Strain S1 Research Articles

Probiotic properties, whole-genome sequence analysis, and safety assessment of BreviBacillus borstelensis S8

In previous studies, BreviBacillus borstelensis S8 exhibited favorable enzyme production characteristics. In this study, the whole genome of Br borstelenensis S8 was sequenced to analyze its genomic sequence information and functions, investigating its genotype and potential probiotic properties. The whole genome sequence of S8 was determined to be 5,559,180 bp with an average GC content of 51.5%. A total of 5406 protein-coding genes were identified and annotated using NCBI Prokaryotic Genome Annotation. By comparing it with the complete genome sequences of Bacillus cereus strains, 1911 orthologous gene families were recognized. Genome analysis revealed the absence of active antibiotic resistance genes, virulence factor genes, and toxin-encoding genes. Safety assessments included hemolytic assays, antibiotic resistance tests, and evaluations of biogenic amine production. Functional gene analysis highlighted S8's metabolic pathways based on its genomic features. The presence of adhesion-related genes, stress response genes (e.g., acid and bile salts), antimicrobial gene clusters, and genes involved in vitamin and essential amino acid synthesis in the genome showcased the probiotic potential of S8. Simultaneously, analysis of hemolytic activity, biogenic amines production ability, acid and bile salt tolerance for the strain S8 also demonstrated its probiotic traits. The genomic exploration and probiotic activity of S8 revealed its potential as a probiotic, offering valuable genomic insights and a foundation for its safe utilization in the food industry.

Synergistic suppression of grey mold on cut rose flower by combined application of methyl jasmonate and lipopeptides from Bacillus altitudinis

Roses are the most popular cut flowers that are widely used in various occasions and celebrations, yet can also be easily infected by grey mold caused by Botrytis cinerea; a notorious pathogen in the cut flower industry. A wide range of methods, including fungicides, have been used to control B. cinerea. Environmentally benign methods are increasingly sought after to minimize disease incidence. In this study, we evaluated 10 strains of Bacillus across eight species for their action against B. cinerea. Strains TM22A (Bacillus altitudinis) and S2 (Bacillus subtilis) were identified to be effective in disease suppression. Lipopeptides (LPs) were isolated from two strains, and their activity for inhibiting the growth of B. cinerea was tested. Additionally, methyl jasmonate (MeJA), a natural growth regulator, was evaluated for disease suppression using a in planta assay. Subsequently, LPs from TM22A and MeJA, alone or in combination, were evaluated for disease occurrence in cut rose flowers. Results showed that either LPs or MeJA at concentrations of 250 μmol/L significantly reduced lesion size and disease incidence. The combined application of LPs from TM22A with MeJA further reduced the disease occurrence in cut rose flowers. Furthermore, the activities of POD, PPO, PAL, CAT, and SOD enzymes were much higher in flowers treated with combined application of LPs from TM22A with MeJA. The enhanced suppression of B. cinerea is attributed to the antifungal effects of LPs and MeJA and activation of antioxidant enzymes. This study suggests that the combined application of LPs from TM22A with MeJA could be an environmentally friendly approach for control of grey mold disease in cut roses.

Evaluation of Probiotics Ability to Enhance Population Density, Growth Rate, and Neonates Production of Moina micrura in Different Environmental Parameters

