Comparison of Antibacterial Activities of Six Bacillus amyloliquefaciens Strains

In this study, six bacterial strains were isolated from the sediment, probiotic fermentation products, and lake sediments, they were identified as Bacillus amyloliquefaciens using genetic evolution analysis, which were named B3, B4, B5, XD3, YF6, and YF8. The comparison of the antibacterial activity, hemolytic activity, and antibiotic sensitivity of six Bacillus amyloliquefaciens strains laid a foundation for the development and application of antimicrobial peptide products. A surface activity assay was used to determine the production of biosurfactants in six Bacillus amyloliquefaciens strains. With Staphylococcus aureus and Escherichia coli as indicator bacteria, their antibacterial activity was determined using the agar diffusion method;the same diffusion method was used to determine the antibiotic susceptibility of Bacillus amyloliquefaciens. The results showed that the six Bacillus amyloliquefaciens strains had obvious biosurface activity, and the bacteria inhibited Staphylococcus aureus and Escherichia coli, from strong to weak: YF8, XD3, B3, B4, YF6, and B5. Strain YF8 had the best broad-spectrum bacteriostatic effect, followed by strain XD3. All Bacillus amyloliquefaciens strains were susceptible to 16 common drugs, except for Bacillus amyloliquefaciens strain YF8, which was intermediate to neomycin. The study shows that Bacillus amyloliquefaciens and secondary metabolites have the ability to produce a variety of active peptides, exert a certain inhibitory effect on common pathogens, and have the potential to develop as animal probiotics.

基于代谢组学和转录组学揭示郫县豆瓣中两株优势产风味菌株Bacillus amyloliquefaciens与Candida versatilis互作机制

为揭示郫县豆瓣内源性优势菌株Bacillus amyloliquefaciens与Candida versatilis的互作机制,文章基于非靶向代谢组学和转录组学技术,探究了两株菌在单独培养和共培养体系下代谢物和基因表达水平的差异。结果表明,与单独培养B.amyloliquefaciens与C.versatilis相比,共培养体系下分别有193种和297种显著差异代谢物,集中在脂类和类脂分子、有机酸及其衍生物、氨基酸及其衍生物等。通过转录组学和差异代谢通路分析,B.amyloliquefaciens在共培养体系下上调表达基因801个,集中在生物过程中;而下调表达基因805个,以分子功能为主。C.versatilis对B.amyloliquefaciens的生长代谢具有双向作用,其中糖酵解/糖异生、脂肪酸降解等途径显著增强,而脂肪酸合成、氨基酸相关代谢显著减弱。研究结果可揭示两株菌的互作机制,并为改善郫县豆瓣风味及品质提供理论指导。

Modulations of genes related to gut integrity,apoptosis,and immunity underlie the beneficial effects of Bacillus amyloliquefaciens CECT 5940 in broilers fed diets with different protein levels in a necrotic enteritis challenge model

Background:The ban of in-feed antimicrobial additives has negatively affected the poultry industry by causing necrotic enteritis(NE)to emerge in the flocks.Alternatives such as Bacillus probiotics have shown to be effective on eliminating the negative effects of this disease.Two experiments were conducted to investigate the effect of Bacillus amyloliquefaciens CECT 5940(BA)in broiler chickens under NE challenge and/or fed diets with different protein levels.Methods:In both experiments,480 day-old mix-sexed Ross-308 broilers were arranged in a 2×2 factorial arrangement of treatments.In experiment 1,the factors were NE challenge(yes or no)and probiotic(yes or no).In experiment 2,the factors were dietary crude protein levels(standard or reduced)and probiotic(yes or no)and were used under NE challenge condition.Oral administration of Eimeria oocysts(day 9)followed by inoculation with Clostridium perfringens(day 14 and 15)was used to induce NE challenge.On day 16,two birds from each treatment were gavaged with fluorescein isothiocyanate-dextran(FITC-d)and blood samples were collected for gut integrity evaluation,and jejunal samples were collected for gene expression assay.Results:In experiment 1,BA supplementation decreased caspase-3(CASP3)(P<0.001)and caspase-8(CASP8)(P<0.05)and increased occludin(OCLD)(P<0.05)expression regardless of the challenge.Additionally,BA supplementation downregulated interfron-γ(IFN-γ)expression(P<0.01)and upregulated immunoglobulin-G(IgG)(P<0.01)and immunoglobulin-M(IgM)(P<0.05)only in challenged birds.In experiment 2,the expression of genes encoding mucin-2(MUC2)(P<0.001),tight junction protein-1(TJP1)(P<0.05)and OCLD(P<0.05)were upregulated by the addition of BA in the diet,regardless of the crude protein level.Further,BA supplementation downregulated INF-γ(P<0.01)and upregulated immunoglobulin-A(IgA)(P<0.05),IgM(P<0.05)and IgG(P<0.01)regardless of the crude protein level.Conclusion:These findings suggest that supplementation of BA in broiler diets can improve gut health by modulation of genes related to the mucosal barrier,tight junction,and immunity in broilers challenged by unfavourable conditions such as NE challenge.

