Influence of dietary Bacillus coagulans and/or Bacillus licheniformis-based probiotics on performance, gut health, gene expression, and litter quality of broiler chickens.

Probiotics are non-pathogenic microorganisms that are potentially important non-antibiotic alternatives. This study aimed to compare novel multi-strain and single-strain Bacillus probiotics and their respective influences on broiler chickens' performance, gut health, litter quality, immune response, and NBN and TLR gene expression. A total of 1200 Arbor-Acres 1-day-old broiler chicks were randomly allocated into three treatments (T1 was a control, T2 was supplemented with a combined Bacillus coagulans (2 × 109 cfu/g) and Bacillus licheniformis (8 × 109 cfu/g) probiotic strains (0.2 kg/ton of feed), and T3 was supplemented with Bacillus licheniformis (3.2 × 109 cfu/g) probiotic (0.5 kg/ton of feed) with eight replicas of each. Supplementing the broiler diet with either the single-strain (T3) or the multi-strain (T2) Bacillus-based probiotic raised the overall birds' body weight, body weight gain, feed conversion ratio, and European production efficiency factor compared to the control (T1), with a significant enhancement achieved by the multi-strain Bacillus product (P = 0.005). T2 and T3 exhibited significantly improved cholesterol, Alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase, and alkaline phosphatase levels than the control (P ≤ 0.05). The transcript levels of both NBN and TLR genes were upregulated in the liver in the T2 and T3 groups. The T2 group experienced significant reductions in gut bacterial counts, especially for Clostridia, and recorded the lowest litter moisture and nitrogen. In conclusion, supplementing broiler diets with probiotics of multiple Bacillus strains increased production profitability by promoting bird growth, improving feed intake, enhancing gut mucosa and immune organs, and upregulating genes responsible for immunity. All these inhibit the overgrowth of enteric pathogens and sustain litter quality.

Multistep Metabolic Engineering of Bacillus licheniformis To Improve Pulcherriminic Acid Production.

The cyclodipeptide pulcherriminic acid, produced by Bacillus licheniformis, is derived from cyclo(l-Leu-l-Leu) and possesses excellent antibacterial activities. In this study, we achieved the high-level production of pulcherriminic acid via multistep metabolic engineering of B. licheniformis DWc9n*. First, we increased leucine (Leu) supply by overexpressing the ilvBHC-leuABCD operon and ilvD, involved in Leu biosynthesis, to obtain strain W1, and the engineered strain W2 was further attained by the deletion of gene bkdAB, encoding a branched-chain α-keto acid dehydrogenase in W1. As a result, the intracellular Leu content and pulcherriminic acid yield of W2 reached 147.4 mg/g DCW (dry cell weight) and 189.9 mg/liter, which were 227.6% and 48.9% higher than those of DWc9n*, respectively. Second, strain W3 was constructed through overexpressing the leucyl-tRNA synthase gene leuS in W2, and it produced 367.7 mg/liter pulcherriminic acid. Third, the original promoter of the pulcherriminic acid synthetase cluster yvmC-cypX in W3 was replaced with a proven strong promoter, PbacA, to produce the strain W4, and its pulcherriminic acid yield was increased to 507.4 mg/liter. Finally, pulcherriminic acid secretion was strengthened via overexpressing the transporter gene yvmA in W4, resulting in the W4/pHY-yvmA strain, which yielded 556.1 mg/liter pulcherriminic acid, increased by 337.8% compared to DWc9n*, which is currently the highest pulcherriminic acid yield to the best of our knowledge. Taken together, we provided an efficient strategy for enhancing pulcherriminic acid production, which could apply to the high-level production of other cyclodipeptides.IMPORTANCE Pulcherriminic acid is a cyclodipeptide derived from cyclo(l-Leu-l-Leu), which shares the same iron chelation group with hydroxamate sidephores. Generally, pulcherriminic acid-producing strains could be the perfect candidates for antibacterial and anti-plant-pathogenic fungal agents. In this study, we obtained the promising W4/pHY-yvmA pulcherriminic acid-producing strain via a multistep metabolic modification. The engineered W4/pHY-yvmA strain is able to achieve 556.1 mg/liter pulcherriminic acid production, which is the highest yield so far to the best of our knowledge.