Salinity, light intensity, and oxygen concentration are key environmental factors that significantly affect biological processes and the composition and dispersion of Moina biomass. Evaluating the effectiveness of probiotic enrichment in improving population density, growth rate, and neonate production can provide valuable details on the effectiveness of probiotics in enhancing the resilience and viability of Moina micrura under suboptimal circumstances. The purpose of this research project is to assess the efficacy of two probiotics, Bacillus pocheonensis strain S2 and Lysinibacillus fusiformis strain A1, in improving the population density, growth rate, and reproductive output in M. micrura across various environmental conditions. Moina micrura were treated with each probiotic at a volume of 5 x 105 CFU/ml under different levels of salinity (0, 2, 4, and 6 ppt), light intensity (800, 1,000, 1,500, and 2,000 lux), and oxygen concentration (80, 70, 60, and 50%). The results indicated that M. micrura treated with L. fusiformis A1 at 0 ppt attained the highest population density (6 ± 0.90 Ind./ml), growth rate (0.355 ± 0.030 µ), and number of offspring production (5 ± 0.75 Ind./ml). The highest point of population density (5 ± 0.07 Ind./ml), growth rate (0.381 ± 0.002 µ) and number of offspring (7 ± 0.41 Ind./ml) of M. micrura were obtained while treated with B. pocheonensis S2 at light intensity of 1,500 lux. Similarly, the highest population density (5 ± 0.60 Ind./ml), growth rate (0.365 ± 0.190 µ), and offspring production (2 ± 0.25 Ind./ml) of M. micrura were observed during enrichment with B. pocheonensis S2 at 70% oxygen concentration. Therefore, these results suggested that the optimum conditions for enriching M. micrura with B. pocheonensis S2 are salinity of 0 ppt, 70% oxygen concentration, and a light intensity level of 1,500 lux.

The quorum sensing SinI/SinR-TraI/TraR systems promote Pb stabilization by Ensifer adhaerens S24 in the Pb-polluted aquatic environment

In this study, the Pb-resistant Ensifer adhaerens strain S24, which contains quorum sensing (QS) systems responsible for N-acyl homoserine lactone (AHL) production, was investigated for QS system-mediated Pb stabilization and the underlying mechanisms. Whole-genome sequence analysis revealed the QS SinI/R and TraI/R systems in strain S24. Subsequently, strains S24 and the S24∆sinI/R, S24∆traI/R, S24∆traI/R/sinR, and S24∆sinI/R-traI/R/sinR mutants were constructed and compared for QS SinI/SinR-TraI/TraR system-mediated Pb stabilization in the solution and the mechanisms involved. After 5 days of incubation, strain S24 significantly decreased the Pb concentration in the Pb-contaminated solution compared with the mutants. The S24∆sinI/R-traI/R/sinR mutant exhibited reduced Pb stabilization and AHL activity than the other mutants. The S24∆sinI/R-traI/R/sinR mutant had significantly greater Pb concentrations in the solution and lower cell surface-adsorbed and extracellular precipitated Pb (PbS) contents as well as lower expression of H2S-producing genes of metC and sseA than did strain S24. Furthermore, the S24∆sinI/R-traI/R/sinR mutant displayed reduced interactions between the hydroxyl, amino, carboxyl, and ether groups and Pb, compared with strain S24. These findings implied the vital role of the SinI/SinR-TraI/TraR systems in strain S24 for Pb stabilization through enhanced cell surface adsorption and extracellular precipitation in Pb-polluted aquatic environments.

Mitigating the detrimental impacts of low- and high-density polyethylene microplastics using a novel microbial consortium on a soil-plant system: Insights and interactions

The accumulation of polyethylene microplastics (PE-MPs) in soil has raised considerable concerns; however, the effects of their persistence and mitigation on agroecosystems have not been explored. This study aimed to assess the detrimental effects of PE-MPs on a soil-plant system and evaluate their mitigation using a novel microbial consortium (MC). We incorporated low-density polyethylene (LDPE) and high-density polyethylene (HDPE) at two different concentrations, along with a control (0 %, 1 %, and 2 % w/w) into the sandy loam soil for a duration of 135 days. The samples were also treated with a novel MC and incubated for 135 days. The MC comprised three bacterial strains (Ralstonia pickettii (MW290933) strain SHAn2, Pseudomonas putida strain ShA, and Lysinibacillus xylanilyticus XDB9 (T) strain S7–10F), and a fungal strain (Aspergillus niger strain F1–16S). Sunflowers were subsequently cultivated, and physiological growth parameters were measured. The results showed that adding 2 % LDPE significantly decreased soil pH by 1.06 units compared to the control. Moreover, adding 2 % HDPE resulted in a more significant decrease in soil electrical conductivity (EC) relative to LDPE and the control. A dose-dependent increase in dissolved organic carbon (DOC) was observed, with the highest DOC found in 2 % LDPE. The addition of higher dosages of LDPE reduced soil bulk density (BD) more than HDPE. The addition of 2 % HDPE increased the water drop penetration time (WDPT) but decreased the mean weight diameter of soil aggregates (MWD) and water-stable aggregates (WSA) compared to LDPE. The results also revealed that higher levels of LDPE enhanced soil basal respiration (BR) and microbial carbon biomass (MBC). The interaction of MC and higher MP percentages considerably reduced soil pH, EC, BD, and WDPT but significantly increased soil DOC, MWD, WSA, BR, and MBC. Regarding plant growth, incorporating 2 % PE-MPs significantly reduced physiological responses of sunflower: chlorophyll content (Chl; −15.2 %), Fv/Fm ratio (-25.3 %), shoot dry weight (ShD; −31.3 %), root dry weight (RD; −40 %), leaf area (LA; −38.4 %), and stem diameter (StemD; −25 %) compared to the control; however, the addition of novel MC considerably reduced and ameliorated the harmful effects of 2 % PE-MPs on the investigated plant growth responses.

