Cereal-based fermented foods as microbiota-directed products for improved child nutrition and health in sub-Saharan Africa.

Several strategies, programs and policies have long been developed and implemented to alleviate child malnutrition in sub-Saharan African countries. However, stunting and wasting still persist at an alarming rate, suggesting that alternative strategies are needed to induce faster progress toward the 2030 SDGs targets of reducing malnutrition. Gut microbiota-directed intervention is now being recognized as an unconventional powerful approach to mitigate malnutrition and improve overall child health. In an African setting, manufactured probiotic and synbiotic foods or supplements may not be successful owing to the non-affordability and high attachment of African populations to their food tradition. This review analyses the potential of indigenous fermented cereal-based products including porridges, doughs, beverages, bread- and yoghurt-like products, to be used as microbiota-directed foods for over 6 months children. The discussion includes relevant strategies to effectively enhance the beneficial effects of these products on gut microbiota composition for improved child health and nutrition in sub-Saharan Africa. Characterization of probiotic features and general safety of food processing in sub-Saharan Africa as well as randomized clinical studies are still lacking to fully ascertain the health effects and suitability of these fermented foods in preventing and treating child malnutrition and diarrhea.

The impact of botanical fermented foods on metabolic syndrome and type 2 diabetes: a systematic review of randomised controlled trials.

Our systematic review assessed the impact of botanical fermented food (BFF) consumption on glucose, lipid, anthropometric, inflammatory and gut microbiota parameters, in adults with metabolic syndrome (MetS), MetS components or type 2 diabetes mellitus (T2DM). Embase, MEDLINE, Cochrane CENTRAL and Google Scholar were searched with no language limits, from inception to 31 August 2022, for eligible randomised controlled trials (RCTs). Two independent reviewers screened 6873 abstracts and extracted relevant data. Risk of bias (ROB) was assessed using the Cochrane Collaboration's ROB2 tool. The final review included twenty-six RCTs, with thirty-one reports published between 2001 and 2022. Significant (p < 0·05) within-group and between-group changes in cardiometabolic outcome means were reported in twenty-three and nineteen studies, respectively. Gut microbiota composition was assessed in four studies, with two finding significant between-group differences. No significant difference between groups of any measured outcomes was observed in five studies. There were fourteen studies at low ROB; ten were of some concern; and two were at high ROB. In 73% of included studies, BFF consumption by participants with obesity, MetS or T2DM led to significant between-group improvements in discrete cardiometabolic outcomes, including fasting blood glucose, lipid profile, blood pressure, waist circumference, body fat percentage and C-reactive protein. BFF consumption increased the abundance of beneficial gut bacteria, such as Bifidobacterium and LAB, whilst reducing potential pathogens such as Bacteroides. To determine the clinical significance of BFFs as therapeutic dietary adjuncts, their safety, tolerability and affordability must be balanced with the limited power and magnitude of these preliminary findings.

Diversity and succession of contaminating yeasts in white-brined cheese during cold storage.

Contamination of white-brined cheeses (WBCs) with yeasts is of major concern in the dairy industry. This study aimed to identify yeast contaminants and characterize their succession in white-brined cheese during a shelf-life of 52 weeks. White-brined cheeses added herbs (WBC1) or sundried tomatoes (WBC2) were produced at a Danish dairy and incubated at 5 °C and 10 °C. An increase in yeast counts was observed for both products within the first 12-14 weeks of incubation and stabilized afterwards varying in a range of 4.19-7.08 log CFU/g. Interestingly, higher incubation temperature, especially in WBC2, led to lower yeast counts, concurrently with higher diversity of yeast species. Observed decrease in yeast counts was, most likely, due to negative interactions between yeast species leading to growth inhibition. In total, 469 yeast isolates from WBC1 and WBC2 were genotypically classified using the (GTG)5-rep-PCR technique. Out of them, 132 representative isolates were further identified by sequencing the D1/D2 domain of the 26 S rRNA gene. Predominant yeast species in WBCs were Candida zeylanoides and Debaryomyces hansenii, while Candida parapsilosis, Kazachstania bulderi, Kluyveromyces lactis, Pichia fermentans, Pichia kudriavzevii, Rhodotorula mucilaginosa, Torulaspora delbrueckii, and Wickerhamomyces anomalus were found in lower frequency. Heterogeneity of yeast species in WBC2 was generally larger compared to WBC1. This study indicated that, along with contamination levels, taxonomic heterogeneity of yeasts is an important factor influencing yeast cell counts, as well as product quality during storage.