Physicochemical Properties and Bioactivities of Rice Beans Fermented by Bacillus amyloliquefaciens

The purpose of this study was to investigate the physicochemical properties and bioactivities of rice beans(Vigna umbellata)fermented by Bacillus amyloliquefaciens.The fermentation conditions were optimized on the basis of the fibrinolytic activity.Under the optimal fermentation conditions,the fibrinolytic activity reached a maximum of 78.0 FU·g^(-1)(4890 IU·g^(-1),fibrin plate method,FU:fibrin degradation unit).The contents of peptides(which increased from 2.1 to 10.9 g per 100 g),total phenolics(from 116.7 to 388.5 mg gallic acid per 100 g),total flavonoids(from 235.5 to 354.3 mg rutin per 100 g),and anthocyanin(from 20.1 to 47.1 mg per 100 g),as well as the superoxide dismutase activity(from 55.3 to 263.6 U·g^(-1))in rice beans were significantly increased after fermentation.The 2,2-diphenyl-1-picrylhydrazyl(DPPH)and 2,2-azinobis(3-ethylbenzothiazoline-6-sulphonic acid)diammonium salt(ABTS)free radical scavenging activities and ferric reducing antioxidant power(FRAP)of fermented rice beans were 1.9–4.8 times higher than those of unfermented rice beans.Moreover,fermentation induced an increase in the dipeptidyl peptidase IV(DPP-IV)inhibition,a-glucosidase inhibition,and anticoagulant activities of rice beans.Rice beans fermented by Bacillus amyloliquefaciens may serve as a functional food with potential benefits for the prevention of thrombotic diseases.

Production,characterization and biological activities of acidic exopolysaccharide from marine Bacillus amyloliquefaciens 3MS 2017

Objective: To evaluate in-vitro antioxidant, anti-inflammatory and antitumor abilities against human breast adenocarcinoma(MCF7) and human prostate cancer(PC3) as well as the suppressor effect of bacterial exopolysaccharide(BAEPS) on Ehrlich ascites carcinoma(EAC). Methods: In-vitro antioxidants characters of BAEPS were determined using various methods, while anti-inflammatory activity was estimated against cyclooxygenase(COX-1 and COX-2). In-vitro study, anticancer against MCF7 and PC3 were assessed by the mitochondrial dependent reduction of yellow MTT. In in-vivo study against EAC progression, mice were inoculated with EAC cells and then were orally administered BAEPS at 200 mg/kg after 24 h(equals to 0.10 of determined LD50)/10 d. Results: BAEPS was acidic exopolysaccharide contained uronic acid(12.3%) and sulfate(22.8%) with constitution of glucose, galactose and glucuronic acid in a molar ratio1.6: 1.0: 0.9, respectively, with a molecular mass of 3.76伊104 g/mol. BAEPS appeared potent antioxidant characters as free radical scavenging, oxygen reactive species scavenging and metal chelation, while its reducing power was low. BAEPS showed selective anti-inflammatory activity against COX-2 than COX-1, COX-2 selective. BAEPS exhibited potent and selective effect to breast cell cancer MCF7, the death percentage was 65.20% with IC_(50)=70 μg/m L and IC_(90)=127.40 μg/m L. BAEPS decreased counted viable EAC cells and induced non-viable cells. BAEPS improved all assessed hematological parameters. These improvements were reflected in the increasing median survival time and significant increment(P<0.05) in life span. Conclusions: BAEPS has anti-tumor activity with a good margin of safety. The anti-tumor activity of BAEPS may be due to its content from sulfated groups and uronic acids and they have antioxidant and anti-inflammatory properties.