Clinical and Histopathological Evolution of Acute Intraperitoneal Infection by Streptococcus agalactiae Serotypes Ib and III in Nile Tilapia

Streptococcus agalactiae is a highly invasive bacterium that causes significant economic losses in tilapia aquaculture around the world. Furthermore, it is a pathogen for mammals, including humans, emphasizing its importance in One Health. The aim of this work was to evaluate the evolution of clinical and histopathological lesions caused by acute infection with two serotypes of S. agalactiae. For this, two strains isolated from natural outbreaks in Brazilian aquaculture farms (S13, serotype Ib; S73, serotype III) were used to challenge juvenile Nile tilapia (Oreochromis niloticus) intraperitoneally. Target organ samples were collected ten times, between 1 and 96 h post-infection, for microbiological and histopathological analyses. Anorexia was the first clinical sign and the first death occurred at 24 and 30 h in the fish infected with strains S13 and S73, respectively. Serotype Ib initially caused more pronounced lesions in the nervous system; however, serotype III lesions progressed more aggressively, reaching the same severity as those of serotype Ib. This trend was repeated in the mortality curve after 32 h. These results elucidated the important stages in the pathogenesis of S. agalactiae serotypes Ib and III in tilapia and suggest “tips and tricks” to improve the positive culture rate in the clinical diagnosis of infections in some tissues.

Role of Arthrospira Platensis in Preventing and Treating High-Fat Diet-Induced Hypercholesterolemia in Adult Rats.

Hyperlipidaemia is a recognised risk factor for cardiovascular disease. In this study, the antihyperlipidaemic properties of spirulina (Arthrospira platensis, strain S2 from Serbia) were tested in adult Wistar rats before and after induction of hypercholesterolaemia by a high-fat diet (HFD) to compare the preventive with the curative effect. Total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), alanine transaminase (ALT) and aspartate transaminase (AST) levels were measured in the blood samples. The chemical composition (lipids, proteins and cholesterol) and the content of bile acids in the faeces of the animals were also analysed. Feeding rats with an atherogenic diet for 10 weeks led to the successful development of hyperlipidaemia, as serum TC and LDL-C levels as well as lipids, cholesterol and bile acids in the animals' faeces were significantly increased. Pre- and post-treatment with spirulina led to a reduction in serum LDL, TC and ALT levels. Administration of spirulina resulted in both a significant increase in primary bile acids excretion and a decrease in bile acids metabolism, with pre-treatment being more effective than post-treatment in some cases. These results suggest that increased excretion of bile acids as well as an effect on the gut microbiota may be the mechanism responsible for the anti-hyperlipidaemic activity of the tested spirulina strain.

Antifungal Properties of Streptomyces bacillaris S8 for Biological Control Applications.