Comparative effects of Bacillus subtilis and Bacillus licheniformis on live performance, blood metabolites and intestinal features in broiler inoculated with Salmonella infection during the finisher phase.

Free of Salmonella infection, a total of 300 broiler chicks (Ross 308) were randomly allotted to six dietary treatments (10 replicates) as follows: Negative control (only the basal diet); positive control (infected only); T1, infection + antibiotic (avilamycin); T2, infection + Bacillus subtilis and T3, Salmonella infection + Bacillus licheniformis. The results revealed that production performance was severely affected in the infected group. Also the supplementation of Bacillus subtilis (T2) significantly (P < 0.01) improved feed intake, body weight and performance efficiency factor as compared to the positive control. In addition, feed conversion ratio was significantly (P < 0.01) improved in T2 and T3 compared to the positive control. The results of intestinal health showed that significantly (P < 0.01) higher villus height and total surface area were found in T2 compared to positive control. The results of blood cholesterol, glucose, globulin and total protein concentration were significantly (P < 0.05) higher in T3 compared to the infected birds (positive control). It was concluded that Bacillus subtilis produced superior results in comparison with Bacillus licheniformis in term of growth and intestinal features in broiler by mitigating the deleterious effects of Salmonella infection.

Bacillus subtilis and Bacillus licheniformis reduce faecal protein catabolites concentration and odour in dogs.

BACKGROUND: Direct-fed microbials (DFM), such as Bacillus subtilis and Bacillus licheniformis, may improve gut functionality of the host by favouring non-pathogenic bacteria and reducing the formation of putrefactive compounds. The aim of this study was to assess the nutrient digestibility, faecal characteristics and intestinal-fermentation products in dogs fed diets with Bacillus subtilis and Bacillus licheniformis. Sixteen dogs were randomly divided into two groups. Every eight dogs were fed with the control diet or the diet with the addition of 62.5 g of DFM (B. subtilis and B. licheniformis)/ton. Diets were provided throughout a 20-day adaptation period, followed by 5 days of total faecal collection. Nutrient digestibility and the metabolisable energy of the diets, plus the dogs' faecal characteristics and intestinal fermentation products were assessed. RESULTS: There were no differences in nutrient digestibility (P > 0.05). However, DFM supplementation improved faecal score and resulted in less fetid faeces (P < 0.001). DFM inclusion reduced (P < 0.05) the biogenic amines concentration: putrescine, spermidine and cadaverine, besides the concentration of phenols and quinoline. CONCLUSIONS: The use of B. subtillis and B. licheniformis as DFM reduce the concentration of nitrogen fermentation products in faeces and faecal odour, but the digestibility of nutrients is not altered in dogs.

Crystal structure of NucB, a biofilm-degrading endonuclease.

Bacterial biofilms are a complex architecture of cells that grow on moist interfaces, and are held together by a molecular glue of extracellular proteins, sugars and nucleic acids. Biofilms are particularly problematic in human healthcare as they can coat medical implants and are thus a potential source of disease. The enzymatic dispersal of biofilms is increasingly being developed as a new strategy to treat this problem. Here, we have characterized NucB, a biofilm-dispersing nuclease from a marine strain of Bacillus licheniformis, and present its crystal structure together with the biochemistry and a mutational analysis required to confirm its active site. Taken together, these data support the categorization of NucB into a unique subfamily of the ßßα metal-dependent non-specific endonucleases. Understanding the structure and function of NucB will facilitate its future development into an anti-biofilm therapeutic agent.

Induced resistance in nectarine fruit by Bacillus licheniformis W10 for the control of brown rot caused by Monilinia fructicola.

