Bacteriocins are antimicrobial peptides with potential applications as therapeutic agents for the treatment of microbial infections. The aim of this work was to investigate the effect of different protectors on the activity of salivaricin CRL1328, a bacteriocin produced by Lactobacillus salivarius CRL1328, during the lyophilization process and subsequent storage at different temperatures for 18 months using statistical models. Different protectors such as mannitol, Tween 80, polyethylene glycol 8000 (PEG), monosodium glutamate (MSG), reconstituted skim milk, sucrose and ascorbic acid were used for the lyophilization and storage of salivaricin. The biplot of principal component analysis was used for the interpretation of the interactions between the different factors studied. The antimicrobial activity of salivaricin was dependent mainly on temperature, and also on the time of storage and protector assayed. The stability of salivaricin was higher at -20°C and 4°C than 25°C and decreased during the time of storage; however, salivaricin was active after 18 months of storage at 25°C. Sucrose, mannitol plus sucrose, PEG plus sucrose and MSG were the most effective agents in protecting the bacteriocin during the lyophilization process. Effective maintenance of the activity of the bacteriocin was observed by storage with sucrose and ascorbic acid at –20°C as well as with PEG plus sucrose at 4°C and –20°C. The results obtained suggest that sucrose alone or combined with PEG can effectively maintain the activity of salivaricin during lyophilization and storage. This study provides useful information for the potential application of salivaricin as a bioactive principle for a pharmaceutical formulation.
Bacillus thuringiensis is a bacterium of great agronomic and scientific interest. The subspecies of this bacterium colonize and kill a large variety of host insects and even nematodes, but each strain does so with a high degree of specificity. Therefore molecular typing and diversity analysis of B. thuringiensis has enormous importance for discrimination of strains isolated from different sources. In this study, 113 native B. thuringiensis isolates collected from diverse habitats and locations in India and 27 B. thuringiensis type strains obtained from the Bacillus Genetic Stock Centre (BGSC), Ohio State University, USA and used as reference, were analyzed for molecular typing. Genotypic data of 140 B. thuringiensis isolates and type strains was generated by using REP-PCR and ERIC-PCR primers and unweighted pair group method with arithmetic mean (UPGMA) analysis using NTSYSpc2.2 and grouped into 4 main clusters. All the groups have isolates from diverse origins. No group was found to represent any specific origin or location. The observed patterns of REP-PCR and ERIC-PCR pattern were discriminatory enough to reveal differences in the B. thuringiensis isolates and reference strains. The resolution power and marker index of the ERIC-PCR (RP 9.39, MI 6.34) was found to be higher than that of the REP-PCR (RP 6.20, MI 4.48). The REP-PCR and ERIC-PCR markers have been found to be useful for discrimination of B. thuringiensis isolates and reference strains. ERIC-PCR was the more informative of the two techniques. This study showed that the B. thuringiensis isolates collected from diverse habitats in India had a high degree of genetic diversity.
Various traditional fermented yak milk and raw milk foods could be considered as an abundant resource for obtaining novel lactic acid bacteria (LAB) with unique properties. Eighty-eight samples of yak milk products were collected from Gansu Province in China. Three hundred and nineteen strains of LAB isolated from these samples were identified by phenotypic methods, 16S rRNA gene sequence analysis and PCR-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) technology. Among the isolates, one hundred and sixty-four isolates (51.41% of the total) were classified under Lactobacilli, and one hundred and fifty-five (48.59%) belonged to cocci. All the isolates were classified to six genera (Lactobacillus, Lactococcus, Leuconostoc, Streptococcus, Enterococcus and Weissella) and twenty-one species. Lactobacillushelveticus (87 strains), Leuconostocmesenteroides subsp. mesenteroides (49 strains), Streptococcusthermophilus (39 strains), Lactobacilluscasei (31 strains) and Lactococcuslactis subsp. lactis (19 strains) were considered as the predominant populations in the yak milk products. The results showed that there were abundant genus and species LAB existing in yak milk products in Gansu Province in China. The obtained LAB pure cultures may be a valuable source for further starter selection.
Sorghum presents a sustainable feedstock for Mediterranean buffaloes due to its reduced water and nitrogen requirements compared with maize, which is currently fed primarily. We investigated the effects of feeding sorghum as opposed to maize on Mediterranean buffalo rumen microbial diversity and milk fatty acid content. Four cannulated lactating Mediterranean buffalo cows were fed a basal diet for one month before switching either to maize or sorghum-silage based diets for a 3-month period. Buffaloes were then changed over to the contrasting diet for a further one month. Rumen and milk samples were collected at the end of each month. DGGE- and T-RFLP-based dendrograms generated from rumen samples did not show an effect of diet on rumen bacterial diversity. Milk samples also did not differ in terms of their fatty acid content post sorghum feeding as compared with maize feeding. Thus, sorghum provides an environmentally beneficial alternative to maize for feeding Mediterranean buffalo with little effect on rumen microbial diversity or milk fatty acid composition compared with maize feeding.
The objective of this study is to determine whether DNA signature recovery of Bacillus anthracis strains from different environmental substrates correlates with pathogen cell surface hydrophobicity and induction of host cell death. We compared recovery of DNA signatures from a panel of B. anthracis strains collected from two environmental substrates, non-porous surfaces and soil, using real-time qPCR. We further assessed both cell surface hydrophobicity of the B. anthracis strains by contact angle measurements and host cell viability in response to B. anthracis infection in a mouse macrophage cell model system. Our studies demonstrated correlation between reduced B. anthracis sample recovery from environmental substrates and increased cell surface hydrophobicity. Surprisingly, the most hydrophilic strain, K4596, which exhibited the highest level of recovery from the environmental surfaces, induced the highest level of host cell cytotoxicity compared to more hydrophobic B. anthracis strains in the panel. Our results suggest that cell surface hydrophobicity may play a leading role in mediating pathogen adherence to environmental surfaces. These findings can contribute to the optimization of pathogen detection efforts by understanding how bacterial parameters such as hydrophobicity and induction of host cell death affect bacterial adherence to environmental surfaces.