Soybean (Glycine max), a crucial global crop, experiences yearly yield reduction due to diseases such as anthracnose (Colletotrichum truncatum) and root rot (Fusarium spp.). The use of fungicides, which have traditionally been employed to control these phytopathogens, is now facing challenges due to the emergence of fungicide-resistant strains. Streptomyces bacillaris S8 strain S8 is previously known to produce valinomycin t through a nonribosomal peptide synthetase (NRPS) pathway. The objective of this study was to evaluate the antifungal activity of S. bacillaris S8 against C. truncatum and Fusarium sp., assessing its efficacy against soybean pathogens. The results indicate that strain S8 effectively controlled both above-ground and underground soybean diseases, using the NRPS and NRPS-related compound, suggesting its potential as a biological control in plant-microbe interactions. These findings underscore the pivotal role of the stain S8 in fostering healthy soybean microbial communities and emphasize the significance of microbiota structure studies in unveiling potent biocontrol agents.

Organic matter degradation in the deep, sulfidic waters of the Black Sea: insights into the ecophysiology of novel anaerobic bacteria

BackgroundRecent studies have reported the identity and functions of key anaerobes involved in the degradation of organic matter (OM) in deep (> 1000 m) sulfidic marine habitats. However, due to the lack of available isolates, detailed investigation of their physiology has been precluded. In this study, we cultivated and characterized the ecophysiology of a wide range of novel anaerobes potentially involved in OM degradation in deep (2000 m depth) sulfidic waters of the Black Sea.ResultsWe have successfully cultivated a diverse group of novel anaerobes belonging to various phyla, including Fusobacteriota (strain S5), Bacillota (strains A1T and A2), Spirochaetota (strains M1T, M2, and S2), Bacteroidota (strains B1T, B2, S6, L6, SYP, and M2P), Cloacimonadota (Cloa-SY6), Planctomycetota (Plnct-SY6), Mycoplasmatota (Izemo-BS), Chloroflexota (Chflx-SY6), and Desulfobacterota (strains S3T and S3-i). These microorganisms were able to grow at an elevated hydrostatic pressure of up to 50 MPa.Moreover, this study revealed that different anaerobes were specialized in degrading specific types of OM. Strains affiliated with the phyla Fusobacteriota, Bacillota, Planctomycetota, and Mycoplasmatota were found to be specialized in the degradation of cellulose, cellobiose, chitin, and DNA, respectively, while strains affiliated with Spirochaetota, Bacteroidota, Cloacimonadota, and Chloroflexota preferred to ferment less complex forms of OM. We also identified members of the phylum Desulfobacterota as terminal oxidizers, potentially involved in the consumption of hydrogen produced during fermentation. These results were supported by the identification of genes in the (meta)genomes of the cultivated microbial taxa which encode proteins of specific metabolic pathways. Additionally, we analyzed the composition of membrane lipids of selected taxa, which could be critical for their survival in the harsh environment of the deep sulfidic waters and could potentially be used as biosignatures for these strains in the sulfidic waters of the Black Sea.ConclusionsThis is the first report that demonstrates the cultivation and ecophysiology of such a diverse group of microorganisms from any sulfidic marine habitat. Collectively, this study provides a step forward in our understanding of the microbes thriving in the extreme conditions of the deep sulfidic waters of the Black Sea.Exyq52VguUrmavAnxe2qURVideo

Antagonistic and enzymatic activities of Bacillus species isolated from the fish gastrointestinal tract as potential probiotics use in Artemia culture.

Probiotics have been used successfully in aquaculture to enhance disease resistance, nutrition, and/or growth of cultured organisms. Six strains of Bacillus were isolated from the intestinal tracts of fish and recognised by conventional biochemical traits. The six isolated strains were Bacillus cereus and Bacillus subtilis using MALDI-TOF-MS technique. The probiotic properties of these Bacillus strains were studied. The tested bacillus strains exhibit antibacterial activity against the different pathogens. The strain S5 gave the important inhibition zones against most pathogens (20.5, 20.33, 23, and 21 mm against Vibrio alginolyticus, Vibrio parahaemolyticus, Staphylococcus aureus, and Salmonella typhimurium, respectively). According to our results, all Bacillus strains have extracellular components that can stop pathogenic bacteria from growing. The enzymatic characterization showed that the tested strains can produce several biotechnological enzymes such as α-glucosidase, naphtol-AS-BI-Phosphohydrolase, esterase lipase, acid phosphatase, alkaline phosphatase, amylase, lipase, caseinase, and lecithinase. All Bacillus strains were adhesive to polystyrene. The adding Bacillus strains to the Artemia culture exerted significantly greater effects on the survival of Artemia. The challenge test on Artemia culture showed that the protection against pathogenic Vibrio was improved. These findings allow us to recommend the examined strains as prospective probiotic options for the Artemia culture, which will be used as food additives to improve the culture conditions of crustacean larvae and marine fish.

