the knowledge of LAB and yeast community of Chinese sauerkraut.

Comparison of the effects of acetic acid bacteria and lactic acid bacteria on the microbial diversity of and the functional pathways in dough as revealed by high-throughput metagenomics sequencing.

Knowledge of the effects of various strains of acetic acid bacteria (AAB) on sourdough remains limited. In this study, the diversity of microbial taxa in sourdoughs fermented by different starters was assessed and their functional capacity was evaluated via high-throughput metagenomics sequencing. Results showed that Erwinia (29.43%), Pantoea (45.89%), and Enterobacter (9.16%) were predominant in the blank CK treatment. Lactobacillus (91.40%), Saccharomyces (6.13%), as well as the AAB genus Acetobacter (0.61%) were the dominant microbial genera in the sourdoughs started by yeast and a strain of lactic acid bacteria (YL treatment). By contrast, the dominant genera in the sourdoughs started by yeasts and various LAB and AAB strains (YLA treatment) were Komagataeibacter (0.39%) except for the inoculated Lactobacillus (68.37%), Acetobacter (20.17%), and Saccharomyces (8.31%) species. Functional prediction of these changes in microbial community and diversity revealed that various metabolism-related pathways, including alanine, aspartate, and glutamate metabolism (21.95%), as well as amino acid biosynthesis (19.14%), were predominant in the sourdoughs started by yeast and an AAB strain (YA treatment). Moreover, arginine biosynthesis (11.65%) were the dominant pathways in the YL treatment. The fermented dough added with sourdoughs started with yeast + AAB and yeast + AAB + LAB strains had substantially higher contents (more than 48.58% in total) of essential amino acids than the dough added with sourdoughs started with yeast + LAB strain. These results demonstrated that amino acid biosynthesis has a beneficial effect on sourdoughs inoculated with an AAB strain.

Identification and characterisation of the lactic acid bacteria associated with the traditional fermentation of dairy fermented product.

The aim of this research was to identify the key lactic acid bacteria associated with the fermentation of dairy traditional fermented products for developing starter cultures for controlled fermentation. A total of 100 lactic acid bacteria (LAB) were isolated from dairy traditional fermented products. Samples were obtained from eight producers in the South East of Nigeria. Isolates were identified by phenotypic and genotypic techniques including rep-PCR genotyping and sequencing of the 16S rRNA, pheS and rpoA genes. Isolates were characterised for antimicrobial activity against foodborne pathogens, exopolysaccharide (EPS) production and survival at low pH and in the presence of bile salts. All isolates clustered into 11 distinct rep-PCR groups and were identified as Lactobacillus fermentum (40%), Lactobacillus delbrueckii (23%), Streptococcus thermophilus (22%), Streptococcus infantarius (10%), Lactobacillus senioris (2%), Leuconostoc pseudomesenteriodes (2%) and Enterococcus thailandicus (1%). Lactobacillus fermentum showed a broad spectrum antimicrobial activity and survival at low pH, while Lactobacillus delbrueckii was able to tolerate low pH and produce EPS. All isolates survived in vitro exposure to 1% (w/v) bile salts over a 3-h period. L. fermentum, L. delbrueckii and S. thermophilus could be used to simulate the fermentation of dairy traditional fermented products.

In vitro characterization of chicken gut bacterial isolates for probiotic potentials.

Probiotics often play an important role in improving gut health in chickens through multiple mechanisms, including enhancement of tight junctions, nutrient acquisition, niche colonization, or coaggregation with enteric pathogens. The objective of this study was to characterize lactic acid bacteria (LAB) isolated from the gut of healthy broiler chickens for a number of phenotypes that might be indicative of good probiotic potentials. A total 40 bacterial isolates were isolated from 3-week-old chickens using Man, Rogosa and Sharpe (MRS) agar plates. The bacterial isolates were evaluated in vitro for motility, autoaggregation, pathogen inhibition, pH of overnight culture, growth on different agar plates, and their impact on gut integrity. Selected isolates were genotyped by sequencing the 16S-23S rRNA gene intergenic region. Based on the phenotype and genotype, we identified 20 potential probiotic (PP) isolates that belong to LAB. Multivariate analysis showed that PP isolates were positively correlated with parameters such as growth on MRS agar plate (pH 5.5), pathogen inhibition, and autoaggregation. However, growth on MacConkey agar plates, supernatant pH, motility, and transepithelial electrical resistance were negatively correlated with the PP isolates. Furthermore, in vivo study needs to be performed for evaluation of the utility of these probiotic candidates in poultry production.

Unique niche-specific adaptation of fructophilic lactic acid bacteria and proposal of three Apilactobacillus species as novel members of the group.

BACKGROUND: Fructophilic lactic acid bacteria (FLAB) found in D-fructose rich niches prefer D-fructose over D-glucose as a growth substrate. They need electron acceptors for growth on D-glucose. The organisms share carbohydrate metabolic properties. Fructobacillus spp., Apilactobacillus kunkeei, and Apilactobacillus apinorum are members of this unique group. Here we studied the fructophilic characteristics of recently described species Apilactobacillus micheneri, Apilactobacillus quenuiae, and Apilactobacillus timberlakei. RESULTS: The three species prefer D-fructose over D-glucose and only metabolize D-glucose in the presence of electron acceptors. The genomic characteristics of the three species, i.e. small genomes and thus a low number of coding DNA sequences, few genes involved in carbohydrate transport and metabolism, and partial deletion of adhE gene, are characteristic of FLAB. The three species thus are novel members of FLAB. Reduction of genes involved in carbohydrate transport and metabolism in accordance with reduction of genome size were the common characteristics of the family Lactobacillaceae, but FLAB markedly reduced the gene numbers more than other species in the family. Pan-genome analysis of genes involved in metabolism displayed a lack of specific carbohydrate metabolic pathways in FLAB, leading to a unique cluster separation. CONCLUSIONS: The present study expanded FLAB group. Fructose-rich environments have induced similar evolution in phylogenetically distant FLAB species. These are examples of convergent evolution of LAB.

