Potential Probiotic Bacteria Research Articles

Probiotic potential of bacterial endophytes isolated from leaves of <em style="mso-bidi-font-style: normal;">Murraya</em><em style="mso-bidi-font-style: normal;"> </em><em style="mso-bidi-font-style: normal;">koenigii</em><em style="mso-bidi-font-style: normal;"> </em>L.

Probiotics are live microorganisms which confer health benefits to the host when administered in adequate amounts. Endophytes, which live in plant tissues between the plant cells, can act as potential probiotic bacteria. The leaves of Murraya koenigii L. is a common cuisine and herbal ingredient used in indigenous medicine. The main objective of the current study was to assess probiotic characteristics of isolated bacterial endophytes of M. koenigii leaves. Young and mature leaves of M. koenigii were collected from sites in the wet and dry zones of Sri Lanka. The endophytes were isolated using three techniques: placing leaf segments, leaf macerations and preparation of pour plates on three different media viz., nutrient agar, Luria-Bertani (LB), and De Man, Rogosa and Sharpe agar (MRS). They were identified from morphological characteristics and preliminary biochemical tests. Probiotic characterization of the isolates was carried out by using a series of standard tests including resistance to low pH, tolerance of bile salts, antimicrobial activity (Escherichia coli and Pseudomonas aeruginosa), antibiotic resistance (Gentamycine), anti-haemolytic activity, and non-DNase activity. Ten endophytic bacteria (two bacilli, eight cocci including two Gram positive cocci, and eight Gram negative cocci and bacilli) were isolated from the young and mature leaf samples collected. While probiotic characterization tests were positive, four isolates showed antibiotic susceptibility. These results indicate the possibility that M. koenigii leaves possess bacterial endophytes with probiotic potential.

Exploring autochthonous strains with probiotic potential: A comprehensive characterization of functional properties and their application in fermented blueberry-watermelon smoothies

Fruit-fermented smoothies enriched with probiotics present a promising alternative for individuals who avoid dairy products. However, identifying suitable probiotic strains poses a critical challenge. This study aimed to characterize five potential probiotic strains: Hanseniaspora opuntiae BIOTEC045, Pediococcus pentosaceus BIOTEC046 and BIOTEC047, and Lactiplantibacillus plantarum LpAv and 73a. The objective was to assess their aggregation and antipathogenic abilities, safety, stress tolerance, and other functional properties. LpAv and 73a displayed the most favorable characteristics. They exhibited high auto-aggregation (>65%), resistance to simulated gastrointestinal conditions (with only 0.3–1.5 log reductions), efficient enzyme production (high levels of β-galactosidase and β-glucosidase), capability to utilize various prebiotics (with maximum optical density values of 1.62–1.81), and production of gamma-aminobutyric acid (1.2 mM), among other features. LpAv and 73a were selected for fermenting blueberry-watermelon smoothies (BWS). During a 7-day storage period at 5 °C, both strains maintained high viability (>9 log colony forming units/mL), while the control samples exhibited spoilage microbial load, highlighting the effectiveness of lactic acid fermentation as a viable bio-preservation technique. Additionally, the BWS retained bioactive compounds and antioxidant activity throughout storage after fermentation, with total phenolic and anthocyanin contents of 0.68–0.73 mg/g and 0.16–0.17 mg/g, respectively. These findings confirm the suitability of BWS as a matrix for fermentation with the selected potential probiotic bacteria. Additionally, the comprehensive characterization of the strains provides valuable insights into their functional properties and contributes to the understanding of their potential health benefits.

Elucidating the Mechanisms of Cell-to-Cell Crosstalk in Probiotics Co-culture: A Proteomics Study of Limosilactobacillus reuteri ZJ625 and Ligilactobacillus salivarius ZJ614.

