MRS Medium Research Articles

Bio preservation capacity of potentially probiotic Lacticaseibacillus strains in fermented sausage

SummaryBio preservation constitutes a well‐acknowledged natural method for controlling the growth of undesirable bacteria in meat products. The aim was to evaluate four different potentially probiotic Lacticaseibacillus strains as bio preservatives and their application in fermented meat sausage. The strains were inoculated for 24 h and used a concentration of 0.1% of the total mass into which it was directly inserted. Regarding the inhibition zone, only the commercial strains exhibited effect on the tested pathogens. They also displayed better growth results on MRS medium, which simulates meat product additives, during the 24‐h period. The pH, sodium nitrite and sodium chloride proved to be limiting factors for the growth of the potentially probiotic Lacticaseibacillus strains. Lc. paracasei BGP1 significantly reduced water activity and pH – both fundamental factors for food conservation‐ when incorporated. Lc. paracasei BGP1 demonstrated its efficacy as bio preservative for use in fermented meat sausage. Technological parameters were evaluated, showing positive responses to the application of acid lactic bacteria. These results indicate the potential of lactic acid bacteria to inhibit certain microorganisms, offering a perspective on inhibition when used in fermented meat products. Hence, further studies could be conducted, primarily concerning Listeria sp.

Characterization of Lactobacillus Isolated from the Microflora of Different Fermentation Processes and Doderlein Flora

Characterization of Lactobacillus Isolated from the Microflora of Different Fermentation Processes and Doderlein Flora

Aguamiel Enhance Proteolytic Activity and Survival of Lactiplantibacillus pentosus ABHEAU-05 during Refrigerated Storage of a Fermented Milk

Different carbon sources, such as prebiotics, have promoted probiotics’ survival during refrigerated fermented milk storage. These compounds stimulate both the metabolic response and the resistance of probiotics to adverse conditions, such as low temperatures. That is why the objective of this study was to evaluate the kinetic parameters of growth, the proteolytic profile, and the survival of Lactiplantibacillus pentosus ABHEAU-05 in fermented milk added with aguamiel as a prebiotic source during refrigerated storage. Inulin was used for control experiments. A 12% w/v powdered skimmed milk solution was inoculated with L. pentosus ABHEAU-05 (106 CFU/mL). It was fermented at 37 °C until a pH of 4.5, and the kinetic parameters were calculated. Analysis of survival and proteolytic profile during refrigeration storage (4 °C for 21 days) was carried out. The survival of the microorganism was determined by viable count on MRS agar, the production of free amino groups by the TNBS method, and the accumulation of low molecular weight peptides by polyacrylamide gel electrophoresis (SDS-PAGE). The pH of 4.5 was reached 26 h before the control. The maximum concentration of viable cells was 108 CFU/mL at the fermentation’s end and maintained throughout the refrigerated storage. With the analysis of the proteolytic profile, high metabolic activity was demonstrated during fermentation and refrigerated storage in milk with aguamiel. The accumulation of low molecular weight peptides and the generation of free amino groups were higher than the control results. It was verified that aguamiel is a carbon source with the potential for developing and maintaining the probiotic L. pentosus ABHEAU-05 in fermented milk.

Probiotic Lactobacillus strains isolated from date waste and wastewater for the kidney stone, intestinal oxalate-degradation and antioxidant activity

Lactic acid bacteria (LAB) are probiotic organisms and are promising alternatives to manage kidney stone problems. Oxalate digesting LAB was screened from date wastes and date wastewater. LAB was isolated using De Man, Rogosa and Sharpe agar (MRS agar) medium and was subjected to oxalate degradation assay in a calcium oxalate plate. The strain was further subjected to assay for its capacity to tolerate acidic pH value and bile salt. Then, anti-urolithiatic activity assay was carried out by in vitro analysis. Based on preliminary characteristics, among 62 oxalate degrading strains, twenty-four strains exhibited a greater than 10 mm zone. The potent bacterial strains were identified as Lactobacillus cassei, L. acidophilus, L. gasseri and L. fermentum. Isolates from date wastes showed better antagonistic properties towards bacterial pathogens. At 50 µg Calcium oxalate concentration, 75.6 ± 0.865 degradation was achieved by L acidophilus. LAB isolated from date waste showed promising anti-urolithiasis activity. Hence, date waste may serve as a potential source for LAB probiotics to treat hyperoxaluria.