生物炭环境协同生防菌Bacillus amyloliquefaciens P4防控香芋软腐病的研究

为挖掘香芋软腐病的生物防治资源,探讨生物炭环境协同解淀粉芽孢杆菌(Bacillus amyloliquefaciens)对香芋软腐病病原菌(Dickeya fangzhongdai)的拮抗作用以及生物防治效果,利用平板对峙法从健康香芋植株中筛选出一株拮抗Dickeya fangzhongdai ZXC1的菌株P4并对其进行分类鉴定;采用竞争性培养的方法验证生防菌的拮抗效果;利用对峙平板法,根据抑菌圈大小明确生物炭是否能够促进生防菌分泌拮抗物质;利用计数统计法测定不同浓度的生物炭环境中致病菌及拮抗菌的数目变化;不同含量生物炭环境下进行盆栽试验,分析生物炭环境与生防菌作用下香芋软腐病病原菌种群数量变化,并统计香芋的发病情况。结果表明,P4菌株对香芋软腐病病原菌具有良好的抑制作用;适量生物炭能够加强生防菌对香芋软腐病的防治效果,并且能直接抑制病原菌致病;生物炭直接刺激生防菌增强其分泌拮抗物质的效果很微弱;生防菌与病原菌的数目随生物炭浓度的上升而增加。P4菌株可作为香芋软腐病的生防菌。加入适量的生物炭能够促进土壤中生防菌生长繁殖并使其占据良好的生态位,加强生防菌对香芋软腐病的生防效果,并且仅加入生物炭也能够抑制病原菌致病。

Evaluation of Plant Growth Promoting Ability of Bacillus amyloliquefaciens Bc2 and Trichoderma harzianum TR In Vivo

Bacillus amyloliquefaciens Bc2 and Trichderma harzianum TR were used to evaluate their growth-promoting activity on cultivated strawberries, under laboratory and field conditions, and we have noticed that the percentage of achene germination is important for ones treated with TR (=97%) followed by those treated with Bc2 strain (=90%) and the control (=84%). Inoculations on field showed that on untreated soil with insecticide, TR is effective and allows the development of plants and extends the duration of flowering and fruiting. On treated soil, Bc2 clearly promotes the growth and development of strawberry seedlings and its role as plant growth promoting microorganisms has been proved.

Improvement of Sporulation Conditions of a New Strain of Bacillus amyloliquefaciens in Liquid Fermentation

The desirable active ingredients for the development of bioproducts based in Bacillus sp. for the control of soil pathogens are the spores because these structures exhibit more resistance and stability to conditions present during the fermentation, formulation, and storage processes. To improve the sporulation of a native strain of Bacillus amyloliquefaciens (Bs006) using liquid fermentation, some modifications in the concentrations of the components in a previously standardized culture media were made. Subsequently, five sporulation inducers (iron nitrate, mixture of salts, peroxide hydrogen, temperature, and initial cell concentration) were evaluated. The treatment with a mixture of salts in combination with an initial cell concentration of 1 × 108 cells/ml was selected because a final spore concentration of 1.05 × 1010 spores/ml after 66 hours with a fully substrate consumption and sporulation efficiency of 88.7% was obtained. To demonstrate the biological activity of B. amyloliquefaciens Bs006 in Cape gooseberry seedlings, a greenhouse bioassay was conducted, and statistical differences in plant growth-promoting parameters compared with previous media were not found. Additionally, the proposed modified media (coded as JM) presented a benefit-cost ratio 2.65 times higher compared with the baseline media.

Biocontrol Potential of Bacillus amyloliquefaciens Bc2 and Trichoderma harzianum TR against Strawberry Anthracnose under Laboratory and Field Conditions

The increasingly strict regulation of the use of phytosanitary products and the will of the populations to move towards sustainable development allow bio-sourced products and more particularly for biocontrol in the field of pesticides to progress. The study carried out here concerns the evaluation of the potency of Bacillus amyloliquefaciens Bc2 and Trichoderma harzianum to fight against strawberry cultures infected with anthracnose (Fragaria × ananassa Duch.). The studies were carried out in the laboratory and in the field. The results indicate that B. amyloliquefaciens Bc2 and Trichoderma harzianum TR are effective for the biological control of anthracnose, gray mold and powdery mildew on strawberries grown in field conditions. The increase in plant size and the number of fruits produced with these control agents has also been observed.

