Glycerol is a main co-product of biodiesel production. Crude glycerol may serve as a cheap and attractive substrate in biotechnological applications, e.g. for the production of valuable chemicals or as an electron donor for reduction processes. In this work, sulfate reduction with glycerol was studied at neutral and acidic pH using bioreactor sludge samples and Tinto River sediments as a source of inoculum, respectively. Communities of sulfate-reducing bacteria (SRB) and fermentative bacteria were co-enriched at both pH values. Molecular analyses revealed that sequences belonging to Desulfomicrobium genus were dominant in the cultures enriched at pH 7, while Desulfosporosinus sequences dominated in the culture enriched at pH 4. Glycerol conversion was coupled to sulfate reduction, but the substrate was incompletely oxidized to acetate in the neutrophilic enrichments, and acetate, lactate, and 1,3-propanediol under low pH conditions. Two strains belonging to Desulfomicrobium and Proteiniphilum genera were isolated from the neutrophilic enrichments, but the first isolate was not able to use glycerol, which suggests a syntrophic relationship between glycerol-degrading fermentative bacteria and SRB. A Clostridium strain able to grow with glycerol was isolated from the low pH enrichment. Our data indicate that glycerol promotes the growth of sulfate-reducing communities to form sulfide, which can be used to precipitate and recover heavy metals.
A total of 6,219 positive clones were obtained by constructing a BAC library of uncultured ruminal fungi of gayal, and two clones (xynF1 and eglF2) with lignocellulolytic enzyme activity were selected. The sequencing results showed that xynF1 and eglF2 had 903-bp, and 1,995-bp, open reading frames likely to encode β-xylanase (XynF1) and β-glucosidase (EglF2), respectively. The amino acid sequence of XynF1 had 99% coverage and 95% homology to the endo-β-1,4-xylanase encoded by the cellulase gene of Orpinomyces sp. LT-3 (GenBank accession No. AEO51791.1). The amino acid sequence of EglF2 had 99% coverage and 93% homology to the β-glucosidase encoded by the cellulase gene of Piromyces sp. E2 (GenBank accession No. CAC34952.1). Analysis using the SMART software showed that XynF1 contains a glycoside hydrolase family 11 functional module and a carbohydrate-binding module, while EglF2 contains a glycoside hydrolase family 1 functional module. XynF1 showed the highest relative enzymatic activity, up to 95%, at 45°C and pH 4.2, while EglF2 showed the highest relative enzymatic activity, up to 95%, at 55°C and pH 6.2. In this study, we achieved efficient expression of the xynF1 and eglF2 genes in Pichia pastoris, which laid a foundation for the practical application of the lignocellulolytic enzymes.
The terrestrial cyanobacterium Nostoc commune forms macroscopic colonies in its natural habitats, and these colonies consist of both cellular filaments and massive extracellular matrixes. In this study, the biochemical features of the extracellular matrix components were investigated. Naturally growing N. commune was tolerant to desiccation, and produced massive extracellular polysaccharides that contained both neutral sugars and glucuronic acid as constituent monosaccharides. The extracellular polysaccharide contents and desiccation tolerance were compared in laboratory culture strains of Nostoc species. The laboratory culture of N. commune strain KU002 was sensitive to desiccation and produced smaller amounts of extracellular polysaccharides, unlike the field-isolated naturally growing colonies. Nostoc punctiforme strain M-15, which is genetically closed to N. commune, was able to tolerate desiccation, although the other Nostoc strains were desiccation-sensitive. A laboratory culture strain of the aquatic cyanobacterium Nostoc sphaericum produced massive extracellular polysaccharides but was sensitive to desiccation, suggesting that extracellular matrix production is not enough to make this strain tolerant to desiccation. WspA (water stress protein) and SodF (superoxide dismutase) were found to be characteristic protein components of the extracellular matrix of N. commune. Because the WspA proteins were heterogeneous, the wspA genes were highly diverse among the different genotypes of N. commune, although the sodF gene was rather conservative. The heterogeneity of the WspA proteins suggests their complex roles in the environmental adaptation mechanism in N. commune.