Limosilactobacillus reuteri ZJ625 and Ligilactobacillus salivarius ZJ614 are potential probiotic bacteria with improved benefits when administered to the host as a multi-strain preparation. To elucidate the mechanisms of cell-to-cell crosstalk between these two strains, we studied their intracellular and extracellular proteomes in co-culture by liquid-chromatography mass-spectrometry (LC-MS) using Dionex Nano-RSLC and fusion mass spectrometer. The experiment consisted of five biological replicates, and samples were collected during the mid-exponential growth phase. The quantitative proteomic profiles revealed several differentially expressed proteins (DEPs), which are down- or up-regulated between and within groups for both the intracellular and extracellular proteomes. These DEPs include proteins synthesising autoinducer-2, a sensor compound for cell-to-cell bacterial crosstalk during quorum sensing in mixed culture. Other important DEPs identified include enolase, phosphoglycerate kinase, and l-lactate dehydrogenase, which play roles in carbohydrate metabolism. Proteins associated with transcription, ATP production and transport across the membrane, DNA repair, and those with the potential to bind to the host epithelium were also identified. The post-translational modifications associated with the proteins include oxidation, deamidation, and ammonia loss.Importantly, this study revealed a significant expression of S-ribosylhomocysteine lyase (luxS) involved in synthesising autoinducer-2 that plays important roles in quorum sensing, aiding bacterial cell-to-cell crosstalk in co-cultures. The proteome of L. salivarius ZJ614 was most affected when co-cultured with L. reuteri ZJ625. In contrast, omitting some medium components from the defined medium exerted more effects on L. reuteri ZJ625 than L. salivarius ZJ614.

Evaluation of Rouxiella badensis Subsp Acadiensis (Canan SV-53) as a Potential Probiotic Bacterium.

The advent of omic platforms revealed the significant benefits of probiotics in the prevention of many infectious diseases. This led to a growing interest in novel strains of probiotics endowed with health characteristics related to microbiome and immune modulation. Therefore, autochthonous bacteria in plant ecosystems might offer a good source for novel next-generation probiotics. The main objective of this study was to analyze the effect of Rouxiella badensis acadiensis Canan (R. acadiensis) a bacterium isolated from the blueberry biota, on the mammalian intestinal ecosystem and its potential as a probiotic microorganism. R. acadiensis, reinforced the intestinal epithelial barrier avoiding bacterial translocation from the gut to deep tissues, even after feeding BALB/c mice for a prolonged period of time. Moreover, diet supplementation with R. acadiensis led to increases in the number of Paneth cells, well as an increase in the antimicrobial peptide α defensin. The anti-bacterial effect of R. acadiensis against Staphylococcus aureus and Salmonella enterica serovar Typhimurium was also reported. Importantly, R. acadiensis-fed animals showed better survival in an in vivo Salmonella enterica serovar Typhimurium challenge compared with those that received a conventional diet. These results demonstrated that R. acadiensis possesses characteristics of a probiotic strain by contributing to the reinforcement and maintenance of intestinal homeostasis.

Antibiotic resistance in potential probiotic lactic acid bacteria of fermented foods and human origin from Nigeria

IntroductionProbiotic lactobacilli are generally recognized as safe (GRAS) and are being used in several food and pharma formulations. However, growing concern of antibiotic resistance in bacterial strains of food origin and its possible transmission via functional foods is increasingly being emphasized.ObjectivesThis study screened potential probiotic lactic acid bacteria (LAB) strains for their phenotypic and genotypic antibiotic resistance profiles.MethodsSusceptibility to different antibiotics was assayed by the Kirby Bauer standard disc diffusion protocol. Both conventional and SYBR-RTq-PCR were used for detection of resistance coding genes.ResultsA variable susceptibility pattern was documented against different antibiotic classes. LAB strains irrespective of origin displayed marked phenotypic resistance against cephalosporins, aminoglycosides, quinolones, glycopeptides; and methicillin among beta-lactams with few exceptions. In contrast, high sensitivity was recorded against macrolides, sulphonamides and carbapenems sub-group of beta-lactams with some variations. parC, associated with ciprofloxacin resistance was detected in 76.5% of the strains. Other prevalent resistant determinants observed were aac(6?)Ii (42.1%), ermB, ermC (29.4%), and tetM (20.5%). Six (?17.6%) of the isolates were free from genetic resistance determinants screened in this study.ConclusionStudy revealed presence of antibiotic resistance determinants among lactobacilli from both fermented foods and human sources.