Brown rot caused by Monilinia fructicola has led to considerable preharvest and postharvest losses in all major nectarine fruit-growing areas. In our previous study, we successfully identified a biocontrol strain of bacteria, Bacillus licheniformis W10, that can be used to control brown rot. However, the possible mechanism of the control of brown rot by B. licheniformis W10 is still unclear. Therefore, the objectives of this study were to determine whether B. licheniformis W10 induces resistance by activating defense-related enzymes including antioxidant enzymes in nectarine. Treatment of nectarine fruit with B. licheniformis W10 reduced both M. fructicola-induced oxidative damage and reactive oxygen species (ROS) production. Furthermore, application of B. licheniformis to nectarine fruit resulted in a significant increase in the activity of antioxidant and defense-related enzymes and increase in the expression of the corresponding genes. Overall, our results verified the proposed mechanism of B. licheniformis W10 in controlling M. fructicola via regulation of ROS levels and activation of antioxidant and defense-related enzymes.

The Biological Threat: The Threat of Planetary Quarantine Failure as a Result of Outer Space Exploration by Humans.

The paper presents the results of experiments with spore-forming bacteria and microscopic fungi performed in the framework of the Russian Research Program outside the International Space Station. It has been found that microorganisms not only survive in this extreme environment, but also retain reproductive ability. Moreover, most microorganisms exhibit an increase in biochemical activity and resistance to antimicrobial agents, specifically antibiotics. These findings are of obvious interest to the developers of both planetary quarantine methods and biomedical safety systems for manned space exploration missions. In addition, they demonstrate the necessity of experiments on the exposure of bio-objects to simulated environmental factors beyond Earth's magnetosphere.

Microbiological and real-time mechanical analysis of Bacillus licheniformis and Pseudomonas fluorescens dual-species biofilm.

In natural habitats, bacterial species often coexist in biofilms. They interact in synergetic or antagonistic ways and their interactions can influence the biofilm development and properties. Still, very little is known about how the coexistence of multiple organisms impact the multispecies biofilm properties. In this study, we examined the behaviour of a dual-species biofilm at the air-liquid interface composed by two environmental bacteria: Bacillus licheniformis and a phenazine mutant of Pseudomonas fluorescens. Study of the planktonic and biofilm growths for each species revealed that P. fluorescens grew faster than B. licheniformis and no bactericidal effect from P. fluorescens was detected, suggesting that the growth kinetics could be the main factor in the dual-species biofilm composition. To validate this hypothesis, the single- and dual-species biofilm were characterized by biomass quantification, microscopy and rheology. Bacterial counts and microscale architecture analysis showed that both bacterial populations coexist in the mature pellicle, with a dominance of P. fluorescens. Real-time measurement of the dual-species biofilms' viscoelastic (i.e. mechanical) properties using interfacial rheology confirmed that P. fluorescens was the main contributor of the biofilm properties. Evaluation of the dual-species pellicle viscoelasticity at longer time revealed that the biofilm, after reaching a first equilibrium, created a stronger and more cohesive network. Interfacial rheology proves to be a unique quantitative technique, which combined with microscale imaging, contributes to the understanding of the time-dependent properties within a polymicrobial community at various stages of biofilm development. This work demonstrates the importance of growth kinetics in the bacteria competition for the interface in a model dual-species biofilm.

Dietary probiotics have different effects on the composition of fecal microbiota in farmed raccoon dog (Nyctereutes procyonoides) and silver fox (Vulpes vulpes fulva).

BACKGROUND: The abuse of antibiotics in animal husbandry imposes a serious threat to both animal health and the environment. As a replacement for antibiotics, probiotic products have been widely used in livestock farming to promote growth of animals. However, no products specifically developed for farmed raccoon dogs and foxes are commercially available at the moment. This study was conducted to investigate the effects of mixed probiotics on farmed raccoon dogs and foxes. RESULTS: Two feeding trials on farmed raccoon dogs and foxes were performed. A mixed probiotic preparation composed of Bifidobacterium bifidum, Clostridium butyricum, Bacillus subtilis and Bacillus licheniformis was fed to these two canine species in order to assess whether such a mixed probiotics can be an alternative to antibiotics (control group). The body weight of raccoon dogs exhibited an increasing tendency with mixed probiotics administration, while that of foxes did not. The serum antioxidant activity was evaluated, and a significantly increase of total antioxidative capacity (T-AOC) was observed in both species. Illumina MiSeq was used for the sequencing of 16S rRNA genes to compare the composition of fecal microbiota between the control and mixed probiotics groups. Although α-diversity did not change, ß-diversity of the fecal microbiota showed a distinct dissimilarity between the control and probiotics groups of both raccoon dogs and foxes. Dietary mixed probiotics increased the abundance of the genus Bifidobacterium in the fecal samples of raccoon dogs, and the genus Bacillus in the fecal samples of foxes. The different responses of raccoon dogs and foxes to probiotics might be the result of differences in the composition of the native gut microbiota of the two species. CONCLUSIONS: The mixed probiotics preparation composed of Bifidobacterium bifidum, Clostridium butyricum, Bacillus subtilis and Bacillus licheniformis could be an effective feed additive for the improvement of the health of farmed raccoon dogs, but it may not be suitable for foxes.