High-degree polymerizate IMOs of dextranase hydrolysates enhance Lactobacillus acid metabolism: Based on growth, and metabolomic and transcriptomic analyses

This study aimed to explore the growth and metabolic mechanisms and functional properties of the combination of high-degree polymerizate (DP) isomaltooligosaccharides (IMOs) and Lactobacillus to provide a basis for their subsequent applications in functional foods in improving human health. Metabolomic and transcriptomic analyses were used to select the important metabolic pathways, regulatory genes, and metabolites involved in the growth of Lactobacillus by high DP IMOs. Lactobacillus paracasei JLPF-176 grew well in the MRS medium with high DP IMOs. The best effect in promoting L. paracasei JLPF-176 biofilm formation was 58.94%. When high DP IMOs hydrolyzed for 48 h were added to the MRS medium, 684 genes were upregulated and 276 genes were downregulated in L. paracasei JLPF-176, in important pathways such as d-amino acid metabolism, tryptophan metabolism, arginine and proline metabolism. HMDB compound classification statistics revealed 421 organic acids and derivatives, accounting for the highest number of 28.39%. The number of genes annotated in amino acid metabolism was 107.

Isolation of Health-benefiting Bacteria from Probiotic Foods

Probiotic Foods are known to have many health benefits such as bolstering intestinal health, reducing risk to many diseases, and reducing antibiotic resistance, to name a few. Because these foods have so many benefits, I hypothesized that by isolating bacteria from them, these health benefits could be extended to all kinds of different foods. The bacteria of interest were isolated from Great Value Greek Plain Nonfat Yogurt by using MRS medium to encourage the growth of lactic acid bacteria. After several strains had been isolated, they were used to ferment orange juice and carrot juice for about three days. By using HPLC (High-Performance Liquid Chromatography), several differences between the original juices and the fermented juices were found. E. Coli Bacteria were tested against antibiotics as well as salt concentrations after being exposed to one of two treatments: fermented juice and non-fermented juice. Through this experiment, I have been able to observe positive results from three bacterial strains. I was able to observe a decrease in antibiotic resistance to ampicillin, hygromycin, carbenicillin, and kanamycin when I exposed E. Coli bacteria to juices fermented by my isolated bacteria.

Production of nisin from Lactococcus lactis in acid-whey with nutrient supplementation

The production of Nisin, an FDA-approved food preservative, was attempted by Lactococcus lactis subsp. lactis ATCC® 11454 using the underutilized milk industry effluent, acid-whey, as a substrate. Nisin production was further improved by studying the effect of supplementation of nutrients and non-nutritional parameters. The addition of yeast extract (6% w/v) as nitrogen source and sucrose (4% w/v) as carbon source were found to be suitable nutrients for the maximum nisin production. The changes in the medium pH due to lactic acid accumulation during batch fermentation and its influence on the production of nisin were analyzed in the optimized whey medium (OWM). The production characteristics in OWM were further compared with the nisin production in MRS media. The influence of nisin as an inducer for its own production was also studied and found that the addition of nisin at 0.22 mg/ml promote the nisin production. The analysis of consumption of various metal ions present in the OWM during the nisin production was also analyzed, and found that the copper ions are the most consumed ion. The highest nisin yield of 2.6 × 105 AU/mL was obtained with OWM.

Preferential sugar utilization by bifidobacterial species.