Improving dryland maize productivity and water efficiency with heterotrophic ammonia-oxidizing bacteria via nitrification and cytokinin activity

A two-year field experiment conducted under dryland conditions in semi-humid and drought-prone regions of China aimed to assess the effect of ammonia-oxidizing bacterial on maize water use efficiency and yield. A heterotrophic ammonia-oxidizing bacteria (HAOB) strain S2_8_1 was used. Six treatments were applied: (1) no irrigation + HAOB strain (DI), (2) no irrigation + blank culture medium (DM), (3) no irrigation control (DCK), (4) irrigation + HAOB (WI), (5) irrigation + blank culture medium (WM), and (6) irrigation control (WCK). Results revealed that HAOB treatment increased maize growth, yield, and water use efficiency over controls, regardless of whether the year was wet or dry. This improvement was attributed to the accelerated nitrification in the rhizosphere soil due to HAOB inoculation, which subsequently led to increased levels of leaf cytokinins. Overall, these findings suggest that HAOB inoculation holds promise as a strategy to boost water use efficiency and maize productivity in dryland agriculture.

Non-Pigmented Antarctic Yeasts and Their Resistance to Toxic Metals

Despite the key role in biogeochemical processes and in the functioning of terrestrial ecosystems, yeasts of Antarctic regions still remain insufficiently studied. The study and analysis of the composition of Antarctic microbial communities remains relevant and is carried out using molecular biological approaches. The investigation of their resistance to toxic metal ions is essential to select industrially promising strains that can contribute to the development of new methods of metals detoxification via microorganisms. Aim. To determine the taxonomic position of non-pigmented Antarctic yeasts and investigate their resistance to toxic metal ions. Methods. The objects of the research are yeasts isolated from Antarctic phytocenoses. They were grown on malt wort (pH 5.0–5.5, temperature 18–20 °C). Isolation of genomic DNA was performed via the commercial DNA-sorb kit. Amplification of DNA was carried out using primers NL1 and NL4. Phylogenetic analysis was conducted by construction of trees (dendrograms) showing the position of the studied strains among closely related and typical species. The resistance of yeasts to toxic metal ions was established by cultivation in the concentration gradient of Ni2+, Co2+, CrO42-, and Сu2+. The ecophysiological traits of the isolated yeast strains including psychro- and halotolerance were determined. Results. Phylogenetic analysis showed a high percentage of similarity (99.5–99.6 %) of sequences of 18S rRNA genes of Antarctic yeast strains with the yeast sequences from the GenBank database. Psychrotolerant and halotolerant Antarctic yeast strains S11 and S12 were identified as Leucosporidium scottii and Debaryomyces hansenii, respectively. The studied yeast strains were found to be the most resistant to metal ions Ni2+ and Co2+. Strain of L. scottii S11 grew at 800 mg/L of Co2+, and D. hansenii S12 – at 750 mg/L of Ni2+. The yeasts were the least resistant to CrO42-: the L. scottii S11 and D. hansenii S12 strains grew at concentrations of 25 mg/L and 150 mg/L, respectively. In the presence of Cu2+, they grew at the same concentration – 600 mg/L. The combined action of toxic metal ions resulted in the increased toxic effects on the studied yeasts. Conclusions. The nucleotide sequences of 18S rRNA gene fragment of yeast strains S11 and S12 were included in the GenBank database under the numbers LT220858 and LT220859. Metal-resistant psychrotolerant yeast strains can be used to evaluate the metals content in polar regions as well as to bioremediate metal-contaminated ecosystems. However, further research is needed to develop and optimize bioremediation processes.