Bacillus licheniformis, a potential probiotic, inhibits obesity by modulating colonic microflora in C57BL/6J mice model.

AIMS: This study evaluated the effects of a potential probiotic, Bacillus sp., on the growth, serum and hepatic triglyceride, histological features of liver tissues and colonic microflora in high-fat diet-induced obese mice. METHODS AND RESULTS: Sixty male C57BL/6J mice were randomly divided into five groups: mice fed a low-fat diet (Cont), mice fed a high-fat diet (Hf), Hf and orally challenged with Bacillus subtilis (Bs), B. licheniformis (Bl) and a mixture of B. subtilis and B. licheniformis (Bls). Gavage feeding was provided at week 9 and the experiment was continued for 8 weeks. Treatment with B. licheniformis and a mixture of Bacillus sp. attenuated body weight gain at the end of study and enhanced glucose tolerance by sensitizing insulin action in the Hf-fed mice. Lower serum and hepatic triglyceride and epididymal fat weight were observed in Bl and Bls groups than that of Hf group. Lesser hepatic fat deposition was observed in the Bl and Bls groups than in the Hf group. High-throughput sequencing showed that Bacillus sp. supplementation dramatically changed the colonic bacterial community in obese mice. CONCLUSIONS: Bacillus licheniformis reduced body weight and improved glucose tolerance, obesity and insulin resistance in Hf-fed mice by changing colonic microbiota composition. SIGNIFICANCE AND IMPACT OF THE STUDY: Orally administration of Bacillus licheniformis may reduce body weight and decrease fat deposition by modulating colonic bacterial community in Hf model.

Sporulating behavior of Bacillus licheniformis strains influences their population dynamics during raw milk holding.

To understand the role of strain variability, population dynamics of 2 strains of Bacillus licheniformis, ATCC 6634 and ATCC 14580, were modeled as a function of temperature (4.0-12.0°C) and duration (0-72 h) using regression analysis. Based on the initial spiking of vegetative cells (approximately 4.0 log cfu/mL) and spores (approximately 2.0 log cfu/mL), regression equations, elucidating B. licheniformis growth behavior during raw milk holding at low temperature, were obtained. Contour plots were developed to determine the time-temperature combinations, keeping the population changes to less than 1.0 log. In vegetative cell spiking study of B. licheniformis ATCC 6634 (S1), cell population changes remained below 1.0 log up to 72 h at 8°C. For B. licheniformis ATCC 14580 (S2), 1.0 log shift was not observed only after 80 h at 8°C, indicating higher multiplication potential of S1 as compared with S2. As S2 was a readily sporulating strain, the vegetative spiking study showed spore formation at different storage temperatures. Evidence of some parallel germination was observed for this strain at 8°C or higher, when raw milk samples were spiked with spores. The experimental values obtained for sporeformers and spore counts were validated with contour plot-generated values. Overall, for raw milk samples predominated by the low sporulating strain, the contour plots suggested holding at 8°C or below for up to 72 h. In the case of the readily sporulating strain (S2), raw milk could be held at 8°C for 80 h, where little or no sporulation is observed. Sporulation behavior, germination and multiplication ability, strain variability, and temperature and duration of holding raw milk influenced the population dynamics of Bacillus species. However, in the presence of equivalent numbers of both types of sporulating strains in raw milk, despite strain variability, holding milk at 8°C for not more than 72 h would keep any cell population changes below 1.0 log. In addition, under these storage conditions, the population would remain as vegetative cells that are likely to be inactivated by pasteurization. The contour plots, so generated, would help predict the population shifts and define optimum holding conditions for raw milk before further processing.

