Use of bacterial endospore with longer shelf-life in improved retting of jute

Aim: The aim of the present study was to evaluate the retting of jute (Corchorus olitorius L. and C. capsularis L.) using the endospores of microbial consortium of three strains of Bacillus pumilus with extended shelf-life. Methodology: Endospore and vegetative cells of Bacillus pumilus were tested for viability by introducing them into different temperature, pH, UV radiation and antibiotics. Laboratory, as well as field-trials of jute retting was performed with 6 and 18-months-old endospores and vegetative cells of Bacillus pumilus with estimation of enzymatic activities for comparison of their retting efficiency. Results: Endospores of Bacillus pumilus recorded very high colony forming unit (109 to 108ml-1) compared to their vegetative cells (106 to 104ml-1) after 6 to 18 months of their preservation. Endospores also showed higher resistance to temperature, pH, UV irradiation and antibiotic than their vegetative forms. High colony forming unit and higher release of pectinolytic and xylanolytic enzymes during retting of jute by endospores resulted in complete of jute retting in 10 days with good quality jute fibre compared to talc based formulation. Interpretation: It can be concluded from the study that endospores remained highly efficient in rejuvenating higher CFU and quantitatively larger pool of enzymes to accelerate retting of jute after prolonged preservation. Therefore, the endospores of Bacillus pumilus can be used cost effectively in place of their talc based formulation for higher shelf life of the product, faster retting and better fibre quality of jute.

Two methods for isolation of endophytic and edaphic Bacillus spp. from sugarcane fields / Dois métodos para isolamento de Bacillus spp. endofíticos e edáficos de canaviais

Bacillus has been widely studied and used for the control of pests and diseases. The adapted protocol proposed by POLANCZYK (2004) proved to be more efficient than the one by the World Health Organization (WHO, 1985) to isolate edaphic strains of Bacillus. However, it has not been assessed for isolation of endophytic strains, which are much less abundant in the nature and more difficult to be isolated. This study aimed to compare two methodological procedures for isolation of Bacillus, established by the WHO (1985) and by POLANCZYK (2004), regarding their efficiency for isolation of endophytics and edaphics Bacillus strains from inside the root tissue of sugarcane, as well as from the associated soil sample, collected from 11 locations; and to compare the density of bacteria in both environments. Endophytic and edaphic strains of Bacillus were isolated by both procedures. However, the isolation protocol performed by POLANCZYK (2004) made more efficient by having a greater number of colony forming units (CFU) per gram of soil and root indicating that this procedure is more useful, especially for isolation of endophytic strains of Bacillus, which are much less abundant in the nature than edaphic strains, being therefore more difficult to be isolated. Using the Polanczyk protocol (2004), Bacillus strains were recovered from all roots (endophytic) and soil (edaphic) samples of all the 11 fields, suggesting that the plant root may be another important source for isolation of Bacillus besides the soil. Higher densities of Bacillus were isolated from the edaphic environment compared with the endophytic environment, with significant differences when isolated by Polanczyk method (2004).(AU)

Bacillus tem sido amplamente estudado e usado para o controle de pragas e doenças. O protocolo adaptado proposto por POLANCZYK (2004) mostrou-se mais eficiente do que o da Organização Mundial de Saúde (WHO, 1985) para isolar cepas edáficas de Bacillus. No entanto, não foi avaliado quanto ao isolamento de estirpes endofíticas, que são muito menos abundantes na natureza e mais difíceis de isolar. Este estudo teve como objetivos comparar dois procedimentos metodológicos para o isolamento de Bacillus, o estabelecido pela OMS (WHO, 1985) e o de POLANCZYK (2004), quanto a sua eficiência para o isolamento de estirpes endofitas e edáficas de Bacillus originárias do interior do tecido radicular de cana-de-açúcar, bem como de amostras de solos associados, coletada de 11 locais; e comparar a densidade de bactérias em ambos os ambientes. As cepas endofíticas e edáficas de Bacillus foram isoladas por ambos os procedimentos. No entanto, o protocolo de isolamento realizado por POLANCZYK (2004) demonstrou-se mais eficiente por gerar maior número de unidades de formação de colônias (CFU) por grama de solo e raiz, indicando que esse procedimento é mais útil, especialmente para isolamento de estirpes endofíticas de Bacillus, que são muito menos abundantes na natureza do que as cepas edáficas, sendo, portanto, mais difíceis de serem isoladas. Usando o protocolo de POLANCZYK (2004), as cepas de Bacillus foram isoladas de todas as amostras de raízes (endofíticas) e de solo (edáficas) de todos os 11 campos, sugerindo que a raiz da planta pode ser outra fonte importante de isolamento de Bacillus além do solo. As densidades mais altas de Bacillus foram isoladas do ambiente edáfico em comparação com o ambiente endofítico, com diferenças significativas quando isoladas pelo método de POLANCZYK (2004).(AU)

The genotypic diversity and lipase production of some thermophilic bacilli from different genera