Methods of selection of probiotic microorganisms with high adhesive properties

The ability to adhere to the intestinal epithelium is a classic criterion for the selection of potential probiotic bacteria, which can lead to temporary colonization, which will promote immunomodulatory effects, as well as stimulate the intestinal barrier and metabolic functions. In this research paper, several methods of selection of probiotic microorganisms are considered in order to determine the most useful and proliferative strains for subsequent use in clinical practice in the correction of metabolic disorders and relief of inflammatory processes of the gastrointestinal tract.

Antimycotic effect of 3-phenyllactic acid produced by probiotic bacterial isolates against Covid-19 associated mucormycosis causing fungi.

The Covid-19 associated mucormycosis (CAM) is an emerging disease affecting immunocompromised patients. Prevention of such infections using probiotics and their metabolites persist as effective therapeutic agents. Therefore, the present study emphasizes on assessment of their efficacy and safety. Samples from different sources like human milk, honey bee intestine, toddy, and dairy milk were collected, screened and characterized for potential probiotic lactic acid bacteria (LAB) and their metabolites to be used as effective antimicrobial agents to curtail CAM. Three isolates were selected based on probiotic properties and characterized as Lactobacillus pentosus BMOBR013, Lactobacillus pentosus BMOBR061 and Pediococcus acidilactici BMOBR041 by 16S rRNA sequencing and MALDI TOF-MS. The antimicrobial activity against standard bacterial pathogens showed ˃9 mm zone of inhibition. Furthermore, the antifungal activity of three isolates was tested against Aspergillus flavus MTCC 2788, Fusarium oxysporum, Candida albicans and Candida tropicalis where the results showed significant inhibition of each fungal strain. Further studies were carried out on lethal fungal pathogens like Rhizopus sp. and two Mucor sp. which are associated with post Covid-19 infection in immunosuppressed diabetic patients. Our studies on CAM inhibitory effect of LAB revealed the efficient inhibition against Rhizopus sp. and two Mucor sp. The cell free supernatants of three LAB showed varied inhibitory activity against these fungi. Following the antimicrobial activity, the antagonistic metabolite 3-Phenyllactic acid (PLA) in culture supernatant was quantified and characterized by HPLC and LC-MS using standard PLA (Sigma Aldrich). The isolate L. pentosus BMOBR013 produced highest PLA (0.441 g/L), followed by P. acidilactici BMOBR041 (0.294 g/L) and L. pentosus BMOBR061 (0.165 g/L). The minimum inhibitory concentration of HPLC eluted PLA on the Rhizopus sp. and two Mucor sp. was found to be 180 mg/ml which was further confirmed by inhibition of total mycelia under live cell imaging microscope.

Characterization of Short-chain fatty acid-producing and cholesterol assimilation potential probiotic Lactic acid bacteria from Chinese fermented rice