Analysis and modeling of two-dimensional piezoelectric semiconductor shell theory

In recent years, there has been a surge in research on piezoelectric semiconductors (PSs). However, studies specifically focused on two-dimensional PS structures remain scarce. In this paper, based on the principle of the virtual work method, the condition of charge continuity and the first-order thickness shear theory, we have established a two-dimensional model to investigate the electromechanical properties of two-dimensional PS shells. The basic field quantities, including the displacement, the electric potential and the carrier concentration perturbations were Fourier expanded along the width direction, and then the governing equations were successfully derived. The differential quadrature method was used to obtain numerical results for the PS shells with different loading modes and structural configurations. It was found that the inhomogeneous shear strain S13 is the key to the potential change in the PS shell structure. The emergence of the potential barriers and wells in the shell structure of PS is well explained. In addition, the position of the potential barriers can be modulated by varying the magnitude of the load, which provides a reference for the design of thin-film devices.

Characterization of a hemolytic and antibiotic-resistant Pseudomonas aeruginosa strain S3 pathogenic to fish isolated from Mahananda River in India.

Virulent strain Pseudomonas aeruginosa isolated from Mahananda River exhibited the highest hemolytic activity and virulence factors and was pathogenic to fish as clinical signs of hemorrhagic spots, loss of scales, and fin erosions were found. S3 was cytotoxic to the human liver cell line (WRL-68) in the trypan blue dye exclusion assay. Genotype characterization using whole genome analysis showed that S3 was similar to P. aeruginosa PAO1. The draft genome sequence had an estimated length of 62,69,783 bp, a GC content of 66.3%, and contained 5916 coding sequences. Eight genes across the genome were predicted to be related to hemolysin action. Antibiotic resistance genes such as class C and class D beta-lactamases, fosA, APH, and catB were detected, along with the strong presence of multiple efflux system genes. This study shows that river water is contaminated by pathogenic P. aeruginosa harboring an array of virulence and antibiotic resistance genes which warrants periodic monitoring to prevent disease outbreaks.

Degradation of pyrene by Xanthobacteraceae bacterium strain S3 isolated from the rhizosphere sediment of Vallisneria natans: active conditions, metabolite identification, and proposed pathways.

Polycyclic aromatic hydrocarbons (PAHs) were typical environmental contaminants that accumulated continuously in sediment. Microbial degradation is the main way of PAH degradation in the natural environment. Therefore, expanding the available pool of microbial resources and investigating the molecular degrading mechanisms of PAHs are critical to the efficient control of PAH-polluted sites. Here, a strain (identified as Xanthobacteraceae bacterium) with the ability to degrade pyrene was screened from the rhizosphere sediment of Vallisneria natans. Response surface analysis showed that the strain could degrade pyrene at pH 5-7, NaCl addition 0-1.5%, and temperature 25-40°C, and the maximum pyrene degradation (~ 95.4%) was obtained under the optimum conditions (pH 7.0, temperature 28.5°C, and NaCl-free addition) after 72h. Also, it was observed that the effect of temperature on the degradation ratio was the most significant. Furthermore, eighteen metabolites were identified by mass spectrometry, among which (2Z)-2-hydroxy-3-(4-oxo-4H-phenalen-3-yl) prop-2-enoic acid, 7-(carboxymethyl)-8-formyl-1-naphthyl acetic acid, phthalic acid, naphthalene-1,2-diol, and phenol were the main metabolites. And the degradation pathway of pyrene was proposed, suggesting that pyrene undergoes initial ortho-cleavage under the catalysis of metapyrocatechase to form (2Z)-2-hydroxy-3-(4-oxo-4H-phenalen-3-yl) prop-2-enoic acid. Subsequently, this intermediate was progressively oxidized and degraded to phthalic acid or phenol, which could enter the tricarboxylic acid cycle. Furthermore, the pyrene biodegradation by the strain followed the first-order kinetic model and the degradation rate changed from fast to slow, with the rate remaining mostly slow in the later stages. The slow biodegradation rate was probably caused by a significant amount of phenol accumulation in the initial stage of degradation, which resulted in a decrease in bacterial activity or death.

Traditional Bulgarian Fermented Foods as a Source of Beneficial Lactic Acid Bacteria.