Technologically relevant Bacillus species and microbial safety of West African traditional alkaline fermented seed condiments.

Fermented food condiments serve as a major source of nutrients to many homes in West Africa, especially among the rural poor who use these condiments as a cheap source of protein substitute for milk and other animal protein sources. Traditional fermented West African condiments are produced by spontaneous fermentation of legumes and protein-rich seeds of both cultivated and wild plant species. These fermented condiments are culturally accepted and widely produced in the West African sub-region, and rely on indigenous microbiota responsible for taste, texture, aroma development and the overall unique product characteristics. Detailed understanding of fermentation microbiota and their unique technological and functional properties are fundamental in developing products with enhanced quality and safety, as well as development of specific locally adapted starter cultures. Technologically relevant Bacillus spp., mainly Bacillus subtilis, are the predominant fermentative bacteria responsible for the natural fermentation of condiments across West Africa. Other species of Bacillus including B. amyloliquefaciens, B. licheniformis, B. pumilus, B. megaterium, B. sphaericus, B. cereus, B. badius and B. fusiformis are also frequently involved in the fermentation process. These bacterial species are responsible for flavor development, bio-conversion of complex food molecules, and production of antimicrobial compounds that impact shelf-life and safety, and in some instances, may confer host-beneficial health effects beyond basic nutrition. First, this review provides currently available information on the technologically relevant Bacillus species isolated from fermented food condiments in nine (9) West African countries. In addition, perspectives on harnessing the potentials of the technologically beneficial bacterial strains in fermented condiments in West Africa for enhanced food safety, quality and overall food security is presented.

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 potential of Saccharomyces cerevisiae and Kluyveromyces marxianus isolated from West African spontaneously fermented cereal and milk products.

The yeast species Saccharomyces cerevisiae and Kluyveromyces marxianus are associated with fermentation of West African indigenous foods. The aim of this study was to characterize potential probiotic properties of S. cerevisiae and K. marxianus isolates from the West African milk products lait caillé and nunu and a cereal-based product mawè. The strains (14 in total) were identified by 26S rRNA gene sequencing and characterized for survival at gastrointestinal stress (bile salts and low pH) and adhesion to Caco-2 intestinal epithelial cells. Selected yeast isolates were tested for their effect on the transepithelial electrical resistance (TEER), using the intestinal epithelial cell line Caco-2 and for maintenance of intracellular pH (pHi ) during perfusion with gastrointestinal pH (3.5 and 6.5). All tested yeasts were able to grow in bile salts in a strain-dependent manner, exhibiting a maximum specific growth rate (µmax ) of 0.58-1.50 h-1 . At pH 2.5, slow growth was observed for the isolates from mawè (µmax of 0.06-0.80 h-1 ), whereas growth of yeasts from other sources was mostly inhibited. Yeast adhesion to Caco-2 cells was strain specific and varied between 8.0% and 36.2%. Selected strains of S. cerevisiae and K. marxianus were able to maintain the pHi homeostasis at gastrointestinal pH and to increase TEER across the Caco-2 monolayers, indicating their potential to improve intestinal barrier functions. Based on overall results, strains of K. marxianus and S. cerevisiae from mawè exhibited the highest probiotic potential and might be recommended for further development as starter cultures in West African fermented products.

The Effects of NaCl and Temperature on Growth and Survival of Yeast Strains Isolated from Danish Cheese Brines.