Bacillus subtilis B21 and Bacillus licheniformis B26 improve intestinal health and performance of broiler chickens with Clostridium perfringens-induced necrotic enteritis.

This study investigated the influence of Bacillus-based probiotics on performance and intestinal health in broiler challenged with Clostridium perfringens-induced necrotic enteritis. One-day-old Arbor Acre (n = 480) were randomly assigned to four treatments with 10 cages of 12 birds: (a) basal diet negative control (NC), with no probiotics nor antibiotics formulated to contain 2,930 and 3,060 kcal/kg with 24.07 and 15.98% CP, for starter and finisher diet, respectively, (b) basal diet + enramycin (5 mg/kg), an antibiotic growth promoter (AGP); (c) basal diet + Bacillus subtilis B21 at 2 × 109 CFU per g (BS); (d) basal diet + Bacillus licheniformis B26 at 2 × 109 CFU per g (BL); growth performance, intestinal morphology, intestinal lesion scores, short-chain fatty acids (SCFAs) and mucosal barrier tight junction's (TJ) mRNA expression were assessed. NC- and BL-fed groups showed higher (p = 0.005) average daily feed intake from d1 to d21 than AGP and BS, whereas BS- and AGP-fed groups showed higher average daily weight gain from d22 to d42 and d1 to d42 of age. Higher mortality rate of (12.5%) and lower of (5.5%) were recorded in AGP and NC fed-groups respectively, lesion score was higher in BS and BL than in AGP, while no lesion was observed in NC group, results revealed higher duodenum and jejunum villus height to crypt depth (VH:CD) compared with NC and BS. Probiotics-fed groups showed higher total (SCFAs), acetic and butyric acid concentrations at d21 post-challenge (PC) than other groups. The expression of claudin-1 was upregulated in duodenum (d7) PC and in jejunum (d7) and (d21) PC in BL group, while at d21 PC, the expression of occludens was higher in jejunum and ileum by AGP and BL. The present study indicated both BS and BL have some similarity with AGP in preventing or partially preventing NE effect in broilers.

Biocontrol traits of Bacillus licheniformis GL174, a culturable endophyte of Vitis vinifera cv. Glera.

BACKGROUND: Bacillus licheniformis GL174 is a culturable endophytic strain isolated from Vitis vinifera cultivar Glera, the grapevine mainly cultivated for the Prosecco wine production. This strain was previously demonstrated to possess some specific plant growth promoting traits but its endophytic attitude and its role in biocontrol was only partially explored. In this study, the potential biocontrol action of the strain was investigated in vitro and in vivo and, by genome sequence analyses, putative functions involved in biocontrol and plant-bacteria interaction were assessed. RESULTS: Firstly, to confirm the endophytic behavior of the strain, its ability to colonize grapevine tissues was demonstrated and its biocontrol properties were analyzed. Antagonism test results showed that the strain could reduce and inhibit the mycelium growth of diverse plant pathogens in vitro and in vivo. The strain was demonstrated to produce different molecules of the lipopeptide class; moreover, its genome was sequenced, and analysis of the sequences revealed the presence of many protein-coding genes involved in the biocontrol process, such as transporters, plant-cell lytic enzymes, siderophores and other secondary metabolites. CONCLUSIONS: This step-by-step analysis shows that Bacillus licheniformis GL174 may be a good biocontrol agent candidate, and describes some distinguished traits and possible key elements involved in this process. The use of this strain could potentially help grapevine plants to cope with pathogen attacks and reduce the amount of chemicals used in the vineyard.

In vitro antagonistic activity and the protective effect of probiotic Bacillus licheniformis Dahb1 in zebrafish challenged with GFP tagged Vibrio parahaemolyticus Dahv2.