In Vitro Antagonistic Activity of Trichoderma harzianum and Bacillus amyloliquefaciens against Colletotrichum acutatum

Biological control of plant diseases makes extensive use of the antagonists. The effectiveness of the control depends on the choice of an effective antagonist strains from criteria of implying a good knowledge of biological peculiarities of the material used. In this study, the antagonistic activities of nine isolates of Bacillus amyloliquefaciens and one isolate of Trichoderma harzianum were tested in vitro against seven strains of Colletotrichum acutatum, the causal agent of strawberry anthracnose. It was found that T. harzianum and B. amyloliquefaciens strains showed the ability to inhibit the mycelial growth of the pathogen by dual culture technique by more than 50 percent. The nonvolatile substances produced by the antagonists showed high inhibition percentages that are more than 99 percent, meanwhile their volatile compounds inhibited the mycelial growth of C. acutatum strains with low inhibition percentages starting from 30 percent except the substances produced by B. amyloliquefaciens Bc2 that showed a high inhibition percentage of more than 70 percent.

Improvement of Bio-Efficacy to Reduce Bacterial Wilt Complex Disease in Tomatoes through Bacillus amyloliquefaciens and Pseudomonas fluorescens under Field Conditions

Bacterial wilt complex disease of tomato(Solanum lycopersicum L.)was incited jointly by bacterial wilt pathogen Ralstonia solanacearum and Meloidogyne incognita worldwide.Bio-efficacy of bacterial antagonists i.e.B.amyloliquefaciens DSBA-11 and P.fluorescens DTPF-3 was studied against the wilt disease complex in tomato at National Phytotran facility Indian Agricultural Research Institute(IARI),New Delhi,at 26±2°C.Minimum wilt disease incidence(26.00%)with the highest bio-control efficacy(64.15%),less juvenile population(19.33 J2/g of soil)of M.incognita was recorded in the combined application of DTPF-3+DSBA-11 after 30 d of inoculation under glasshouse conditions.In a field study,minimum bacterial wilt disease incidences 19.0%and 20.4%were recorded in the bleaching powder treatment followed by mixed application of DSBA-11+DTPF-3,19.6%and 21.2%wilt incidence in 2014 and 2015 respectively.However,a reduction of root-knot gall index was recorded a maximum of 59.76%and 69.62%in DSBA-11+DTPF-3 treated plants followed by 54.88%and 60.13%over control in DTPF-3 treatment in 2014 and 2015 respectively.The yield of tomato fruit was increased over control by 17.48%and 16.97%in 2014 and 2015 respectively under field conditions.A combination of P.fluorescens DTPF-3+B.amyloliquefaciens DSBA-11 suppressed bacterial wilt and root-knot diseases and also increased the yield of the tomato fruit significantly(p<0.05)under field conditions.

Bacillus amyloliquefaciens 40 regulates piglet performance,antioxidant capacity,immune status and gut microbiota

Probiotics can improve animal growth performance and intestinal health.Bacillus species,Lactobacillus species,Bifidobacterium species,yeast etc.are the common types of probiotics.However,understanding the effects of probiotics on the immune status and gut microbiota of weaning piglets and how the probiotics exert their impact are still limited.This study aimed to investigate the effects of Bacillus amyloliquefaciens 40(BA40)on the performance,immune status and gut microbiota of piglets.A total of 12 litters of newborn piglets were randomly divided into 3 groups.Piglets in control group were orally dosed with phosphate buffered saline;BA40 group and probiotics group were orally gavaged with resuspension BA40 and a probiotics product,respectively.The results showed that BA40 treatment significantly decreased(P<0.05)the diarrhea incidence(from d 5 to 40),diamine oxidase,D-lactate,interleukin(IL)-1βand interferon-γconcentrations compared with control group and probiotics group.Meanwhile BA40 dramatically increased the total antioxidant capacity,IL-10 and secretory immunoglobulin-A concentrations in contrast to control group.For the microbial composition,BA40 modulated the microbiota by improving the abundance of Bacteroides,Phascolarctobacterium(producing short-chain fatty acids)and Desulfovibrio and reducing the proliferation of pathogens(Streptococcus,Tyzzerella,Vellionella and paraeggerthella).Meanwhile,a metabolic function prediction explained that carbohydrate metabolism and amino acid metabolism enriched in BA40 group in contrast to control group and probiotics group.For correlation analysis,the results demonstrated that BA40-enriched Phascolarctobacterium and Desulfovibrio provide insights into strategies for elevating the health status and performance of weaned piglets.Altogether,BA40 exerted stronger ability in decreasing diarrhea incidence and improved antioxidant activity,gut barrier function and immune status of piglets than the other treatments.Our study provided the experimental and theoretical basis for the application of BA40 in pig production.