Yeasts play an important role in cheese making, by contributing to microbial community establishment and improving flavor. This study aimed at investigating the impact of NaCl and temperature on growth and survival of 20 strains belonging to the yeast species Candida intermedia (2 strains), Debaryomyces hansenii (11), Kluyveromyces lactis (1), Papiliotrema flavescens (1), Rhodotorula glutinis (1), Sterigmatomyces halophilus (2) and Yamadazyma triangularis (2) isolated from Danish cheese brines. All yeasts could grow in Malt Yeast Glucose Peptone (MYGP) medium with low NaCl (≤ 4%, w/v) concentrations at 25 °C and 16 °C. Further, none of the strains, except for one strain of D. hansenii (KU-9), were able to grow under a condition mimicking cheese brine (MYGP with 23% (w/v) NaCl and 6.3 g/L lactate) at 25 °C, while all yeasts could grow at 16 °C, except for the two strains of C. intermedia. In the survival experiment, D. hansenii, S. halophilus and Y. triangularis survived in MYGP with 23% (w/v) NaCl throughout 13.5 days at 25 °C, with Y. triangularis and S. halophilus being the most NaCl tolerant, while the remaining yeasts survived for less than 7 days. These results enable the selection of relevant yeasts from cheese brines for potential use in the cheese industry.

Technological properties of indigenous Lactococcus lactis strains isolated from Lait caillé, a spontaneous fermented milk from Burkina Faso.

The experiments reported in this research paper aimed to determine the technological properties of indigenous Lactococcus lactis strains isolated from Lait caillé, a spontaneous fermented milk, from the perspective of starter culture development. Fermentations were conducted to determine the acidification patterns. The ropy character, growth in 0.04 g/ml NaCl and citrate metabolism were additionally tested. Furthermore, the rheological properties of samples from selected strains and the impact of cold storage were evaluated. Based on the rate of acidification, the indigenous strains were divided into 2 groups depending on their fermentation time, i.e. 10-13 h (fast acidifier), and up to 72 h (slow acidifier), respectively. The physiological tests suggested that most of these strains produced exopolysaccharides but none could ferment citrate. The flow properties of the samples inoculated by the fast acidifier strains showed a time-dependent shear thinning behaviour, while their viscoelastic properties corresponded structurally to those of weak gels. Cold storage decreased the viscosity and CFU counts for most of the indigenous strains tested. This study is a step towards the definition of starter cultures for African spontaneous fermented milks such as Lait caillé.

Identification of the predominant microbiota during production of lait caillé, a spontaneously fermented milk product made in Burkina Faso.

The spontaneously fermented curdled milk product from Burkina Faso, lait caillé is prepared by traditional processing from raw unpasteurised milk. The fermentation lasts 1-3 days. This study aims to identify the predominant microbiota involved in lait caillé fermentation from cow milk. A survey on lait caillé end-products from local markets showed pH ranges of 3.5 to 4.2. Counts of total lactic acid bacteria (LAB) were 7.8 ± 0.06 to 10.0 ± 0.03 log CFU/g and yeast counts were 5.3 ± 0.06 to 8.7 ± 0.01 log CFU/g, together with considerate amounts of Enterobacteriaceae < 3.00 to 8.4 ± 0.14 log CFU/g. Sampling throughout the entire fermentation of lait caillé was performed at a traditional house-hold production site. A drop in pH from 6.7 ± 0.01 at 0 h to 4.3 ± 0.08 in the end-product (59 h) was found. Total LAB counts increased to 8.6 ± 0.02 log CFU/g in the end-product, while yeast and Enterobacteriaceae counts reached 6.4 ± 0.11 and 6.7 ± 0.00 log CFU/g, respectively. LAB and yeasts isolated during the fermentation were clustered by (GTG)5 repetitive-PCR fingerprinting followed by 16S and 26S rRNA gene sequencing, respectively. Microbial successions were observed with Leuconostoc mesenteroides being the predominant LAB followed by Pediococcus pentosaceus and Weissella paramesenteroides at the onset, while Lactococcus lactis and Enterococcus spp. where the predominant LAB after 7 h of fermentation. During the first 18 h Candida parapsilosis was the dominant yeast species, while from 35 h to the end-product, Saccharomyces cerevisiae predominated. The microbial safety risk pointed out in this study, showed the need for implementation of good manufacturing practices including pasteurisation and use of well-defined starter cultures.

Modulation of gut microbiota from obese individuals by in vitro fermentation of citrus pectin in combination with Bifidobacterium longum BB-46.