Probiotic strains are phrased as beneficial for life because this bacterial association has substantial effects on human and animal health when administrated orally, among which bio-active metabolite short-chain fatty acids production and cholesterol assimilation are most significant. All the short-chain fatty acids, mainly acetate, propionate, and butyrate reveal valuable effects on human health. In this research, the ability of 9 the potential lactic acid bacteria food-isolated probiotics was investigated to produce bio-active metabolite short-chain fatty acids directly or indirectly, and cholesterol-lowering ability through various mechanisms. Their 16S rRNA gene sequences were analyzed. short-chain fatty acids were quantified by gas chromatography, while cholesterol assimilation was measured using the o-phthalaldehyde method with slight modifications. All isolated strains through this study have the ability to optimize cholesterol content in the culture broth, short-chain fatty acid production, and survival in extreme alkaline and acidic conditions. The highest resistance to bile salt concentrations, γ-hemolysis, and strong antibacterial activity against pathogenic indicators, especially Escherichia coliATCC25922. The highest Lipid accumulation inhibitory effect was noticed in Lentilactobacillus hilgardii LMG 07934 as a potential probiotic isolate on the basis of cholesterol assimilation capacity (i.e., 91.4%), along with the ability to produce abundant i.e., butyric acid, propionic acid, and acetic acid, followed by Lacticaseibacillus paracasei ATCC 334 which has 85.3% cholesterol assimilation ability. These findings demonstrated that these isolated strains from Chinese fermented rice can be used as a potential probiotic and alternate therapeutic supplement because of their ability to produce short-chain fatty acids and their cholesterol-lowering ability.

Draft Genome Sequence of Lactiplantibacillus plantarum Strain ISO1, a Potential Probiotic Bacterium Isolated from the Milk of South African Saanen Goats.

Lactiplantibacillus plantarum strain ISO1, a potential probiotic with wide food and biotechnological applications, was isolated from the raw milk of South African Saanen goats. Here, we report the draft genome sequence of this bacterium. Sequencing yielded a 3.23-Mb genome and 2,967 protein-coding sequences.

In <i>Vitro </i>Evaluation of Putative Probiotic Candidates Isolated from Various Origins

Present study describes isolation of potential probiotic lactic acid bacteria from chicken crop, human feces, buttermilk and chilly. The isolated <i>Lactobacillus</i> strains survive, tolerate and grow in MRS medium spiked with bile, salt and having acidic pH. The <i>Lactobacillus</i> isolates possess several probiotic properties, viz. (i) ability to bind gastrointestinal mucosa, up to ≥80% cells adhered mucin, (ii) 50% cells retained viability during oro-gastro-intestinal transit, (iii) all the isolates exhibited broad anti-microbial spectrum against food spoilage and gastro-intestinal pathogens,<i> Limosilactobacillus fermentum</i> SBM showed maximum inhibition, (iv) ability to produce enzymatic activities like L-asparaginase, β-galactosidase and bile salt hydrolase (BSH) activities, <i>Limosilactobacillus fermentum</i> SBM showed maximum L-asparaginase activity (2.567 U/ml), and<i> Lactiplantibacillus pentosus</i> GCHI showed maximum β-galactosidase activity (296±0.1 Miller’s Unit), (v) <i>Lactiplantibacillus pentosus</i> GCHI aggregated up to ≥92% after 24 h, and (vi) the <i>Lactobacillus</i> isolates were susceptible towards nucleic acid synthesis inhibitors and cell wall synthesis inhibitor antibiotics. These <i>Lactobacillus</i> strains do not possess haemolytic, mucin degrading and DNase activities indicating their safety. Further characterization of these strains indicated potential probiotic properties and their suitability in food formulations as probiotics. The study presents an interesting illustration of mining of potential probiotic strains from nature exhibiting health benefits for human being and animals.

Screening and activity of potential gastrointestinal probiotic lactic acid bacteria against Yersinia ruckeri O1b.