In vitro properties of potential probiotic lactic acid bacteria originating from Ghanaian indigenous fermented milk products.

Fermented milk products are a major source of health-promoting microorganisms known as probiotics. To characterize the probiotic properties of lactic acid bacteria isolated from Ghanaian traditionally fermented milk, thirty (30) isolates comprising Enterococcus faecium (1), Lactobacillus fermentum (14), Lb. plantarum (2) and Pediococcus acidilactici (13) identified by 16S rRNA gene sequencing, were tested for survival at low pH (2.5) and bile salts (0.3% (w/v)), hydrophobicity, co-aggregation, auto-aggregation and antimicrobial activities against selected pathogens. Safety of potential probiotic bacteria was assessed by hemolytic activity on blood agar and susceptibility to nine different antibiotics. Majority (90%) of the strains showed survival rates above 80% at pH (2.5) and in bile salts (0.3% (w/v)). Hydrophobicity ranged from 5 to 61% while cell auto-aggregation ranged from 41 to 80% after 24 h. Co-aggregation with E. coli (3.7-43.9%) and S. Typhimurium (1.3-49.5%) were similar for the LAB strains at 24 h. Cell- free supernatants of all LAB strains inhibited E. coli while S. Typhimurium was not sensitive to cell-free supernatants of five Pd. acidilactici strains: OS24h20, OS18h3, OY9h19, OS9h8 and 24NL38. None of the LAB strains showed ß-hemolysis but 38% of strains showed α-hemolysis. Susceptibilities to antibiotics were strain-specific; only four strains, two Lb. fermentum and two Pd. acidilactici were susceptible to all nine antibiotics tested. Based on high survival rates in bile salts, low pH and generally good hydrophobicity, auto-aggregation, co-aggregation and inhibitory activities, 15 out of 30 strains tested were considered qualified candidates for development of probiotic cultures for fermented milk products in sub-Saharan Africa.

Multifunctional properties and safety evaluation of lactic acid bacteria and yeasts associated with fermented cereal doughs.

Spontaneous cereal fermentations involve diverse lactic acid bacteria (LAB) and yeasts which may include multifunctional and safe or unsafe strains. This study assessed acidification ability, safety, antifungal activity and free amino acids release ability of LAB and yeasts previously isolated from spontaneously fermented cereal doughs in Benin. Fourteen LAB and thirteen yeast strains were studied in liquid media and/or in a model cereal dough prepared in laboratory conditions. Antifungal activity was assessed against Candida glabrata in liquid medium. Amino acids were determined by pre-column derivatization and separation with reversed-phase HPLC. Antimicrobial susceptibility was analysed by minimum inhibitory concentration determination. The acidification ability was higher for LAB compared to yeast strains. All LAB strains retarded the growth of C. glabrata Cg1 with the highest inhibition recorded for Weissella confusa Wc1 and Wc2. The highest free amino acid content was found in the doughs fermented with Pichia kudriavzevii Pk2 and Pk3. All the LAB strains were susceptible to ampicillin, chloramphenicol, erythromycin, but displayed phenotypic resistance to kanamycin, streptomycin and tetracycline. Positive PCR amplicon of resistance genes were detected in the following cases: 2 LAB strains were positive for kanamycin (aph(3)III), 5 strains were positive for streptomycin (aadA and/or strA and/or strB) and 3 strains were positive for tetracycline (tet (L) and/or tet (M)). For yeasts, most of the P. kudriavzevii strains were resistant to amphotericin B, fluconazole and itraconazole opposite to K. marxianus and Saccharomyces cerevisiae strains which were susceptible. The results obtained are valuable for selecting safe and multifunctional strains for cereal fermentation in West Africa.

Probiotic properties of lactic acid bacteria isolated from traditionally prepared dry starters of the Eastern Himalayas.

The Himalayan people prepare dry and oval to round-shaped starter cultures to ferment cereals into mild-alcoholic beverages, which contain lactic acid bacteria (LAB) as one of the essential microbiota. There is no report on probiotic characters of LAB isolated from dry starters. Hence, we screened the probiotic and some functional properties of 37 LAB strains isolated from dry starters of the Eastern Himalayas viz. marcha, phab, paa, pee and phut. About 38% of the LAB strains showed high survival rate (> 50%) at pH 3 and 0.3% bile salts. Enterococcus durans BPB21 and SMB7 showed the highest hydrophobicity percentage of 98%. E. durans DMB4 and SMB7 showed maximum cholesterol assimilation activity. About 65% of the LAB strains showed the ability to produce ß galactosidase. Majority of the strains showed phytase activity, whereas none of the strain showed amylase activity. About 86% of LAB strains showed an optimum tolerance of 10% ethanol concentration. Genetic screening of some probiotic and functional marker genes have also been analysed. The occurrence of clp L gene, agu A gene (survival of gastrointestinal tract conditions), apf, mub1 and map A gene (adhesion genes) was higher compared to other genes. The occurrence of bsh gene (bile salt tolerance) was detected in Pediococcus pentosaceus SMB13-1 and Enterococcus faecium BPB11. Gene ped B for pediocin with amplicon size of 375 bp was detected in E. durans DMB13 and Pediococcus acidilactici AKB3. Detection of nutritional marker gene rib A and fol P in some strains showed the potential ability to synthesize riboflavin and folic acid. LAB with probiotic and functional properties may be explored for food industry in future.