This study aimed to evaluate the effects of three treatments, i.e., Bifidobacterium longum BB-46 (T1), B. longum BB-46 combined with the pectin (T2), and harsh extracted pectin from lemon (T3) on obesity-related microbiota using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME®). The effects of the treatments were assessed by the analysis of the intestinal microbial composition (using 16S rRNA gene amplicon sequencing) and the levels of short-chain fatty acids (SCFAs) and ammonium ions (NH4+). Treatments T2 and T3 stimulated members of the Ruminococcaceae and Succinivibrionaceae families, which were positively correlated with an increase in butyric and acetic acids. Proteolytic bacteria were reduced by the two treatments, concurrently with a decrease in NH4+. Treatment T1 stimulated the production of butyric acid in the simulated transverse and descending colon, reduction of NH4+ as well as the growth of genera Lactobacillus, Megamonas, and members of Lachnospiracea. The results indicate that both B. longum BB-46 and pectin can modulate the obesity-related microbiota; however, when the pectin is combined with B. longum BB-46, the predominant effect of the pectin can be observed. This study showed that the citric pectin is able to stimulate butyrate-producing bacteria as well as genera related with anti-inflammatory effects. However, prospective clinical studies are necessary to evaluate the anti/pro-obesogenic and inflammatory effects of this pectin for future prevention of obesity.

The effect of pectins on survival of probiotic Lactobacillus spp. in gastrointestinal juices is related to their structure and physical properties.

Pectins are plant polysaccharides used in food industry as gelling and stabilizing agents. This study investigated the ability of pectins to improve survival of probiotic species Lactobacillus fermentum PCC, L. reuteri RC-14, L. rhamnosus LGG and L. paracasei F-19 in simulated gastric solution in relationship to their structural and physical properties. Electrostatic interactions between pectins and bacteria were evaluated by the Zeta-potential approach. Bacterial survival was assessed by flow cytometry and plate counting. L. fermentum PCC and L. reuteri RC-14 were more resistant to gastric conditions; their survival rate was further improved in the presence of five out of ten tested pectins. Additionally, two of the pectins had a positive effect on viability of the less resistant L. rhamnosus LGG and L. paracasei F-19. The beneficial effect was generally observed for the high-methoxylated pectins, indicating that substituted polygalacturonic acid in the backbone is essential for bacterial protection. Other pectin features associated with improved survival, included less negative Zeta-potential, higher molecular weight, as well as lower values of hydrodynamic sizes, viscosity and degree of branching. The study indicates that pectins have a potential to protect probiotic bacteria through the gastro-intestinal transit and identifies the features linked to their functionality.

Occurrence of lactic acid bacteria and yeasts at species and strain level during spontaneous fermentation of mawè, a cereal dough produced in West Africa.

Mawè is a West African spontaneous fermented cereal-based dough. Different types of mawè exist varying in type of cereal and/or production condition, with fermentations lasting 24-48 h. With the aim of obtaining a comprehensive understanding of the microbial ecology of mawè processing, a microbiological characterisation was performed for four mawè types, produced at eight sites in Benin. At the onset of the fermentations lactic acid bacteria (LAB) and yeast counts were on average 7.5 ±â€¯1.03 and 4.8 ±â€¯0.79 Log10 cfu/g, which increased to 9.2 ±â€¯0.38 and 7.4 ±â€¯0.42 Log10 cfu/g, respectively, at the end of the fermentations. LAB (n = 321) and yeasts (n = 298), isolated during the fermentations, were identified. The predominant LAB and yeast species were Lactobacillus fermentum and Pichia kudriavzevii, respectively, followed by Kluyveromyces marxianus, all present throughout the mawè fermentations. Further, microbial successions took place with Weissella confusa occurring mostly at the onset, while Pediococcus acidilactici and Saccharomyces cerevisiae were mainly associated with the end of the fermentations. Species diversity was influenced both by type of cereal and production condition. The dominating strain clusters of L. fermentum and P. kudriavzevii were ubiquitous and strain diversities were influenced by type of cereal and production site.

Prevalence, virulence factor genes and antibiotic resistance of Bacillus cereus sensu lato isolated from dairy farms and traditional dairy products.