Aim: Bifidobacteria benefit host health and homeostasis by breaking down diet- and host-derived carbohydrates to produce organic acids in the intestine. However, the sugar utilization preference of bifidobacterial species is poorly understood. Thus, this study aimed to investigate the sugar utilization preference (i.e., glucose or lactose) of various bifidobacterial species. Methods: Strains belonging to 40 bifidobacterial species/subspecies were cultured on a modified MRS medium supplemented with glucose and/or lactose, and their preferential sugar utilization was assessed using high-performance thin-layer chromatography. Comparative genomic analysis was conducted with a focus on genes involved in lactose and glucose uptake and genes encoding for carbohydrate-active enzymes. Results: Strains that preferentially utilized glucose or lactose were identified. Almost all the lactose-preferring strains harbored the lactose symporter lacS gene. However, the comparative genomic analysis could not explain all their differences in sugar utilization preference. Analysis based on isolate source revealed that all 10 strains isolated from humans preferentially utilized lactose, whereas all four strains isolated from insects preferentially utilized glucose. In addition, bifidobacterial species isolated from hosts whose milk contained higher lactose amounts preferentially utilized lactose. Lactose was also detected in the feces of human infants, suggesting that lactose serves as a carbon source not only for infants but also for gut microbes in vivo. Conclusion: The different sugar preference phenotypes of Bifidobacterium species may be ascribed to the residential environment affected by the dietary habits of their host. This study is the first to systematically evaluate the sugar uptake preference of various bifidobacterial species.

Bionanotechnology of Selenite Ions Recovery into Nanoselenium by Probiotic Strains of Lactobacteria and Tolerance of Lactobacteria to Sodium Selenite

Green synthesis of nanoparticles (NPs) using living cells is a promising and new tool in bionanotechnology. Chemical and physical methods are used to synthesize NPs, but biological methods are preferred because of their environmentally friendly, clean, safe, cost-effective, simple, and efficient sources for high productivity and purity. Aim. To investigate the processes of bioreduction of selenite ions into nanoselenium by probiotic strains of lactobacilli Lactobacillus plantarum IMV B-7679 and L. casei IMV B-7280. Methods. Cultivation of lactobacilli L. plantarum IMV B-7679 and L. casei IMV B-7280 was carried out in vials (500 cm3) on a rotary shaker (220 rpm) at 30 °C for 2 days on the Man, Rogosa, and Sharpe (MRS) broth nutrient medium. Sodium selenite was additionally added to the environment in different concentrations from 1 to 30 ppm by Se. The number of viable bacterial cells in 1 mL of suspension was determined by the method of limiting dilutions in the case of sowing aliquots on a nutrient medium containing 0.2% agar-agar. Cultures of L. plantarum IMV B-7679 or L. casei IMV B-7280 were grown in the liquid MRS broth medium with low pH in the presence or absence of Na2SeO3. The concentration of sodium selenite ranged from 1 to 30 ppm by Se level. The number of microorganisms was determined by inoculation (0.1 mL of suspension) in dense media on cups with MRS agar medium, and the seeding dose was 107 cells/Petri dish. The tolerance of lactobacilli to the selenite ions was evaluated by the decrease in the number of CFU when sowing aliquots taken from culture samples grown in the presence or absence of selenite. The results of the experiments were presented in CFU and transferred to Log CFU/cm3. The characteristics of Nano-Se were studied using transmission electron microscopy (TEM). Results. It was found that after 48 h incubation in an MRS medium with the addition of sodium selenite from 1 to 30 ppm, the culture of L. plantarum IMV В-7679 was the most resistant. Thus, enrichment of the culture medium with 30 ppm of Se in the Na2SeO3 composition led to a decrease in the number of L. plantarum IMV B-7679 to 5.17 ± 0.09 Log CFU/cm3 against 4.41 ± 0.11 Log CFU/cm3 for L. casei IMV B-7280 in the control. The use of lower concentrations (1—3 ppm of Se in Na2SeO3) did not affect the change in morphology and cultural properties of L. plantarum IMV B-7679. The ability of L. casei IMV B-7280 and L. plantarum IMV B-7679 cultures to grow on MRSA nutrient medium in the presence of 3 ppm Se was shown. Higher tolerance to sodium selenite was found for L. plantarum IMV B-7679. Thus, increasing the concentration to 30 ppm of Se in the form of Na2SeO3 led to a decrease in the viability of only the culture of L. casei IMV B-7280. That is, the studied lactobacilli showed different ability to grow in the presence of selenite ions. The formation of round electron-dense granules sizing from 30 nm to 250 nm was observed using TEM. Both probiotic strains showed the ability to restore selenite ions with the accumulation of intracellular Nano-Se and the release of Nano-Se into the culture medium, which was accompanied by color shifts from yellowish to red-brown. The partial destruction of L. casei IMV B-7280 cells under the influence of oxyanions was revealed, which was accompanied by the release of culture-synthesized electron-dense Nano-Se particles. Conclusions. The optimal conditions for the growth of L. plantarum IMV B-7679 and L. casei IMV B-7280 in the presence of Na2SeO3 were established, and it was proved that lactobacilli have different abilities to grow in the presence of selenite ions. The obtained data indicate that the investigated probiotic strains showed the ability to restore selenite ions along with the accumulation of intracellular Nano-Se and the release of Nano-Se into the culture medium.