Metabolic engineering of Bacillus amyloliquefaciens for efficient production ofα-glucosidase inhibitor1-deoxynojirimycin

Owing to the feature of strongα-glucosidase inhibitory activity,1-deoxynojirimycin(1-DNJ)has broad application prospects in areas of functional food,biomedicine,etc.,and this research wants to construct an efficient strain for 1-DNJ production,basing on Bacillus amyloliquefaciens HZ-12.Firstly,using the temperature-sensitive shuttle plasmid T2(2)-Ori,gene ptsG in phosphotransferase system(PTS)was weakened by homologous recombination,and non-PTS pathway was strengthened by deleting its repressor gene iolR,and 1-DNJ yield of resultant strain HZ-S2 was increased by 4.27-fold,reached 110.72 mg/L.Then,to increase precursor fructose-6-phosphate(F-6-P)supply,phosphofructokinase was weaken,fructose phosphatase GlpX and 6-phosphate glucose isomerase Pgi were strengthened by promoter replacement,moreover,regulator gene nanR was deleted,1-DNJ yield was further increased to 267.37 mg/L by 2.41-fold.Subsequently,promoter of 1-DNJ synthetase cluster was optimized,as well as 5′-UTRs of downstream genes in synthetase cluster,and 1-DNJ produced by the final strain reached 478.62 mg/L.Last but not the least,1-DNJ yield of 1632.50 mg/L was attained in 3 L fermenter,which was the highest yield of 1-DNJ reported to date.Taken together,our results demonstrated that metabolic engineering was an effective strategy for 1-DNJ synthesis,this research laid a foundation for industrialization of functional food and drugs based on 1-DNJ.

Accelerating the menaquinone‑7 production in Bacillus amyloliquefaciens by optimization of the biosynthetic pathway and medium components

Menaquinone-7(MK-7)has an important role in preventing diseases such as cardiovascular disease and osteoporosis.In this study,a combination strategy of strain improvement and medium optimization is investigated to increase MK-7 production in Bacillus amyloliquefaciens.Conventional breeding method was first used to modify the biosynthetic pathway to construct a MK-7 high-producing strain by atmospheric and room temperature plasma mutagenesis and protoplast fusion.The resulted strain Ba-4 with resistance to sulfaguanidine,1-hydroxy-2-naphthoic acid,menadione,2-deoxy-d-glucose and rifampicin as well as sensitive toβ-fluoropyruvate produced 73.57±1.61 mg/L of MK-7,which was 1.36 times more than that of the parent strain H.β.D.R.-5(i.e.,31.12±1.40 mg/L).Subsequently,single-factor optimization and response surface method-ology(RSM)were used to optimize the medium components for increasing MK-7 production by strain Ba-4.Strain Ba-4 produced 90.43±1.32 mg/L of MK-7 under the single-factor optimized medium.Moreover,the results of response surface methodology indicated that glycerol,soy peptone and Tween-80 had significant effects on MK-7 production,and the highest MK-7 production(i.e.,95.03±1.01 mg/L)was obtained under the optimized medium,which was 0.29 times higher than that of the initial medium.These results confirmed that the conventional breeding methods and fermenter control system are effective strategies in improving MK-7 production by B.amyloliquefaciens.