BACKGROUND: B. cereus are of particular interest in food safety and public health because of their capacity to cause food spoilage and disease through the production of various toxins. The aim of this study was to determine the prevalence, virulence factor genes and antibiotic resistance profile of B. cereus sensu lato isolated from cattle grazing soils and dairy products in Ghana. A total of 114 samples made up of 25 soil collected from cattle grazing farm land, 30 raw milk, 28 nunu (yoghurt-like product) and 31 woagashie (West African soft cheese). Ninety-six B. cereus sensu lato isolates from 54 positive samples were screened by PCR for the presence of 8 enterotoxigenic genes (hblA, hblC, hblD, nheA, nheB, nheC, cytK and entFM), and one emetic gene (ces). Phenotypic resistance to 15 antibiotics were also determined for 96 B. cereus sensu lato isolates. RESULTS: About 72% (18 of 25 soil), 47% (14 of 30 raw milk), 35% (10 of 28 nunu) and 39% (12 of 31 woagashi) were positive for B. cereus sensu lato with mean counts (log10 cfu/g) of 4.2 ± 1.8, 3.3 ± 2.0, 1.8 ± 1.4 and 2.6 ± 1.8 respectively. The distribution of enterotoxigenic genes revealed that 13% (12/96 isolates) harboured all three gene encoding for haemolytic enterotoxin HBL complex genes (hblA, hblC and hblD), 25% (24/96 isolates) possessed no HBL gene, whereas 63% (60/96 isolates) possessed at least one of the three HBL genes. All three genes encoding for non-haemolytic enterotoxin (nheA, nheB and nheC) were detected in 60% (57/96) isolates, 14% (13/96) harboured only one gene, 19% (18/96) whereas 8% possessed none of the NHE genes. The detection rates of cytk, entFM, and ces genes were 75, 67 and 9% respectively. Bacillus cereus s. l. isolates were generally resistant to ß-lactam antibiotics such as ampicillin (98%), oxacillin (92%), penicillin (100%), amoxicillin (100%), and cefepime (100%) but susceptible to other antibiotics tested. CONCLUSIONS: Bacillus cereus s. l. is prevalent in soil, raw milk and dairy products in Ghana. However, loads are at levels considered to be safe for consumption. Various enterotoxin genes associated with virulence of B. cereus are widespread among the isolates.

Influence of extracellular pH on growth, viability, cell size, acidification activity, and intracellular pH of Lactococcus lactis in batch fermentations.

In this study, we investigated the influence of three extracellular pH (pHex) values (i.e., 5.5, 6.5, and 7.5) on the growth, viability, cell size, acidification activity in milk, and intracellular pH (pHi) of Lactococcus lactis subsp. lactis DGCC1212 during pH-controlled batch fermentations. A universal parameter (e.g., linked to pHi) for the description or prediction of viability, specific acidification activity, or growth behavior at a given pHex was not identified. We found viability as determined by flow cytometry to remain high during all growth phases and irrespectively of the pH set point. Furthermore, regardless of the pHex, the acidification activity per cell decreased over time which seemed to be linked to cell shrinkage. Flow cytometric pHi determination demonstrated an increase of the averaged pHi level for higher pH set points, while the pH gradient (pHi-pHex) and the extent of pHi heterogeneity decreased. Cells maintained positive pH gradients at a low pHex of 5.5 and even during substrate limitation at the more widely used pHex 6.5. Moreover, the strain proved able to grow despite small negative or even absent pH gradients at a high pHex of 7.5. The larger pHi heterogeneity at pHex 5.5 and 6.5 was associated with more stressful conditions resulting, e.g., from higher concentrations of non-dissociated lactic acid, while the low pHi heterogeneity at pHex 7.5 most probably corresponded to lower concentrations of non-dissociated lactic acid which facilitated the cells to reach the highest maximum active cell counts of the three pH set points.

In vitro investigation of Debaryomyces hansenii strains for potential probiotic properties.