METABOLISM AND FUNCTIONAL HETEROGENEITY OF FERMENTED MILK ORIGIN LACTIC ACID BACTERIA FOR LACTOSE INTOLERANCE

Lactic acid bacteria (LAB) are cosmopolitan in distribution with multiple ecological niches. LAB shows diverse applications in improvement of health by biochemical interference or imuno modulation to overcome several clinical circumstances. Lactose intolerance (LI) is one such situation, where individual show lactose maldigestion after the consumption of dairy products. It is necessary to overcome such a condition by employing indigenous beneficial bacteria or their products. Therefore, in this study we have isolated and characterized LAB from fermented milk samples, from remote villages of districts of south India. Traditionally fermented milk samples (68) were collected, cultivated on MRS medium, identified by biochemical and carbohydrate metabolic activity and correlated with Bergey's manual of systematic bacteriology. When matched with other LAB, Lactiplantibacillus isolates were able to reduced pH of medium significantly and reached pH of 4.6 in 48 hours. On MRS agar, 450 different bacterial isolates were isolated, recognized as presumptive LAB and classified up to the level of genera as Lactiplantibacillus (285), Lactococcus (70), Pediococcus (19), Streptococcus (20), and Enterococcus (16). Later, β-galactosidase screening was carried out using MRS/X-gal agar medium. Out of 450 LAB isolates only Lactiplantibacillus isolates were potential β-galactosidase producers. GRAS organisms such as LAB are multifaceted diverse group of bacteria localized in varieties of fermented foods/in the intestine and recognized as probiotics. Distinct contribution of LAB in health care and disorder management made this organism as a choice for alternate therapy; hence functionality of LAB can be promoted for LI management.

Production of Lactic Acid by lactobacilli using Legume Flours

Due to their high nutritional value, richness in carbohydrates and good source of protein, legumes are considered essential for human diet. Legumes flours were used as fermentable material for lactic acid production. Hence, previously isolated lactobacilli strains were used in this study. The strains showed strong microbial growth and their survival in glucose-containing MRS medium and were described using a modified Gompertz equation. Lactobacilli exhibited the shortest latency phase in MRS-glucose medium. While the highest lactic acid produced was 15.40g/l by Lpb. pentosus U1 isolate after 48hours. Furthermore, these strains were evaluated in flours samples for acidification capacity. The results showed a decrease in pH values after 12hours of fermentation, indicating the fastest acidification with maximum lactic acid generation of 31.66, 25.33, 31.00 and 31.66g/l for black gram flour by Lpb. plantarum M1, Lpb. pentosus U1, Lpb. plantarum VIP1 and Lev. brevis TIP1 isolates. Black gram flour was considered the best among the used substrates for enhancing lactic acid production. Overall, this study can be termed as a preliminary step for fermentation using the desired organism in preparing legume-based food at the industrial level and benefiting consumers.