Yersiniosis of cultured Atlantic salmon is a recurrent fish health management challenge in many continents. The causative organism, Yersinia ruckeri, can reside latently in the gut and lead to acute infection and disease during hatchery and sea-transfer stages. One potential prevention approach is the administration of probiotic bacteria to suppress gut colonization of Y. ruckeri. Our study aimed to isolate and identify anti-Yersinia activity among lactic acid bacteria (LAB) isolated from the gastrointestinal tract (GIT) of aquatic animals. Of the 186 aquatic GIT isolates examined, three strains showed diffusible antimicrobial activity towards Y. ruckeri O1b. Analysis of 16 s rRNA gene sequences indicated the three bacterial strains were Enterococci, related to Enterococcus sp. (99%), Enterococcus thailandicus (99%), and Enterococcus durans (99%). Anti-Yersinia activity was maintained at neutral pH (~6.5-7.0), and in-vitro environmental tolerance assays showed the three strains could withstand simulated salmonids gastrointestinal tract conditions of: low pH (3.4) and 3% bile salt content. All three Enterococci strains showed higher adhesion to the intestinal mucus of Atlantic salmon than Y. ruckeri O1b (E. durans 24%, E. enterococcus sp. 25% and E. thailandicus 98%, compared to Y. ruckeri O1b 5%). However, only Enterococcus sp. and E. thailandicus were able to grow in the salmon intestinal mucus broth while E. durans showed no growth. Anti-Yersinia activity was completely inactivated by proteinase-K treatment, suggesting that the active compound/s are proteinaceous and may be bacteriocin-like inhibitory substances (BLIS). Our data indicate that Enterococcus sp. MA176 and E. thailandicus MA122 are potential probionts for the prevention of yersiniosis in salmonids. Further in-vivo studies are required to determine whether these bacteria reduce the incidence of yersiniosis in Atlantic salmon.

Isolation and Characterization of Lactobacillus crispatus, Lactococcus lactis, and Carnobacterium divergens as Potential Probiotic Bacteria from Fermented Black and Green Olives (Olea europaea): An Exploratory Study.

Table olives are becoming well recognized as a source of probiotic bacteria that might be used to create a health-promoting fermented food product by traditional procedures based on the activities of indigenous microbial consortia present in local environments. Methodology. In the present study, the characterization of probiotic bacteria isolated from mince, chunks, and brine of fermented green and black olives (Olea europaea) was done based on morphological, biochemical, and physiological characteristics. Bacterial isolates demonstrated excellent survival abilities at 25, 37, and 45°C and at a variable range of pH. However, the optimum temperature is 37 and the optimum pH is 7 for all three isolates. An antimicrobial susceptibility pattern was found among these isolates through the disc diffusion method. Most of the isolates were susceptible to streptomycin, imipenem, and chloramphenicol, whereas, amoxicillin showed resistance to these isolates, and variable results were recorded for the rest of the antibiotics tested. The growth of the isolates was optimum with the supplementation of 3% NaCl and 0.3% bile salt. The isolated bacteria were able to ferment skimmed milk into yogurt, hence making it capable of producing organic acid. Isolates of Lactobacillus crispatus MB417, Lactococcus lactis MB418 from black olives, and Carnobacterium divergens MB421 from green olives were characterized as potential candidates for use as starter cultures to induce fermentation of other probiotic food products.

SURVIVABILITY OF ENVIRONMENTAL BIFIDOBACTERIUM SPP. AND LACTIC ACID BACTERIA IN CURD CHEESE (TVAROG) PRODUCED USING ENVIRONMENTAL CULTURE

Background. Qualitative and quantitative composition of human intestinal microbiota depends on many factors. Those which are the most often mentioned include genetic and environmental determinants. Recent studies suggest that the structure of intestinal microbiota population differs not only regionally but also locally. Its composition determines the proper functioning of the entire body. In order to favorably colonize the intestine, a probiotic therapy can be implemented, significantly modulating the composition of intestinal microbiota. Owing to a correlation between the genetic diversity of LAB and their location, there is a need to isolate potential probiotic bacteria from the surrounding environment. Dietary supple ments and medicinal products containing probiotics available on the market often comprise too few mi croorganisms or a smaller number of species than declared on the packaging. The aim of the study was to determine the survival rate of environmental bacteria of the genus Bifidobacterium spp. and LAB, charac teristic of farms in the Świętokrzyskie Province, in curd cheeses produced from low-pasteurized milk and stored at 3 ± 2 °C for 14 days. Microbiological analyses were performed in accordance with ISO stand ards. Results and conclusions. Curd cheeses ensured a high number of bacteria of the genus Bifidobacte rium spp. and lactic acid bacteria meeting the requirements for probiotics. Environmental strains of Bifidobacterium spp. were characterized by good survivability in the tvarogs, exceeding their initial num ber after 14 days of storage. The curd cheeses were similar to each other in terms of overall acceptability during the sensory analysis throughout the entire storage period. The tested cheeses are in line with the current nutritional trends aimed at enriching the diet with natural products containing microorganisms characteristic of a given environment.