In this study, 23 Debaryomyces hansenii strains, isolated from cheese and fish gut, were investigated in vitro for potential probiotic properties i.e. (1) survival under in vitro GI (gastrointestinal) conditions with different oxygen levels, (2) adhesion to Caco-2 intestinal epithelial cells and mucin, and (3) modulation of pro- and anti-inflammatory cytokine secretion by human monocyte-derived dendritic cells. As references two commercially available probiotic Saccharomyces cerevisiae var. boulardii (S. boulardii) strains were included in the study. Our results demonstrate that the different D. hansenii yeast strains had very diverse properties which could potentially lead to different probiotic effects. One strain of D. hansenii (DI 09) was capable of surviving GI stress conditions, although not to the same degree as the S. boulardii strains. This DI 09 strain, however, adhered more strongly to Caco-2 cells and mucin than the S. boulardii strains. Additionally, two D. hansenii strains (DI 10 and DI 15) elicited a higher IL-10/IL-12 ratio than the S. boulardii strains, indicating a higher anti-inflammatory effects on human dendritic cells. Finally, one strain of D. hansenii (DI 02) was evaluated as the best probiotic candidate because of its outstanding ability to survive the GI stresses, to adhere to Caco-2 cells and mucin and to induce a high IL-10/IL-12 ratio. In conclusion, this study shows that strains of D. hansenii may offer promising probiotic traits relevant for further study.

Technological properties and probiotic potential of Lactobacillus fermentum strains isolated from West African fermented millet dough.

BACKGROUND: Throughout Africa, food fermentations are still driven by indigenous microorganisms which influence the nutritional, organoleptic and safety of the final products. However, for improved safety, consistent quality and beneficial health effects, a trend has emerged which involves the isolation of indigenous strains from traditional fermented products to be used as functional starter cultures. These functional starter cultures possess inherent functional characteristics and can contribute to food quality and safety by offering one or more organoleptic, nutritional, technological or health advantage (probiotics). With the aim of selecting potential probiotic starter cultures, Lactobacillus fermentum strains isolated from fermented millet dough were investigated for technological properties and probiotic traits in-vitro. RESULTS: A total of 176 L. fermentum strains were assessed for technological properties including rate of acidification, exopolysaccharide production and amylase activity. Following this, 48 strains showing desirable technological properties were first screened for acid resistance. Sixteen acid resistant strains were assessed for additional probiotic properties including resistance to bile salts, bile salt hydrolysis, antimicrobial property, haemolysis and antibiotics resistance. L. fermentum strains clustered into 3 groups represented by 36 %, 47 % and 17 % as fast, medium and slow acidifiers respectively. About 8 %, 78 % and 14 % of the strains showed strong, weak and no exopolysaccharides production respectively. Amylase activity was generally weak or not detected. After exposure of 48 L. fermentum strains to pH 2.5 for 4 h, 16 strains were considered to be acid resistant. All 16 strains were resistant to bile salt. Four strains demonstrated bile salt hydrolysis. Antimicrobial activity was observed towards Listeria monocytogenes and Staphylococcus aureus but not E. coli and Salmonella enteritidis. Lactobacillus fermentum strains were generally susceptible to antibiotics except 6 strains which showed resistance towards streptomycin, gentamicin and kanamycin. CONCLUSION: In vitro determination of technological and probiotic properties have shown strain specific difference among L. fermentum strains isolated from fermented millet dough. Sixteen (16) L. fermentum strains have been shown to possess desirable technological and probiotic characteristics in vitro. These strains are therefore good candidates for further studies to elucidate their full potential and possible application as novel probiotic starter cultures.

Hanseniaspora jakobsenii sp. nov., a yeast isolated from Bandji, a traditional palm wine of Borassus akeassii.

Investigation of the microbial diversity of Bandji, a traditional palm wine from Burkina Faso (West Africa) revealed the presence of two yeast isolates (YAV16 and YAV17T) with unusual phenotypic and genotypic characteristics. The isolates divide by bipolar budding with no production of ascospores. Phylogenetic analysis of concatenated sequences of the 26S rRNA gene D1/D2 and internal transcribed spacer (ITS) regions indicated that the novel species was most closely related to Kloeckera lindneri and Hanseniaspora valbyensis. The new isolates differed from K. lindneri NRRL Y-17531T and H. valbyensis CBS 479T by substitutions in the D1/D2 region of 12 and 16 nt respectively. The divergence in the ITS region from the closely related species was characterized by substitutions of 45-46 nt. Repetitive palindromic PCR (rep-PCR) profiles of YAV16 and YAV17T were also significantly different from those of K. lindneri MUCL 31146T ( = NRRL Y-17531T), H. valbyensis NCYC 17T ( = CBS 479T) and other species of the genus Hanseniaspora. Based on the results of the phenotypic and genotypic characterizations, it was concluded that the new isolates represent a novel species for which the name Hanseniaspora jakobsenii sp. nov. is proposed with YAV17T ( = CBS 12942T = DSM 26339T = NCYC 3828T; MycoBank number MB 805785) as the type strain.