In vitro antagonistic activity and the protective effect of probiotic Bacillus licheniformis Dahb1 in zebrafish (Danio rerio) challenged with GFP tagged Vibrio parahaemolyticus Dahv2 was studied. The cell free extract of probiotic B. licheniformis Dahb1 at 100 µg mL-1 showed growth inhibition of V. parahaemolyticus Dahv2 in vitro. B. licheniformis Dahb1 also inhibited the biofilm growth of GFP tagged V. parahaemolyticus Dahv2 at 100 µg mL-1 in vitro. The growth and survival of zebrafish was tested using probiotic B. licheniformis Dahb1. Weight (1.28 g) of zebrafish that received the cell free extract was much higher than in control (1.04 g). The mortality of zebrafish infected with GFP tagged V. parahaemolyticus Dahv2 at 107 Cfu mL-1 (Group IV) was 100%, whereas a complete survival of zebrafish that received the cell free extract of B. licheniformis Dahb1 at 107 Cfu mL-1 (Group VII) was observed after 30 days. The number of GFP tagged V. parahaemolyticus Dahv2 colonies in the intestine and gills significantly reduced after treatment with the cell free extract of B. licheniformis Dahb1. Furthermore, a significant decrease in the fluorescent colonies of GFP tagged V. parahaemolyticus Dahv2 was observed after treatment with the cell free extract of B. licheniformis Dahb1 under confocal laser scanning microscopy (CLSM). In conclusion, the cell free extract of B. licheniformis Dahb1 could prevent Vibrio infection by enhancing the growth and survival of zebrafish.

In situ analysis of Bacillus licheniformis biofilms: amyloid-like polymers and eDNA are involved in the adherence and aggregation of the extracellular matrix.

AIMS: This study attempts to determine which of the exopolymeric substances are involved in the adherence and aggregation of a Bacillus licheniformis biofilm. METHODS AND RESULTS: The involvement of extracellular proteins and eDNA were particularly investigated using DNase and proteinase K treatment. The permeability of the biofilms increased fivefold after DNase I treatment. The quantification of the matrix components showed that, irrespective to the enzyme tested, eDNA and amyloid-like polymers were removed simultaneously. Size-exclusion chromatography analyses supported these observations and revealed the presence of associated nucleic acid and protein complexes in the biofilm lysates. These data suggest that some extracellular DNA and amyloid-like proteins were closely interlaced within the matrix. Finally, confocal laser scanning microscopy imaging gave supplementary clues about the 3D organization of the biofilms, confirming that eDNA and exoproteins were essentially layered under and around the bacterial cells, whereas the amyloid-like fractions were homogeneously distributed within the matrix. CONCLUSION: These results confirm that some DNA-amyloid complexes play a key role in the modulation of the mechanical resistance of B. licheniformis biofilms. SIGNIFICANCE AND IMPACT OF THE STUDY: The study highlights the need to consider the whole structure of biofilms and to target the interactions between matrix components. A better understanding of B. licheniformis biofilm physiology and the structural organization of the matrix will strengthen strategies of biofilm control.

Probiotics and Probiotic Metabolic Product Improved Intestinal Function and Ameliorated LPS-Induced Injury in Rats.

In the present study, we sought to determine the effects of Bacillus subtilis (BAS) and Bacillus licheniformis (BAL) in rats after lipopolysaccharide (LPS)-induced acute intestinal inflammation. We also determined whether the B. subtilis metabolic product (BASM) is as effective as the live-cell probiotic. 60 male SD rats were randomly assigned to five groups and administered a diet containing 0.05% B. licheniformis (BAL group), 0.05% B. subtilis (BAS group), 0.5% B. subtilis metabolic product (BASM group), or a basic diet (PC group and NC group) for 40 days. On day 40, BAL, BAS, BASM, and NC groups were injected with 4 mg/kg body weight LPS. 4 h later, all rats were anesthetized and sacrificed. The results showed that the administration of B. licheniformis and B. subtilis improved intestinal function as evidenced by histology, increased enzyme activity, and mucosal thickness. They also increased the number of intraepithelial lymphocytes and decreased mucosal myeloperoxidase activity and plasma TNF-α. In addition, the cecal content of B. subtilis-treated rats had significantly increased microbial diversity, decreased numbers of Firmicutes, and increased numbers of Bacteroidetes as compared to rats fed basic diets. Similar to BAS group, the cecal content of B. licheniformis-treated rats decreased the number of Firmicutes. Administration of B. subtilis metabolic product had similar effects on intestinal function, inflammation response, and microbial diversity as B. subtilis but these effects were attenuated. In conclusion, administration of probiotic strains B. licheniformis or B. subtilis improved intestinal function, ameliorated the inflammation response, and modulated microflora after LPS-induced acute inflammation in rats. Non-living cells also exerted probiotic properties but live cells tended to function better.