Inhibition of Aspergillus carbonarius growth and Ochratoxin A production using lactic acid bacteria cultivated in an optimized medium

The Gram-positive bacteria lactic acid bacteria (LAB) are used in the food industry but are also known for inhibiting certain food spoilage microorganisms, especially fungi. Sources of nitrogen (N) for culture media are generally organic and expensive. Many attempts have been made to formulate economical culture media with alternative N sources obtained from agricultural and industrial byproducts. This study describes the design and optimization of an inexpensive culture medium for Lactiplantibacillus plantarum (formerly Lactobacillus plantarum) MZ809351 strain B31. The culture medium was optimized using statistical experimental designs to identify the factors with the most significant effects on biomass concentration to reduce the overall cost, aiming to obtain a biomass concentration similar to that obtained with the reference LAB culture medium (de Man, Rogosa and Sharpe; MRS). Sodium acetate and magnesium sulfate were the most significant factors (p < 0.005), and their contents were reduced by 22 % and 40 %, respectively, without affecting biomass concentration. Malt germ extract (MGE) was used as an alternative nitrogen source to replace meat extract (ME) and proteose peptone (PP). Through these experiments, the composition of a culture medium that is less expensive than MRS broth was defined, which produced a biomass concentration (3.8 g/L) similar to that obtained with MRS medium. The inhibitory effects of two LAB strains isolated from the Ivory Coast and Mexico on the growth and production of ochratoxin A (OTA) in an ochratoxigenic fungus was tested. The minimum cellular concentration of the LAB to prevent the development of Aspergillus carbonarius Ac 089 and the production of OTA was determined in a model assay in Petri dishes. The conditions to inhibit the germination of A. carbonarius Ac 089 and the production of OTA were found. Using the optimized medium and a ratio of 2 × 104 LAB/spore (1 × 108 CFU/mL) strain B7 (L. plantarum MZ809351) and 2 × 103 LAB/spore (1 × 107 CFU/mL) strain B31 (L. plantarum MN922335) completely inhibited the growth of the fungus. A ratio of 2 × 105 LAB/spore (1 × 109 CFU/mL) was required to inhibit OTA production with strains B7 and B31. This study indicates the potential of cultivating LAB in an optimized and inexpensive culture medium for use as a biological control agent against ochratoxigenic fungi in food.

Genomic and biogenic amine-reducing characterization of Lactiplantibacillus planatraum JB1 isolated from fermented dry sausage

Biogenic amines are naturally present as hazardous, low-molecular organic basic compounds in various foods. Lactic acid bacteria show excellent potential to prevent biogenic amines accumulation. In this study, the amine-reducing mechanism of Lactiplantibacillus planatraum JB1, a strain isolated from fermented sausages, was explored according to its genome, and its ability to degrade biogenic amines was also evaluated in culture medium and dry sausage. The results showed that L. plantarum JB1 possessed a myriad of desirable properties, including tolerance to harsh environments and safety for assessment in vitro. Its genome contained genes encoding transport proteins and multicopper oxidase, which may be involved in biogenic amine degradation. In PBS and MRS medium, L. plantarum JB1 could degrade eight biogenic amines with high degradation rates for tyramine and putrescine. By inoculating sausages with L. plantarum JB1, the total content of biogenic amine was decreased by 32.84% after preparation and 32.07% after storage. Due to its good adaptability, security and amine-reducing capacity, L. plantarum JB1 could be considered a starter culture to ensure the safety of meat products.