Traditional Bulgarian fermented foods are prominent for their uniqueness of local ingredients, production methods, and endemic microbial species. The present research investigated the diversity and beneficial biological potential of lactic acid bacteria (LAB) isolated from various types of unique Bulgarian fermented foods. Species identification was performed via 16S rDNA sequencing. Biological activity was evaluated by determining antibacterial activity (via agar well diffusion assay), H2O2 production, spectrophotometrically determined auto- and co-aggregation, microbial adhesion to hydrocarbon, and biofilm formation. The biosafety of the isolated lactic acid bacteria was established based on hemolytic activity and phenotypic and genotypic antibiotic susceptibility. Forty-five strains were isolated from fermented foods (sauerkraut, fermented green tomatoes, fermented cucumbers, kefir, white cheese, and Izvara (curdled milk)). Five species were detected: Lactiplantibacillus plantarum, Levilactobacillus koreensis, Levilactobacillus brevis, Lactobacillus helveticus, and Levilactobacillus yonginensis. The most prominent species was L. plantarum, at 47%. For the first time, L. koreensis and L. yonginensis, isolated from unique Bulgarian fermented foods, are reported in this study. The antibacterial effect of the cell-free supernatants was evaluated. An antagonistic effect was observed against Escherichia coli (57%) and Salmonella enterica subsp. enterica serotype Enteritidis (19%) for several L. plantarum strains. Only one L. brevis (Sauerkraut, S15) strain showed activity against E. coli. The best autoaggregation ability at hour 4 was observed for L. koreensis (fermented cucumbers, FC4) (48%) and L. brevis S2 (44%). The highest percentage of co-aggregation with Candida albicans, at hou 4 in the experiments, was observed for strains L. koreensis (fermented green tomato, FGT1) (70%), L. plantarum strains S2 (54%), S13 (51%), and S6 (50%), while at hour 24 for strains L. koreensis FGT1 (95%), L. brevis (Kefir, K7) (89%), L. plantarum S2 (72%), and L. koreensis FC2 (70%). Seven of the isolated LAB strains showed hydrophobicity above 40%. Our results showed that the ability of biofilm formation is strain-dependent. No hemolytic activity was detected. The antibiotic resistance to 10 antibiotics from different groups was tested phenotypically and genotypically. No amplification products were observed in any strains, confirming that the isolates did not carry antibiotic-resistance genes. Traditional fermented Bulgarian foods can be considered functional foods and beneficial LAB sources.

Assessing immunogenicity of CRISPR-NCas9 engineered strain against porcine epidemic diarrhea virus

Porcine epidemic diarrhea (PED) caused by porcine epidemic diarrhea virus (PEDV), is an acute and highly infectious disease, resulting in substantial economic losses in the pig industry. Given that PEDV primarily infects the mucosal surfaces of the intestinal tract, it is crucial to improve the mucosal immunity to prevent viral invasion. Lactic acid bacteria (LAB) oral vaccines offer unique advantages and potential applications in combatting mucosal infectious diseases, making them an ideal approach for controlling PED outbreaks. However, traditional LAB oral vaccines use plasmids for exogenous protein expression and antibiotic genes as selection markers. Antibiotic genes can be diffused through transposition, transfer, or homologous recombination, resulting in the generation of drug-resistant strains. To overcome these issues, genome-editing technology has been developed to achieve gene expression in LAB genomes. In this study, we used the CRISPR-NCas9 system to integrate the PEDV S1 gene into the genome of alanine racemase-deficient Lactobacillus paracasei △Alr HLJ-27 (L. paracasei △Alr HLJ-27) at the thymidylate synthase (thyA) site, generating a strain, S1/△Alr HLJ-27. We conducted immunization assays in mice and piglets to evaluate the level of immune response and evaluated its protective effect against PEDV through challenge tests in piglets. Oral administration of the strain S1/△Alr HLJ-27 in mice and piglets elicited mucosal, humoral, and cellular immune responses. The strain also exhibited a certain level of resistance against PEDV infection in piglets. These results demonstrate the potential of S1/△Alr HLJ-27 as an oral vaccine candidate for PEDV control.Key points• A strain S1/△Alr HLJ-27 was constructed as the candidate for an oral vaccine.• Immunogenicity response and challenge test was carried out to analyze the ability of the strain.• The strain S1/△Alr HLJ-27 could provide protection for piglets to a certain extent.