Rhamnolipids from Pseudomonas aeruginosa strain W10; as antibiofilm/antibiofouling products for metal protection.

Industrial biofouling-problems associated with the accumulation of microorganisms from flowing water and fluids on processing surfaces can cause severe problems. A Pseudomonas aeruginosa strain W10 was isolated from industrial setting and found to produce predominantly di-rhamnolipids (Rha-Rha-C10-C10) with a yield of around 10 g L-1 and a critical micelle concentration (CMC) of 80 mg L-1 . P. aeruginosa W10 rhamnolipids were able to disrupt up to 99% of 48 h pre-formed biofilms of the Gram-positive organisms Bacillus licheniformis CAN55, Staphylococcus capitis SH6, and a mixed culture (strains CAN55, SH6, and W10), under static conditions, at concentrations of 0.1, 0.5, and 1 mg ml-1 on a stainless steel surface commonly used in industrial process pipelines. CFU measurements and LIVE/DEAD BacLight staining confirmed these observations. Furthermore, a purified di-rhamnolipid fraction was found to be responsible for the microbial inhibition of B. licheniformis strain CAN55. This study provides evidence that rhamnolipids may have valuable applications in preventing biofilms and biofouling in industrial plants and, in a wider context, may also apply to metal medical devices.

GFP tagged Vibrio parahaemolyticus Dahv2 infection and the protective effects of the probiotic Bacillus licheniformis Dahb1 on the growth, immune and antioxidant responses in Pangasius hypophthalmus.

In this study, the pathogenicity of GFP tagged Vibrio parahaemolyticus Dahv2 and the protective effect of the probiotic strain, Bacillus licheniformis Dahb1 was studied on the Asian catfish, Pangasius hypophthalmus. The experiment was carried out for 24 days with three groups and one group served as the control (without treatment). In the first group, P. hypophthalmus was orally infected with 1 mL of GFP tagged V. parahaemolyticus Dahv2 at two different doses (10(5) and 10(7) cfu mL(-1)). In the second group, P. hypophthalmus was orally administrated with 1 ml of the probiotic B. licheniformis Dahb1 at two different doses (10(5) and 10(7) cfu mL(-1)). In the third group, P. hypophthalmus was orally infected first with 1 mL of GFP tagged V. parahaemolyticus Dahv2 followed by the administration of 1 mL of B. licheniformis Dahb1 (combined treatment) at two different doses (10(5) and 10(7) cfu mL(-1)). The growth, immune (myeloperoxidase, respiratory burst, natural complement haemolytic and lysozyme activity) and antioxidant (glutathione-S-transferase, reduced glutathione and total glutathione) responses of P. hypophthalmus were reduced after post infection of GFP tagged V. parahaemolyticus Dahv2 compared to control. However, after administration with the probiotic B. licheniformis Dahb1 at 10(5) cfu mL(-1), P. hypophthalmus showed significant increase in the growth, immune and antioxidant responses compared to 10(7) cfu mL(-1). On the otherhand, the growth, immune and antioxidant responses of P. hypophthalmus infected and administrated with combined GFP tagged Vibrio + Bacillus at 10(5) cfu mL(-1) were relatively higher than that of GFP tagged V. parahaemolyticus Dahv2 and control groups but lower than that of probiotic B. licheniformis Dahb1 groups. The results of the present study conclude that the probiotic B. licheniformis Dahb1 at 10(5) cfu mL(-1) has the potential to protect the P. hypophthalmus against V. parahaemolyticus Dahv2 infection by enhancing the growth, immune and antioxidant responses. The probiotic B. licheniformis Dahb1 would be effectively used in the treatment of aquatic diseases for improvement of aquaculture industry.