Biological properties and potential probiotic bacteria isolated from honey collected in Chiang Mai, Thailand

Biological properties and potential probiotic bacteria isolated from honey collected in Chiang Mai, Thailand

Safety Assessment of the Potential Probiotic Bacterium Limosilactobacillus fermentum J23 Using the Mexican Fruit Fly (Anastrepha ludens Loew, Diptera: Tephritidae) as a Novel In Vivo Model.

Safety assessment of probiotics is difficult but essential. In this work, the Mexican fruit fly, Anastrepha ludens (Loew) (Diptera: Tephritidae), was used as in vivo model to assess the biosafety of Limosilactobacillus fermentum J23. In the first set of experiments, the strain was orally administered to adult flies through direct feeding, whereas in the second set of experiments, it was supplemented through the larval rearing medium. Data showed that L. fermentum J23 did not lead to increased mortality or treatment-related toxicity signs in adult female and male flies. Ingestion of L. fermentum J23 by adult female flies led to a statistically significant improvement in locomotor activity compared to the control groups (ca. 59% decrease in climbing time, p < 0.0001). A positive trend in lifespan extension under stress (maximum lifespan = 144h) was also observed. When L. fermentum J23 was administered to the larvae, the adult emergence (p = 0.0099), sex ratio (p = 0.0043), and flight ability (p = 0.0009) increased significantly by 7%, 31%, and 8%, respectively, compared to the control diet. No statistical effect between the control diet and the L. fermentum J23-based diet for the number of pupae recovered, pupal weight, duration of the pupal stage, lifespan under stress, and morphological development was observed. We conclude that feeding L. fermentum J23 to the novel experimental model A. ludens had no toxic effects and could be safely considered a potential probiotic for food supplements; however, further studies are still needed to establish its biosafety in humans.

Insights into the bacterial community compositions of peanut worm (Sipunculus nudus) and their association with the surrounding environment

The peanut worm (Sipunculus nudus) is an important economic and ecological fishery resource in China. To determine how S. nudus interacts with its culture environment, in this study, the diversity, composition, and interrelationships of bacterial communities in the intestine, coelomic fluid, and culture environment of S. nudus through high-throughput sequencing using the Illumina HiSeq sequencing platform. Alpha diversity analysis showed that the highest bacterial community richness and diversity were found in the sediments. Species annotation revealed that Proteobacteria dominated in all samples (48.92%–58.36%), and the other dominant phyla were highly variable, indicating a certain independence between the environment and the composition of the bacterial community of S. nudus. The coelomic fluid of S. nudus contained high concentrations of Bacillus and could be a source of potential probiotic bacteria for isolation and culture. Cluster and operational taxonomic unit Venn diagram analyses showed that the bacterial community composition in the intestine was more similar to that in the sediments compared with water. The predicted functional analysis of bacterial communities indicated that the functions of bacterial communities in the different surrounding environment were highly similar and involved in various aspects, such as membrane transport, amino acid metabolism, carbohydrate metabolism, and replication and repair. Our findings have important implications on the establishment of sustainable microecological regulation and management strategies for S. nudus culture and provide a reference for the development and utilization of probiotic bacteria.

Probiotic lactobacilli mediate their immunoregulatory functions in intestinal cells via modulation of H3 histone acetylation.