A Bacillus velezensis strain shows antimicrobial activity against soilborne and foliar fungi and oomycetes.

Biological control uses naturally occurring antagonists such as bacteria or fungi for environmentally friendly control of plant pathogens. Bacillus spp. have been used for biocontrol of numerous plant and insect pests and are well-known to synthesize a variety of bioactive secondary metabolites. We hypothesized that bacteria isolated from agricultural soil would be effective antagonists of soilborne fungal pathogens. Here, we show that the Delaware soil isolate Bacillus velezensis strain S4 has in vitro activity against soilborne and foliar plant pathogenic fungi, including two with a large host range, and one oomycete. Further, this strain shows putative protease and cellulase activity, consistent with our prior finding that the genome of this organism is highly enriched in antifungal and antimicrobial biosynthetic gene clusters. We demonstrate that this bacterium causes changes to the fungal and oomycete hyphae at the inhibition zone, with some of the hyphae forming bubble-like structures and irregular branching. We tested strain S4 against Magnaporthe oryzae spores, which typically form germ tubes and penetration structures called appressoria, on the surface of the leaf. Our results suggest that after 12 hours of incubation with the bacterium, fungal spores form germ tubes, but instead of producing appressoria, they appear to form rounded, bubble-like structures. Future work will investigate whether a single antifungal molecule induces all these effects, or if they are the result of a combination of bacterially produced antimicrobials.

Pathogenic and genomic characterization of rabbit-sourced Pasteurella multocida serogroup F isolates recovered from dead rabbits with respiratory disease.

The 19 rabbit-sourced Pasteurella multocida serogroup F isolates showing high virulence to rabbits were avirulent to the broilers. Notably, polymorphisms were determined in the lipopolysaccharide outer core biosynthetic genes natC and gatF among all serogroup F strains of different hosts. However, the sequences of natC and gatF from rabbit-sourced strains (except for SD11) were identical, which might be responsible for the host specific of the 19 isolates.

In-vitro Inhibitory Activities of Potential Probiotic Isolated from Pangasius nasutus against Aeromonas hydrophila and Streptococcus agalactiae

In aquaculture, using probiotics is crucial for strengthening the immune system and encouraging the growth and survival of many aquatic organisms, including the Pangasius species. This approach is particularly significant given the impact of bacterial diseases on Pangasius survival. This study aimed to assess the effectiveness of probiotics isolated from Pangasius nasutus as alternatives to antibiotics for combating infections caused by Aeromonas hydrophila and Streptococcus agalactiae. Potential bacteria were isolated from the intestine and stomach of healthy P. nasutus. Seventy probiotic strains were successfully isolated and further screened using A. hydrophilaand S. agalactiae as pathogens in an in vitro disc diffusion assay. Preliminary screenings indicated that five probiotic strains inhibited the growth of A. hydrophila. Stomach-derived strain S1 and intestine-derived strain L1 suppressed A. hydrophila growth with inhibition zones of 10.5±1 mm and 8.5±1 mm, respectively. Likewise, strains L2, L8, and L12 from the intestine showed inhibitory zones of 6.0±1 mm, 6.5±1 mm, and 6.0±1 mm, respectively. Of these, only L12 inhibited the growth of S. agalactiae with a clear zone of 5.0±1 mm. In the elimination of pathogenic strains, potential strains S1 and L1 did not grow on the Aeromonas isolation medium. Co-culture assays demonstrated that both potential strains significantly inhibited Aeromonas hydrophila growth at concentrations of 106 and 108 CFU mL-1 over 48- and 96-hour periods, respectively. The potential bacterial strains were identified using 16s rRNA gene sequencing and were classified as follows: S1 - Lactococcus lactis, L1 - Weissella confusa, L2 - Cosenzaea myxofaciens, L8 - Lactococcus garvieae, and L12 - Plesiomonas shigelloides. Strain S1 L. lactis and strain L1 W. confusa are suggested for further evaluation and acquired additional research to fully elucidate their mechanisms and potential as probiotics.