Effects of Bacillus licheniformis on the growth performance and expression of lipid metabolism-related genes in broiler chickens challenged with Clostridium perfringens-induced necrotic enteritis.

BACKGROUND: Necrotic enteritis (NE), caused by Clostridium perfringens, has cost the poultry industry $2 billion in losses. This study aimed to investigate the effect of Bacillus licheniformis as dietary supplement on the growth, serum antioxidant status, and expression of lipid-metabolism genes of broiler chickens with C. perfringens-induced NE. METHODS: A total of 240 one-day-old broilers were randomly grouped into four: a negative control, an NE experimental model (PC), chickens fed a diet supplemented with 30 % of fishmeal from day 14 onwards and challenged with coccidiosis vaccine (FC), and NE group supplied with feed containing 1.0 × 10(6) CFU/g B. licheniformis (BL). RESULTS: Body weight gain, feed conversion ratio, serum antioxidant status, and lipid-metabolism-gene expression were analyzed. In the PC group, FCR increased significantly whereas serum catalase and glutathione peroxidase activity decreased compared with NC group. Dietary B. licheniformis supplementation improved FCR and oxidative stress in experimental avian NE. Using Bacillus licheniformis as a direct-fed microbial (DFM) could also significantly upregulate catabolism-related genes, namely, peroxisome proliferator-activated receptor-α and carnitine palmitoyltransferase-1, in livers and changed the expression of lipid-anabolism genes. CONCLUSION: These results suggested that dietary B. licheniformis supplementation can enhance growth and antioxidant ability, as well as change the expression of genes related to fatty-acid synthesis and oxidation in the livers of NE-infected broilers.

Dietary Bacillus licheniformis improves the effect of Astragalus membranaceus extract on blood glucose by regulating antioxidation activity and intestinal microbiota in InR[E19]/TM2 Drosophila melanogaster.

BACKGROUND: The diabetes mellitus prevalence is rapidly increasing in most parts of the world and has become a vital health problem. Probiotic and herbal foods are valuable in the treatment of diabetes. METHODS AND PERFORMANCE: In this study, Bacillus licheniformis (BL) and Astragalus membranaceus extract (AE) were given with food to InR[E19]/TM2 Drosophila melanogaster, and the blood glucose, antioxidation activity and intestinal microbiota were investigated. The obtained results showed that BA (BL and AE combination) supplementation markedly decreased the blood glucose concentration compared with the standard diet control group, accompanied by significantly increased enzymatic activities of catalase (CAT), decreased MDA levels and prolonged lifespan of InR[E19]/TM2 D. melanogaster. The treatments with BL, AE and BA also ameliorated intestinal microbiota equilibrium by increasing the population of Lactobacillus and significantly decreasing the abundance of Wolbachia. In addition, clearly different evolutionary clusters were found among the control, BL, AE and BA-supplemented diets, and the beneficial microbiota, Lactobacillaceae and Acetobacter, were found to be significantly increased in male flies that were fed BA. These results indicated that dietary supplementation with AE combined with BL not only decreased blood glucose but also extended the lifespan, with CAT increasing, MDA decreasing, and intestinal microbiota improving in InR[E19]/TM2 D. melanogaster. CONCLUSION: The obtained results showed that dietary supplementation with BL and AE, under the synergistic effect of BL and AE, not only prolonged the lifespan of InR[E19]/TM2 D. melanogaster, increased body weight, and improved the body's antiaging enzyme activity but also effectively improved the types and quantities of beneficial bacteria in the intestinal flora of InR[E19]/TM2 D. melanogaster to improve the characteristics of diabetes symptoms. This study provides scientific evidence for a safe and effective dietary therapeutic method for diabetes mellitus.