In many parts of the world Mucuna pruriens is used as an important medicinal, forage and green manure crop. In the present investigation the effect of the addition of CMC in carrier during development of bioformulation on shelflife, plant growth promotive and biocontrol activity against Macrophomina phaseolina was screened taking M. pruriens as a test crop. Ensifer meliloti RMP6Ery+Kan+ and Bradyrhizobium sp. BMP7Tet+Kan+ (kanamycin resistance engineered by Tn5 transposon mutagenesis) used in the study showed production of siderophore, IAA, solubilizing phosphate and biocontrol of M. phaseolina. RMP6Ery+Kan+ also showed ACC deaminase activity. The survival of both the strains in sawdust-based bioformulation was enhanced with an increase in the concentration of CMC from 0 to 1%. At 0% CMC Bradyrhizobium sp. BMP7Tet+Kan+ showed more increase in nodule number/plant (500.00%) than E.meliloti RMP6Ery+Kan+ (52.38%), over the control in M. phaseolina-infested soil. There was 185.94% and 59.52% enhancement in nodule number/plant by RMP6Ery+Kan+ and BMP7Tet+Kan+ with an increase in the concentration of CMC from 0% to 1% in the bioformulations. However further increase in concentration of CMC did not result in enhancement in survival of either the strains or nodule number/plant.
The phenotypic, chemotaxonomic and genetic peculiarities of 5 deep strains of Alteromonas macleodii (isolated from Adriatic and Ionian Sea water from a depth of 1,000–3,500 m) and 5 strains of the same species isolated from the surface layer of Aegean, Andaman, Black Sea and Atlantic Ocean water near the British shore have been studied. Electron microscopy has shown that the deep strains’ cells were, on average, two times longer (2.1±0.2×0.7±0.1 μm) than the surface strains’ (1.1±0.1×0.6±0.1 μm). Using fatty acid analysis (particularly the mono-unsaturated C16:1 and C18:1 fatty acids contents) the deep and surface isolates were clearly separated into two clusters. Distinctions between them were also found in different lectin binding capacity, which is probably determined by the structure of their extracellular polysaccharide matrix. Analysis of the results of PCR with primers to repeated nucleotide sequences revealed a higher level of genetic polymorphism in surface strains in comparison with the deep isolates. This division was confirmed by the cluster analysis method though it was not as clear as in the fatty acids analysis. The described peculiarities are probably reflective of specific conditions in which A. macleodii strains live on the surface or in the depth of the world’s oceans.
Scytonemin is a 544-Da hydrophobic pigment that can absorb UV-A radiation. It is present in cyanobacterial sheaths and is thought to function as a UV protectant. In this study, scytonemin was purified from the terrestrial cyanobacterium Nostoc commune, and its radical-scavenging activity was characterized. The purified scytonemin quenched an organic radical in vitro and accounted for up to 10% of the total activity of an ethanol extract of N. commune. These results suggest that the extracellular UV-absorbing pigment scytonemin has multiple roles, functioning as a UV sunscreen and an antioxidant relevant to anhydrobiosis in N. commune.
Two closely related yeast strains, ST-382 and ST-392, isolated in Thailand showed intermediate relatedness in the DNA-DNA hybridization experiment suggesting that the two strains represent closely related distinct species. In the tree based on the D1/D2 domain sequences of the large subunit rRNA gene, the two strains are located in a subclade in the Wickerhamomyces clade with high bootstrap support. In the D1/D2 domain, the two strains differed by two nucleotides and are assumed to be very closely related. Strain ST-392T (=BCC 15102T = NBRC 107799T = CBS 12176T) forming hat-shaped ascospores is described as Wickerhamomyces tratensis sp. nov. and strain ST-382T (= BCC 15093T = NBRC 107800T = CBS 12175T) is described as Candida namnaoensis sp. nov. because ascospores are not found in this strain. In phenotypic characteristics, W. tratensis and C. namnaoensis are discriminated by the ability of alcoholic fermentation and the assimilation of galactose, D-xylose and D-gluconic acid.
A biosurfactant-producing and hydrocarbon-utilizing bacterium, Pseudomonas aeruginosa USM-AR2, was used to assist conventional distillation. Batch cultivation in a bioreactor gave a biomass of 9.4 g L-1 and rhamnolipid concentration of 2.4 g L-1 achieved after 72 h. Biosurfactant activity (rhamnolipid) was detected by the orcinol assay, emulsification index and drop collapse test. Pretreatment of crude oil TK-1 and AG-2 with a culture of P. aeruginosa USM-AR2 that contains rhamnolipid was proven to facilitate the distillation process by reducing the duration without reducing the quality of petroleum distillate. It showed a potential in reducing the duration of the distillation process, with at least 2- to 3-fold decreases in distillation time. This is supported by GC-MS analysis of the distillate where there was no difference between compounds detected in distillate obtained from treated or untreated crude oil. Calorimetric tests showed the calorie value of the distillate remained the same with or without treatment. These two factors confirmed that the quality of the distillate was not compromised and the incubation process by the microbial culture did not over-degrade the oil. The rhamnolipid produced by this culture was the main factor that enhanced the distillation performance, which is related to the emulsification of hydrocarbon chains in the crude oil. This biotreatment may play an important role to improve the existing conventional refinery and distillation process. Reducing the distillation times by pretreating the crude oil with a natural biosynthetic product translates to energy and cost savings in producing petroleum products.