Characterization of Bacterial Exopolysaccharides Produced from Different Fruit-Based Solid Media

Bacterial exopolysaccharides (EPSs) share their compositional and structural features with plant dietary fibers. Therefore, analysis of bacterial EPSs produced during fermentation of fruit or vegetables interferes with that of plant fibers. To get rid of this effect, bacteria were grown on a solid medium containing fruit juice or purée and EPSs were recovered in a quantitative dependent manner and were quantified with the phenol-sulfuric acid colorimetric method. The protocol was assayed both on MRS medium and fruit-based media, with three bacterial strains from two species, Leuconostoc pseudomesenteroides 56 and Weissella cibaria 21 and 64. With that method, differences in EPS production levels were shown according to the strain and cultivation conditions, such as sucrose content and pH. Complementary analysis with NMR indicated that glucose and sucrose were partly recovered with EPSs, pointing out the requirement for a further purification step. It also showed that EPSs’ ramified structure differed according to the strain and the fruit used in the medium. This method for EPS recovery is helpful to select strains and to pilot EPS production during lactic fermentation of fruit or vegetable foods.

Effects of Bacteriocin Extracted from Lactobacillus rhamnosus on Cutaneous Wound Healing in Mice

Aim: This study evaluates the effects of bacteriocin produced by Lactobacillus rhamnosus on cutaneous wound healing in mice.&#x0D; Study Design: Randomized experimental design was employed in the study.&#x0D; Methodology: Probiotic L. rhamnosus was isolated from nono milk using MRS agar media and identified based on morphological and biochemical characteristics. The potent bacterium was subjected to bacteriocin production. Antimicrobial activity of the bacteriocin was carried out against wound pathogenic bacteria and wound healing effects of the bacteriocins were assessed on mice. 2 mm cutaneous wound was induced on the mice and crude bacteriocin was topically applied on the wounds. White blood cell count was carried out after days 3, 5, 9, and 14. Histopathological analyses were performed after day 14.&#x0D; Results: Crude bacteriocin was effective against wound pathogenic bacteria such as S. aureus, P. aeruginosa and E. coli. Topical application of bacteriocin showed a faster and better wound healing when compared to positive control groups in P. aeruginosa infected wound model. Histology examination revealed an increase wound healing time in L. rhamnosus bacteriocin treated mice. The crude bacteriocins demonstrated excellent wound-healing activity in skin excision albino mice model.&#x0D; Conclusion: L. rhamnosus bacteriocin had various beneficial actions on wounds. It improved wound healing, inhibited wound pathogens and increase wound healing time.

Growth characteristics of lactic acid-producing strains using glucose syrup as a carbon source

This work investigates the growth and productivity characteristics of such lactic-acid producing strains, as Lactobacillus delbrueckii subsp. bulgaricus 19-11 (VKPM B-2368), Lactobacillus acidophilus 5 Ds (VKPM B-2846) and Lactococcus lactis subsp. lactis (VKM B-1662) on standard MRS medium using glucose syrup as a carbon substrate. According to the results of batch cultivation of the selected strains in 5L fermenters for 72 h, the productivity was established to decrease in the Lactobacillus delbrueckii subsp. bulgaricus 19-11 &gt; Lactobacillus acidophilus 5 Ds &gt; Lactococcus lactis subsp. lactis series. L. delbrueckii subsp. bulgaricus 19-11 showed the maximum lactic-acid productivity of 1.94 g/(l×h) with a glucose conversion degree of 87%. After cultivation, a slight decrease in the content of nitrogen, potassium and sodium in the culture liquid of the studied strains was observed. In all strains, the content of other macronutrients (phosphorus, calcium, sulphur, magnesium, barium and iron) increased in proportion to the addition of glucose syrup during cultivation, which is directly related to their significant content in its composition. The Lactobacillus delbrueckii subsp. bulgaricus 19-11 and Lactobacillus acidophilus 5 Ds strains produced racemic (DL) lactic acid, whereas Lactococcus lactis subsp. lactis produced lactic acid with a 73% L-isomer content. The use of glucose syrup in biotechnological processes can contribute to the implementation of waste-free production in the respective enterprises.