The aim of this study was to determine the probiotic and the prebiotic-like properties of Bacillus subtilis BN, a spore-forming bacterium, also known as “natto-kin”, which is used for making the Japanese fermented food, natto. We used the spores and vegetative cells of this strain and compared their effects on the growth of lactobacilli. Culture supernatant from B. subtilis BN was added to a glucose-free MRS medium used to culture lactobacilli. When lactobacilli were cultured in the supernatant-containing medium, growth was improved. This effect resulted from the digestion of starch by amylase, which was secreted by B. subtilis. Moreover, improved amylase-independent growth was also observed. Co-culture with B. subtilis improved the growth of lactobacilli, and this effect was only observed with vegetative cells; spores did not improve the growth of lactobacilli. This effect on growth was lost upon heat treatment of the vegetative cells. These results suggest that the surface protein of B. subtilis BN vegetative cells participates in the improved growth effect of lactobacilli. It is possible that B. subtilis BN could improve the intestinal flora. In addition, B. subtilis BN inhibited the growth of Salmonella enterica. Thus, it was shown that B. subtilis BN has both a probiotic and prebiotic potential. This is the first study demonstrating the selective growth improvement of indigenous intestinal lactobacilli using B. subtilis BN.
Bacteriophages are genetic elements that play key roles in the evolution and diversification of bacterial genomes. The Shiga toxin (Stx)-encoding phage plays an important role in the horizontal transfer of the stx gene. However, the influence of the Stx phage integration on the physiological properties and gene expression pattern of the host have not been clearly resolved. In this study, we constructed the Sp5 lysogen through lysogenisation of E. coli K-12 by Sp5, an Stx2 phage in enterohaemorrhagic E. coli (EHEC) O157:H7 Sakai, and examined the effect of the resulting lysogen on cell motility under various growth conditions. Sp5 lysogenisation decreased cell motility and the expression of fliC, which encodes flagellin, under anaerobic conditions at 37°C. Sp5 also lowered the expression of fliA, which encodes the FliA-sigma factor responsible for the transcription of fliC, and flhD, which facilitates the expression of fliA. Sp5 lysogenisation reduced the amount of FlhD and FlhC expressed from the araBAD promoter, suggesting that one or more genes present in Sp5 represses flhDC at the post-transcriptional level. Flagellin is highly antigenic and triggers an immune response in the host. Thus, Sp5 might enhance its viability by repressing the expression of the flagellar regulon to circumvent the immune response of host cells.
Sustainable renewable energy is being hotly debated globally because the continued use of finite fossil fuels is now widely recognized as being unsustainable. Microalgae potentially offer great opportunities for resolving this challenge. Abscisic acid (ABA), jasmonic acid (JA) and salicylic acid (SA) are involved in regulating many physiological properties and have been widely used in higher plants. To test if phytohormones have an impact on accumulating lipid for microalgae, ABA, JA and SA were used to induce two Chlorella strains in the present study. The results showed 1.0 mg/L ABA, 10 mg/L SA, and 0.5 mg/L JA, led strain C. vulgaris ZF strain to produce a 45%, 42% and 49% lipid content that was 1.8-, 1.7- and 2.0-fold that of controls, respectively. For FACHB 31 (number 31 of the Freshwater Algae Culture Collection at the Institute of Hydrobiology, Chinese Academy of Sciences), the addition of 1.0 mg/L ABA, 10 mg/L SA, and 0.5 mg/L, JA produced 33%, 30% and 38% lipid content, which was 1.8-, 1.6- and 2.1-fold that of controls, respectively. As for lipid productivity, 1.0 mg/L ABA increased the lipid productivity of C. vulgaris ZF strain and FACHB-31 by 123% and 44%; 10 mg/L SA enhanced lipid productivity by 100% and 33%; the best elicitor, 0.5 mg/L JA, augmented lipid productivity by 127% and 75% compared to that of controls, respectively. The results above suggest that the three phytohormones at physiological concentrations play crucial roles in inducing lipid accumulation in Chlorella.