Combinatorial mutagenesis of Bacillus amyloliquefaciens for efficient production of protease

As an important industrial enzyme,protease is widely used in feed,food and other fields.At present,the insufficient protease activity obtained from microorganisms cannot meet the purpose of industrial production.In this study,Bacillus amyloliquefaciens with high protease production was screened from animal feces by plate transparent circle method.To improve the production of protease,atmospheric room temperature plasma(ARTP)mutagenesis was used in the first round,protease activity reached 315.0 U/mL.Then,to enhance production of protease,^(60)Co-γ irradiation was used for combined mutagenesis,leading to protease activity of B.amyloliquefaciens FMME ZK003 up to 355.0 U/mL.Furthermore,to realize the efficient production of protease,after optimization of fermentation conditions,protease activity was increased to 456.9 U/mL.Finally,protease activity of B.amyloliquefaciens FMME ZK003 reached 823.0 U/mL in a 5 L fermenter.These results indicate that B.amyloliquefaciens can efficiently produce protease,which provides a good foundation for the industrial production of protease.

Isolation and Characterization of Bacillus Amyloliquefaciens L-1 for Biocontrol of Pear Ring Rot

Considering the heavy losses caused by pear ring rot(Botryosphaeria berengeriana) disease, the potential biocontrol agent against B. berengeriana was isolated and characterized in this work. Bacteria, isolated from pear orchard rhizosphere soil, were screened for the biological control of pear ring rot caused by B. berengeriana. Among them, strain L-1 showed noticeable inhibitory activities against B. berengeriana and six other commonly occurring postharvest pathogens. Molecular methods indicated strain L-1 was Bacillus amyloliquefaciens. The potential of strain L-1 as an effective biocontrol agent was further estimated. Results showed strain L-1 could successfully colonize in pear wounds, its colonies reached 142.35 folds on 4 days post inoculation, and maintained at a high level during storage. In addition, strain L-1 caused abnormal hyphae growth of B. berengeriana,and its inhibitory percentage against pear ring rot reached 76.55% in vivo on 11 days post inoculation. Strain L-1 significantly induced the peroxidase(POD) and catalase(CAT) activities and delayed the accumulation of malondialdehyde(MDA) in pears. What's more, strain L-1 did not impair the fruit quality. All these results suggest that B. berengeriana L-1 is a promising agent for the biocontrol of pear ring rot.

Hepatoprotective ability of tetramethylpyrazine produced by Bacillus amyloliquefaciens

Tetramethylpyrazine(TTMP),an important aroma compound,was produced by Bacillus amyloliquefaciens XJB-104 with distillers’grains as raw material.The yield of TTMP under optimized fermentation procedure was 3176.52 mg/L.TTMP in the fermentation broth was purified and the purity(>99%)was measured by gas chromatography-mass spectrometry.Furthermore,the hepatoprotective activity of TTMP in ethanol-water system was evaluated by biochemical indicators of liver injury,parameters of antioxidant defense system and inflammatory response,and liver histopathological assessment in mice.Ethanol treatment increased serum levels of alanine aminotransferase,aspartate aminotransferase,alkaline phos-phatase and lactate dehydrogenase,suggesting liver damage.Both high and low dose of TTMP significantly decreased levels of these indicators.Furthermore,the reduction in the activities or concentrations of superoxide dismutase,catalase,reduced glutathione and malondialdehyde in liver tissue caused by ethanol was significantly alleviated by TTMP.Increased inflammatory cytokines including transcription factor,tumor necrosis factor,interleukin-1beta,interleukin,macrophage chemoattractant protein,inducible nitric oxide synthase and cyclooxygenase were also suppressed by TTMP treatment.It is concluded that TTMP in ethanol-water system has potential liver-protective activity,especially when ethanol is consumed at low doses for short periods of time.

Bacillus amyloliquefaciens CECT 5940 improves performance and gut function in broilers fed different levels of protein and/or under necrotic enteritis challenge