Isolation and some basic characteristics of lactic acid bacteria from beetroot (Beta vulgaris L.) —A preliminary study

The carbohydrate content of fruit and vegetable juices has made them a suitable environment for the growth of lactic acid bacteria. Various studies have shown that beetroot can be a good substrate for the production of probiotic drinks for consumers. Therefore, this study was performed to isolate and identify lactic acid bacteria from beetroot juice. Beetroot juice was prepared from two kilograms of washed beetroot with a cold press. Sterile samples of beetroot juice were first enriched in MRS broth and then cultured on MRS agar. Gram and catalase tests were used for the initial detection of lactic acid bacteria. Then, to identify the species of lactic acid bacteria, biochemical confirmation tests such as the melting of gelatin, and the growth of bacteria has been studied in the ideal condition that is 15-45° C, sugars fermentation, gas production from glucose, and growth in at different concentrations of salt were performed. The final verification of species was done using polymerase chain reaction-amplified 16S rRNA-specific DNA. All isolates formed creamy white round colonies on MRS agar medium, were gram-positive, were able to grow in anaerobic conditions, and were also rod-shaped and catalase-negative. Based on biochemical tests, four types of lactic acid bacteria (Lactobacillus plantarum, Lactobacillus helveticus, Lactobacillus salivarius, and Lactobacillus acidophilus) were identified. The findings of this study indicated the presence and diversity of lactic acid bacteria in beet juice, which can be a potential platform for the production of probiotic beetroot juice as a non-dairy product.

Philodulcilactobacillus myokoensis gen. nov., sp. nov., a fructophilic, acidophilic, and agar-phobic lactic acid bacterium isolated from fermented vegetable extracts.

Lactic acid bacteria are commonly in the fermentation industry and pose potential positive effects on health. In this study, a new lactic acid bacterium was isolated from fermented vegetable extracts in Myoko, Niigata, Japan. This bacterium is fructophilic, acidophilic, and hard to grow on agar medium. The isolate is Gram-stain-positive, non-spore-forming, non-motile, rod-shaped, and catalase-negative. Growth occurred at pH 3.5-5.5, with optimal growth at pH 4.5-5.0. The cells formed colonies on a solid MRS medium with 20% (w/v) sucrose and 0.8% (w/v) gellan gum under anaerobic conditions. The bacterium was able to grow on up to 50% (w/v) sucrose but not on d-glucose. Moreover, 16S rRNA gene sequence analysis revealed that the strain was most closely related to Apilactobacillus ozensis (93.1% sequence similarity). The values of average nucleotide identity, digital DNA-DNA hybridization, average amino acid sequence identity, and amino acid identity of conserved genes were calculated between the isolated strain (type strain is WR16-4T = NBRC 115064T = DSM 112857T) and its phylogenetically closest type strains. The average nucleotide identity values (73.36-78.28%) and DNA-DNA hybridization values (16.3-32.9%) were significantly lower than the threshold values for species boundaries. The average amino acid sequence identity values (53.96-60.88%) were significantly below the threshold boundary of genus demarcation (68%). The amino acid identity of conserved genes values compared to strain WR16-4T were the genera Apilactobacillus, Nicoliella spurrieriana SGEP1_A5T, Acetilactobacillus jinshanensis HSLZ-75T, and Fructilactobacillus were 62.51-63.79%, 62.87%, 62.03%, and 58.00-61.04%, respectively. The 16S rRNA gene and core genome phylogenetic trees suggested that this novel strain was most closely related to the type strain of A. jinshanensis HSLZ-75T. Based on the physiological, morphological, and phenotypical characteristics of strain WR16-4T, we propose its classification as a novel genus, Philodulcilactobacillus myokoensis gen. nov., sp. nov.

In vitro assessment of biofunctional properties of Lactiplantibacillus plantarum strain Jb21-11 and the characterization of its exopolysaccharide.