Effect of dissolved oxygen on redox potential and milk acidification by lactic acid bacteria isolated from a DL-starter culture.

Milk acidification by DL-starter cultures [cultures containing Lactococcus lactis diacetylactis (D) and Leuconostoc (L) species] depends on the oxidation-reduction (redox) potential in milk; however, the mechanisms behind this effect are not completely clear. The objective of this study was to investigate the effect of dissolved oxygen on acidification kinetics and redox potential during milk fermentation by lactic acid bacteria (LAB). Fermentations were conducted by single strains isolated from mixed DL-starter culture, including Lactococcus lactis ssp. lactis, Lactococcus lactis ssp. cremoris, and Leuconostoc mesenteroides ssp. cremoris, by the DL-starter culture, and by the type strains. High and low levels of oxygen were produced by flushing milk with oxygen or nitrogen, respectively. The kinetics of milk acidification was characterized by the maximum rate and time of acidification (Vamax and Tamax), the maximum rate and time of reduction (Vrmax and Trmax), the minimum redox potential (Eh7 final), and time of reaching Eh7 final (Trfinal). Variations in kinetic parameters were observed at both the species and strain levels. Two of the Lc. lactis ssp. lactis strains were not able to lower redox potential to negative values. Kinetic parameters of the DL-starter culture were comparable with the best acidifying and reducing strains, indicating their additive effects. Acidification curves were mostly diauxic at all oxygen levels, displaying 2 maxima of acidification rate: before (aerobic maximum) and after (anaerobic maximum) oxygen depletion. The redox potential decreased concurrently with oxygen consumption and continued to decrease at slower rate until reaching the final values, indicating involvement of both oxygen and microbiological activity in the redox state of milk. Oxygen flushing had a negative effect on reduction and acidification capacity of tested LAB. Reduction was significantly delayed at high initial oxygen, exhibiting longer Trmax, Trfinal, or both. Concurrently, anaerobic acidification rate maximum Vamax was decreased and Tamax was extended. Fermentation kinetics in nitrogen-flushed milk was not statistically different from that in untreated milk except for Lc. lactis ssp. lactis CHCC D2, which showed faster reduction time after nitrogen flushing. This study clarifies the relationship between the redox state in milk and acidification kinetics of the predominant subspecies in DL-starter cultures. This knowledge is important for dairies to ensure optimized, fast, and controlled milk fermentations, leading to greater standardization of dairy products.

Expression of Virulence-Related Genes in Listeria monocytogenes Grown on Danish Hard Cheese as Affected by NaCl Content.

Expression of virulence-related genes in Listeria monocytogenes incubated on cheese was assessed by real-time quantitative polymerase chain reaction. The objective of the study was to investigate the impact of sodium chloride concentration in cheese on transcription of virulence genes and, thereby, virulence potential of L. monocytogenes. The expression studies were performed with L. monocytogenes strains characterized by different tolerance to salt stress. Strains ATCC(®) 51779 and DSMZ 15675 were incubated on the Danish hard-cheese type Samsoe, with low (<0.15% [wt/wt]) and high (3.6% [wt/wt]) content of NaCl. Genes differentially expressed (p<0.05) through the 48-h incubation were transcriptional regulators prfA and agrA, genes of the main virulence cluster inlA, hly, actA, involved in invasion of the epithelial cells, and genes bsh, opuC, gadC, clpP, and ami, associated with osmotic stress responses in L. monocytogenes. The more sensitive strain ATCC(®) 51779 was most responsive, showing significant upregulation of prfA, actA, hly, and bsh both at low and high NaCl. Strain DSMZ 15675 was less responsive to NaCl stress, showing reduced or consistent gene transcription at all conditions. Decreased transcription of agrA, ami, gadC, and opuC in both strains was promoted by low NaCl content. The study indicated that virulence gene expression of L. monocytogenes grown in cheese was affected by NaCl content and that effect was more significant in strains sensitive to both hypo- and hyperosmotic stresses.