Two studies were conducted to investigate the effect of Bacillus amyloliquefaciens CECr 5940(BA)as a probiotic on growth performance,amino acid digestibility and bacteria population in broiler chickens under a subclinical necrotic enteritis(NE)challenge and/or fed diets with different levels of crude protein(CP).Both studies consisted of a 2×2 factorial arrangement of treatments with 480 Ross 308 mix-sexed broiler chickens.In study 1,treatments included 1)NE challenge(+/),and 2)BA(1.0×106 CFU/g of feed)supplementation(+/-).In study 2,all birds were under NE challenge,and treatments were 1)CP level(Standard/Reduced[2%less than standard])and 2)BA(1.0×106 CFU/g of feed)supplementation(+/-).After inducing NE infection,blood samples were taken on d 16 for uric acid evaluation,and cecal samples were collected for bacterial enumeration.In both studies,ileal digesta was collected on d 35 for nutrient digestibility evaluation.In study 1,the NE challenge reduced body weight gain(BWG),supressed feed conversion ratio(FCR)and serum uric acid levels(P<0.001).Supplementation of BA increased BWG(P<0.001)and reduced FCR(P=0.043)across dietary treatments,regardless of challenge.Bacillus(P=0.030)and Ruminococcus(P=0.029)genomic DNA copy numbers and concentration of butyrate(P=0.017)were higher in birds fed the diets supplemented with BA.In study 2,reduced protein(RCP)diets decreased BWG(P=0.010)and uric acid levels in serum(P<0.001).Supplementation of BA improved BWG(P=0.001)and FCR(P=0.005)and increased Ruminococcus numbers(P=0.018)and butyrate concentration(P=0.033)in the ceca,regardless of dietary CP level.Further,addition of BA reduced Clostridium perfringens numbers only in birds fed with RCP diets(P=0.039).At d 35,BA sup-plemented diets showed higher apparent ileal digestibility of cystine(P=0.013),valine(P=0.020),and lysine(P=0.014).In conclusion,this study suggests positive effects of BA supplementation in broiler diets via modulating gut microflora and improving nutrient uptake.

Bacillus amyloliquefaciens (IAE635) and their metabolites could purify pollutants, Vibrio spp. and coliformbacteria in coastal aquaculture wastewater

High-density aquaculture often causes the emission of polluted water to the marine environment in the coastal areas of China.To solve the aquaculture-related water quality problems,it is appropriate to adapt eco-friendly methods,such as using microbes and their metabolic products to purify polluted water.In this study,the purifying effects ofBacillus amyloliquefaciens(IAE635)metabolites(poly-γ-glutamic acid;PP)and IAE635 combined with their metabolites(MP)on turbidity,COD,NO3--N,NH4+-N,Vibrio spp.andcoliform bacteria in coastal aquaculture wastewater were conducted in the lab and in situ ponds.The results showed that the removal ofturbidity,COD,NH4+-Nand NO3--Nwith PP and MP was more significant(p<0.05).Compared to Control treatment(Co),the turbidity,COD,NH4+-Nand NO3--N concentrations at 24thhour were evidently reduced by 86.6%,87.5%,83.3%and 58.0%for PP,87.9%,93.5%,86.5%and 78.0%for MP,respectively.The populations of water pathogens under PP and MP were also significantly(p<0.05)removed compared with those of Co;at 24thhour,the Vibriospp.and coliform bacteria were decreased by 68.7%and 66.3%forPP,75.0%and 67.1%for MP,respectively.The water purifying effect of MP was slightly better than that of PP.In situ pond purification test demonstrated that MP significantly lowered the concentrations of turbidity,COD,NH4+-N,NO3--N and NO2--N,which was more effective than EM.A significantly higher(p<0.05)γ-PGA concentration and the total bacterial population for MP compared to PP indicatedthatMPpurifies the coastal aquaculture wastewaterby both flocculation and microbial decomposition.The application of MP will benefit the aquaculture industry by providing a novel method for the removal of chemical pollutants and pathogens.

维氏气单胞菌拮抗菌株J2-2的筛选、鉴定与效果评价

为筛选对维氏气单胞菌(Aeromonas veronii)有潜在抑菌作用的菌株,本研究从湖北公安县泥鳅养殖池塘底泥中分离一株细菌J2-2,对其特性进行了研究,并对抑菌活性物质进行了初步分析。结果显示:菌株J2-2为解淀粉芽孢杆菌(Bacillus amyloliquefaciens),具有抑菌广谱性,对米尔伊丽莎白菌(Elizabethella milieri)、嗜水气单胞菌(A.hydrophila)等6种水产病原菌的抑菌直径在9.6~29.5 mm。菌株J2-2的抑菌物质具有较好的酸碱稳定性、热稳定性和蛋白酶K处理的稳定性。电镜结果显示菌株J2-2的发酵产物对维氏气单胞菌的细胞膜结构具有破坏作用。通过LC-MC分析该拮抗细菌可产生至少15种不同的脂肽类化合物如杆菌霉素等。药敏试验结果显示,菌株J2-2对氨苄西林、青霉素具有耐药性,对克林霉素中度敏感,对大多数抗生素高度敏感。