The microbiota of traditional food provides a rich reservoir of biodiversity to find new strains with interesting features for novel functional food formulation. Therefore, this study aimed to investigate the biofunctional potential of the lactic acid bacteria (LAB) strain Jb21-11 isolated from Jben, a traditional Algerian fresh cheese. This isolate was selected out of a collection of 154 LAB based on its exopolysaccharide (EPS) phenotype and was preliminarily identified by polyphasic characterization as Lactiplantibacillus plantarum (previously known as Lactobacillus plantarum) and its biofunctional properties were then assessed in vitro. The tested strain demonstrated good resistance to gastric juice, acidity around pH 2, and 2% (v/v) bile salts, which are important characteristics for potential biofunctional LAB candidates. It also showed a good production of ropy EPS with 674 mg/L on MRS medium. However, this ability appears to compromise the adhesion of the strain to Caco-2 cells (less than 1%), which according to our results, seems not to be related to autoaggregation and hydrophobicity (44.88 ± 0.028% and 16.59 ± 0.012%). Furthermore, promising antimicrobial activity against three pathogenic bacteria (Escherichia coli, Staphylococcus aureus, and Salmonella) was detected probably due to antimicrobial metabolites excreted during fermentation process into the medium. Moreover, the strain L. plantarum Jb21-11 displayed a therapeutic functionality with both anti-inflammatory and immunomodulatory action using RAW 264.7 cells. The chemical features of the novel ropy Jb21-11-EPS were also investigated revealing the presence of three monosaccharides, namely, mannose, galactose, and glucose, with a molar ratio of 5.42:1.00:4.52 linked together by α- and β-glycosidic bonds, presenting a relatively high molecular weight of 1.08 × 105 Da of interest for a texturing potential. Therefore, the new producing EPS strain Jb21-11 is a promising candidate for use as an adjunct culture for improving the texture of functional food.

Probiotic Lactic Acid Bacteria for Vaginal Application. Optimization of Biomass Production and Freeze-Drying Conditions

Probiotic formula for the vaginal tract must contain high numbers of viable beneficial bacteria that maintain their characteristics during the production and further storage, in order to exert the claimed probiotic effect. Four probiotic strains—Lactobacillus gasseri CRL1320, Limosilactobacillus reuteri CRL1324, Ligilactobacillus salivarius CRL1328 and Lacticaseibacillus rhamnosus CRL1332—originally isolated from vagina of healthy women exhibiting beneficial properties were evaluated. Therefore, the optimization of growth parameters and dry biomass production with high viability while preserving probiotic features of strains is a great challenge. Different growth conditions in MRS medium were set up in a laboratory bioreactor; two initial pH and agitation were recorded speeds during static or controlled fermentations. Production conditions of 37 °C with controlled pH 6.5 and 5.5-MRS with 150 and 75 rpm stirring speeds were used for CRL1329 and CRL1332 strains, respectively, while static and free pH MRS for CRL1324 and CRL1328 probiotics allowed us to obtain maximal cell viability counts. However, during 12 h of fermentation, biomass yields of 19.3, 16.2 and 15.2 g/L were achieved when CRL1329, CRL1328 and CRL1324 probiotic strains were grown in static and free pH MRS. The highest biomass yield for CRL1332 strain was produced under controlled MRS-initial pH 6.5 and 75 rpm fermentation conditions. To preserve probiotic high biomass viability, freeze-drying was carried out in the presence of different cryoprotective agents. Thus, the highest viable numbers (10.9–11.8 log CFU/g) with survival rates between 91.3 and 95.6% were attained in the presence of 10% trehalose (L. reuteri and L. salivarius), lactose (L. rhammosus) and lactose + trehalose + sucrose mix (L. gasseri). When stability during post-freeze-drying storage was evaluated, probiotic strains showed a remarkably higher viability recovery when stored at 4 °C than at 25 °C for 12 and 3 months, respectively. In addition, surface characteristics of vaginal probiotics were affected to different extents during storage depending on the strain, protective agent and storage time/temperature. Critical factors for growth conditions, drying process and storage stability of probiotic lactobacilli strains were optimized in view to preserve cell high viability and surface features for the design of vaginal probiotic formula.