Probiotics maintain intestinal homeostasis through the regulation of the immune response of the host. Hence, the role of histone modifications as epigenetic agents on immune modulations by potential probiotic bacteria has been investigated. Human colonic epithelial cells (Caco-2) pre-treated with class I histone deacetylase (HDAC) specific inhibitor, MS-275, were incubated either with potential probiotic bacteria (Limosilactobacillus fermentum MTCC 5898 and Lacticaseibacillus rhamnosus MTCC 5897) or Escherichia coli (ATCC 14948) as an inflammatory agent. Initially, transcriptional expression of potential immune related genes (IL-6, IL-8 and hBD-2) was analyzed using RT-qPCR and later H3 histone acetylation (H3Ac) at the promoter region of these genes was confirmed with chromatin immunoprecipitation (ChIP) assay respectively. Potentially probiotic L. fermentum (MTCC 5898) significantly suppressed (P<0.05) the inhibitor-mediated elevated expression of immune related genes while another strain L. rhamnosus (MTCC 5897) did not influence these gene expression results. In contrast, as an inflammatory agent, E. coli (ATCC 14948) synergistically augmented the expression of immune related genes. Later, ChIP analysis confirmed the occurrence of H3 acetylation at these genes' promoter region, which was directly related to the transcriptional activity of host epithelial cells stimulated by L. fermentum and E. coli respectively. But in the case of L. rhamnosus, MTCC 5897, acetylation did not follow the transcription pattern and potentiated H3Ac on the promoter regions of these genes. Potential probiotics used in the study were found to regulate the immune response of host cells through histone acetylation in a strain-specific manner.

Drug-Resistant Proteus Virulence Factors Characterization and Their Inhibition Using Probiotic Bacteria

Background: The genus Proteus is a Gram-negative bacterium with a unique characteristic of swarming. Mainly three species are involved in initiating urinary tract infections in the community and in immunocompromised patients, particularly in patients going through long-term catheterization. Due to their strong virulence factors like biofilm formations, protease, and hemolysin, they can lead to lengthening infections in affected individuals. Probiotics are live bacteria and yeasts that are beneficial to human health and can be used as an alternative for the control of nosocomial diseases. Lactobacilli are one of the common probiotics mostly found in yogurt and other fermented foods that have been used as a substitute for infection control. Objectives: The current study was designed to screen potential probiotic bacteria to encounter antibiotic-resistant and virulent Proteus species. Methods: In the current study, using probiotics, already known antibiotic-resistant isolates (n = 25) of Proteus were processed to characterize their virulence factors and their inhibition. Biofilm formation, protease, and hemolysin activities were studied using different phenotypic detection methods. Further, their virulence genes zapA, flg, hmpA, mrp, and rsbA were explored using their genomic DNA. These isolates were found resistant to different classes of antibiotics, and a strategy was designed to inhibit their growth by using probiotic bacteria isolated from the soil. Results: Virulence factors first, all isolates were subjected to biofilm detection, and they were 32% (n = 8) strong, 40% (n = 10) moderate, 16% (n = 4) weak, and 12% (n = 3) non-biofilm producers. All isolates were positive for swarming activity by showing a differentiated ring form of growth. Protease activity showed 56% (n = 14) isolates. Only 24% (n = 6) of isolates were positive for hemolysin. Virulence factors and molecular mechanisms were studied, and gene rsbA responsible for swarming was amplified in 17 (68%) Proteus isolates, and mrp responsible for fimbria was detected in 19 (76%) bacterial isolates. Further, these isolates were subjected to flagella, protease, and hemolysin, and it was revealed that flg 11 (44%), 13 (52%) protease coding zapA, and hmA gene coding hemolysin were amplified in 2 (8%) Proteus isolates. Probiotic bacteria isolated from soil samples were probed for antagonistic activity against Proteus species. The probiotic bacteria were identified as Lactobacillus plantarum, Bacillus subtilis, and B. licheniformis. Due to their strong growth inhibitory effects against Proteus, it is crucial to characterize further the metabolites that have shown suppressive results against Proteus. Conclusions: Findings from the current study will provide new avenues for drug development and also help clinicians manage resistant pathogens in healthcare settings. Probiotic applications for infection control can be useful in treating resistant pathogens. Further purification and characterization of metabolites will provide alternative options for managing resistance issues in microbes.

