Functional annotation of the genome unravels probiotic potential of Bacillus coagulans HS243

Spore forming Bacillus species are widely used as probiotics for human dietary supplements and in animal feeds. However, information on genetic basis of their probiotic action is obscure. Therefore, the present investigation was undertaken to elucidate probiotic traits of B. coagulans HS243 through its genome analysis. Genome mining revealed the presence of an arsenal of marker genes attributed to genuine probiotic traits. In silico analysis of HS243 genome revealed the presence of multi subunit ATPases, ADI pathway genes, chologlycine hydrolase, adhesion proteins for surviving and colonizing harsh gastric transit. HS243 genome harbored vitamin and essential amino acid biosynthetic genes, suggesting the use of HS243 as a nutrient supplement. Bacteriocin producing genes highlighted the disease preventing potential of HS243. Thus, this work established that HS243 possessed the genetic repertoire required for surviving harsh gastric transit and conferring health benefits to the host which were further validated by wet lab evidences.     

Surfactin isoforms from Bacillus coagulans

Bacillus coagulans has been found to produce several surfactins that are powerful lipopeptide surfactants. Four main components with molecular weights 1007, 1021 and 1035 Da were separated. Their structures have been confirmed by spectrometric and spectroscopic studies and by acid hydrolysis. The compounds were found to represent two pairs of surfactin isoforms in which beta-hydroxy-iso-C14 or anteiso-C15 fatty acids are linked to the [Leu7] or [Val7] heptapeptide moiety by both an amide group and a lactone bond.     

Functional Analysis of the Accessory Protein TapA in Bacillus subtilis Amyloid Fiber Assembly

Bacillus subtilis biofilm formation relies on the assembly of a fibrous scaffold formed by the protein TasA. TasA polymerizes into highly stable fibers with biochemical and morphological features of functional amyloids. Previously, we showed that assembly of TasA fibers requires the auxiliary protein TapA. In this study, we investigated the roles of TapA sequences from the C-terminal and N-terminal ends and TapA cysteine residues in its ability to promote the assembly of TasA amyloid-like fibers. We found that the cysteine residues are not essential for the formation of TasA fibers, as their replacement by alanine residues resulted in only minor defects in biofilm formation. Mutating sequences in the C-terminal half had no effect on biofilm formation. However, we identified a sequence of 8 amino acids in the N terminus that is key for TasA fiber formation. Strains expressing TapA lacking these 8 residues were completely defective in biofilm formation. In addition, this TapA mutant protein exhibited a dominant negative effect on TasA fiber formation. Even in the presence of wild-type TapA, the mutant protein inhibited fiber assembly in vitro and delayed biofilm formation in vivo. We propose that this 8-residue sequence is crucial for the formation of amyloid-like fibers on the cell surface, perhaps by mediating the interaction between TapA or TapA and TasA molecules.     

凝结芽孢杆菌在动物饲料中的应用

随着我国“饲料禁抗, 养殖减抗”政策的实施, 饲料及动保企业都在寻找合理有效的抗生素替代品。目前, 人们对以凝结芽孢杆菌为主的饲用微生态制剂进行了很多的实证试验, 替抗效果也逐渐得到行业认可。凝结芽孢杆菌拥有乳酸菌和芽孢菌的双重特性, 能耐受饲料制粒温度存活, 到达胃肠道后萌发并分泌代谢产物, 最终促进畜禽的肠道健康。本文对凝结芽孢杆菌的生物学特性和生物学功能进行阐述, 总结了近年来凝结芽孢杆菌在畜禽、水产和生物饲料中的应用研究, 以期为凝结芽孢杆菌在畜牧养殖中的合理应用提供参考。

     

Cyclomaltodextrin glucanotransferase-producing moderate thermophile,Bacillus coagulans

A new CGTase-producing moderate thermophile was isolated from soil, and was identified as Bacillus coagulans which have not previously been listed as cyclodextrin producing bacteria. The culture filtrate of the isolate as the CGTase source converted about 60% of soluble starch to CD's in 20 h at 50°C, and the ratio of α-: β-: γ-CD produced was 1.0: 0.9: 0.3.     

Growth of Bacillus coagulans in Chemically Defined Media

A nutritional study was made of five strains of Bacillus coagulans obtained from various culture collections. These five strains were descendants of two original isolates; three had been derived from one parent culture in years past and the other two were transfers from another parent culture. Therefore, the five cultures should have represented two distinct groups of genetically identical cultures. Three of the strains obtained from one culture collection had become methyl red-negative and sorbitol-negative and had gained abilities to hydrolyze gelatin and ferment arabinose. Nutritional requirements of the five cultures, determined at 37, 45, and 55 C, differed considerably among strains; however, thiamine and biotin were required by all cultures at all temperatures. Aspartic acid was stimulatory at 37 C and was required at 45 C; folic acid, basic amino acids, and certain other nutrilites were required at 55 C. Adenine supplementation was necessary for two strains at 55 C to prevent autolysis; this phenomenon is discussed. The response of these organisms to both serine and the basic amino acids at the three growth temperatures seems especially significant. The media devised for the growth of the five strains of B. coagulans used in this study permit excellent growth at three incubation temperatures.     

补料分批法高密度培养凝结芽孢杆菌

研究在摇瓶条件下通过补加含有不同浓度的葡萄糖与酵母膏的料液及不同流加方式对凝结芽孢杆菌(Bacillus coagulans)TQ33菌体量和芽孢生成量的影响,确定了补料液中的葡萄糖与酵母膏比例为61(w/w),采用将葡萄糖浓度控制在3.0g/L～6.0g/L的流加方式进行高密度培养凝结芽孢杆菌.在5L自动发酵罐中进行补料分批培养,最终菌体浓度达到4.5×109cfu/mL,芽孢浓度为1.2×109cfu/mL,分别是分批培养的5.9倍和3.8倍.     

Purification and properties of Bacillus coagulans cyclomaltodextrin glucanotransferase

Cyclomaltodextrin glucanotransferase (CGTase), produced in a culture filtrate by Bacillus coagulans, was purified to a single, homogeneous protein. It has a monomeric structure with a molecular weight of 65,000, isoelectric point of 4.6, and contains 2 mol of Ca²⁺ per mol of the enzyme. The enzyme was most active at pH 6.0 and at 70°C. It did not lose its activity by heat treatment at 70°C for 10 min in the presence of CaCl₂ in the pH range of 5.5∼9.5, and by incubation in the pH range of 5.0∼10.5 at 4°C for one month. The enzyme converted about 60% of potato starch to cyclodextrins for 20 h at 50°C, and the ratio of α-: β-: γ-cyclodextrin produced was 8.1:8.9:1.0 B. coagulans CGTase was compared with B. macerans CGTase which was purified by the same method.