Application of 16S rRNA virtual RFLP for the discrimination of some closely taxonomic-related lactobacilli species

BackgroundSeveral species in Lactobacillaceae family were recognized as potential probiotic bacteria. In this group of lactic acid bacteria, species are taxonomically closed and usually share similar 16S rRNA gene, thus, instead of so their identification and discrimination are too difficult. MethodIn the present study, virtual restriction fragment length polymorphism (RFLP) is instead of was used as a tool to discriminate between the closely related species Lactiplantibacillus plantarum (L plantarum), Lactiplantibacillus paraplantarum (L paraplantarum), and Lactiplantibacillus pentosus (L pentosus); Latilactobacillus sakei (L sakei), Latilactobacillus curvatus(L curvatus), and Latilactobacillus graminis (L graminis); Lacticaseibacillus casei (L casei), Lacticaseibacillus paracasei (L paracasei), Lacticaseibacillus zeae, and Lacticaseibacillus rhamnosus; Lactobacillus gasseri (L gasseri) and Lactobacillus johnsonii (L johnsonii). In silico comparative analysis of 16S rRNA sequences digested by 280 restriction enzymes was performed in order to search the key enzymes which gives different profiles. ResultsResults revealed that L casei, L paracasei, L zeae, and Lb rhamnosus could be separated from each other on the basis of AlwI, BpuEI, BsgI, BsrDI, BstYI, EarI, MluCI, and NsPI RFLP. Results showed also that different RFLP patterns were obtained from L sakei, L graminis and L curvatus by using both AflI and NspI endonucleases (in separated restriction) and L plantarum, L paraplantarum, and L pentosus were distinguished each one from the other by MucI, NspI, and TspDTI PCR-RFLP. Lb gasseri and L johnsonii were also separated on the basis of Mse I, Taq I, and Dra I RFLP. ConclusionIn this study, we proved that too closely related species could be separated in virtual analysis on basis of their 16S rRNA RFLP patterns using key restriction enzymes method.

Isolation and identification of potential host probiotic bacteria from Malaysian mahseer, Tor tambroides for aquaculture practices

Abstract. Hossain MK, Iehata S , Noordin NM, Kader MA, Emilia SN, Sung YY, Abol-Munafi AB. 2022. Isolation and identification of potential host probiotic bacteria from Malaysian mahseer, Tor tambroides for aquaculture practices. Biodiversitas 23: 5423-5430. The digestive tract has been revealed as a reservoir of potential probiotics derived from the host. The goal of this study was to isolate, identify, and characterize putative host-associated probiotic bacteria (HAPs) from the digestive tract of Malaysian mahseer Tor tambroides. To evaluate potential T. tambroides HAPs, in-vitro digestive enzyme activity (such as a cellulolytic, proteolytic, and lipolytic activity) and antibacterial activity against two fish pathogens (Vibrio parahaemolyticus and Aeromonas hydrophila) were utilized. Thirty-seven isolates with digestive enzyme activities were tested for their ability to inhibit pathogenic bacteria growth using an agar well-diffusion assay. Three isolates displayed in-vitro suppression of pathogenic bacteria, with two strains (KT03 and KM07) inhibiting the growth of V. parahaemolyticus and one strain (KT27) suppressing the proliferation of both harmful bacteria (V. parahaemolyticus and A. hydrophila). Strains KT03 and KM07 exhibited the most resemblance to Enterococcus faecalis (strains 2674 and FC11682, respectively) based on 16S rRNA sequences, whereas KT27 had a 97% similarity to Aeromonas sp. A8-29. The study’s findings provide valuable data on the prospective use of these three isolates (KT03, KT27, and KM07) as potential HAPs for better understanding their physiological activities, such as growth and disease resistance on T. tambroides.