Abstract Thermophilic 32 isolates and 20 reference bacilli were subjected to Rep-PCR and ITS-PCR fingerprinting for determination of their genotypic diversity, before screening lipase activities. By these methods, all the isolates and references could easily be differentiated up to subspecies level from each other. In screening assay, 11 isolates and 7 references were found to be lipase producing. Their extracellular lipase activities were measured quantitatively by incubating in both tributyrin and olive oil broths at 60 °C and pH 7.0. During the 24, 48 and 72-h period of incubation, the changes in the lipase activities, culture absorbance, wet weight of biomass and pH were all measured. The activity was determined by using pNPB in 50 mM phosphate buffer at pH 7.0 at 60 °C. The lipase production of the isolates in olive oil broths varied between 0.008 and 0.052, whereas these values were found to be 0.002-0.019 (U/mL) in the case of tyributyrin. For comparison, an index was established by dividing the lipase activities to cell biomass (U/mg). The maximum thermostable lipase production was achieved by the isolates F84a, F84b, and G. thermodenitrificans DSM 465T (0.009, 0.008 and 0.008 U/mg) within olive oil broth, whereas G. stearothermophilus A113 displayed the highest lipase activity than its type strain in tyributyrin. Therefore, as some of these isolates displayed higher activities in comparison to references, new lipase producing bacilli were determined by presenting their genotypic diversity with DNA fingerprinting techniques.

Isolation of Antibiotic Producing Bacteria from Soil.

Most antibiotics used today are isolated and extracted from microbial source. The emergence of antibiotic resistance and need for better, broad spectrum antibiotics is always in high demand. In the present study, antibiotic producing bacteria were isolated from a local soil sample. After primary screening, two bacterial strains (strain 1 and 2) were isolated, which showed antimicrobial activity against some common bacteria namely, E. coli, K. pneumonia, S. aureus, P.aeruginosa and M. smegmatis. The isolated strains showed prominent zones of inhibition against E. coli, K. pneumoniae, S. aureus and P. Aeruginosa but not against M. smegmatis. To identify the isolated strains, biochemical tests were performed and it was found that both the strains were Bacillus sp with some differences in cultural characteristics. The strains were also found to be resistant to some antibiotics such as tetracycline. The screen for the presence of any plasmid which might be influencing antibiotic production by these strains, the cultures were used for extraction of plasmids wherein, both the strains were observed to have a plasmid of ~ 10 kb. Further study in identification of these strains will be helpful in improving these strains for antibiotic production.

Anti-staphylococcal and antifungal substances produced by endospore-forming bacilli.

Some bacteria and fungi are related to deterioration and also transmission of foodborne diseases, emphasizing the need to search new substances that may act in the treatment and prevention of the illnesses transmitted by food. Strains from genus Bacillus produce a variety of substances with inhibitory activity that range from antibiotics to bacteriocins. In this work, three strains, identified as B. pasteurii (Pes1) and B. insolitus (Mam2 and Ame3) presented inhibitory action against staphylococcal strains isolated from food. Out of the 33 strains tested, 31 (94.0%) were inhibited by at least one of three main Bacillus producer strains, being most of them inhibited by strain Pes1, that also was able to inhibit filamentous fungi related to food spoilage. The antimicrobial substances produced by Pes1, Mam2 and Ame3 showed to be resistant to proteolytic enzymes, suggesting these substances have not an active proteinaceous compound, as typical bacteriocins. New studies are being performed to extract and characterize these antimicrobial agents to evaluate their potential application in biological control of microorganisms related to spoilage food and foodborne diseases.

The genetic diversity of genus Bacillus and the related genera revealed by 16S rRNA gene sequences and ardra analyses isolated from geothermal regions of turkey

Previously isolated 115 endospore-forming bacilli were basically grouped according to their temperature requirements for growth: the thermophiles (74%), the facultative thermophiles (14%) and the mesophiles (12%). These isolates were taken into 16S rRNA gene sequence analyses, and they were clustered among the 7 genera: Anoxybacillus, Aeribacillus, Bacillus, Brevibacillus, Geobacillus, Paenibacillus, and Thermoactinomycetes. Of these bacilli, only the thirty two isolates belonging to genera Bacillus (16), Brevibacillus (13), Paenibacillus (1) and Thermoactinomycetes (2) were selected and presented in this paper. The comparative sequence analyses revealed that the similarity values were ranged as 91.4-100 %, 91.8- 99.2 %, 92.6- 99.8 % and 90.7 - 99.8 % between the isolates and the related type strains from these four genera, respectively. Twenty nine of them were found to be related with the validly published type strains. The most abundant species was B. thermoruber with 9 isolates followed by B. pumilus (6), B. lichenformis (3), B. subtilis (3), B. agri (3), B. smithii (2), T. vulgaris (2) and finally P. barengoltzii (1). In addition, isolates of A391a, B51a and D295 were proposed as novel species as their 16S rRNA gene sequences displayed similarities ¡Ü 97% to their closely related type strains. The AluI-, HaeIII- and TaqI-ARDRA results were in congruence with the 16S rRNA gene sequence analyses. The ARDRA results allowed us to differentiate these isolates, and their discriminative restriction fragments were able to be determined. Some of their phenotypic characters and their amylase, chitinase and protease production were also studied and biotechnologically valuable enzyme producing isolates were introduced in order to use in further studies.

SPORE CELLULAR FATTY ACIDS ANALYSIS OF SOME AEROBIC ENDOSPORE-FORMING BACILLI / 微生物学通报

A gas chromatographic analysis method was employed to determine the cellular fatty acids (CFAs)profiles of the spores of some aerobic endospore forming bacilli.Purified spore cultures of 51 experimentas strains were processed to acquire whole cell fatty acids methyl esters for the subsequent gas chromatographic analysis,and the corresponding vegetative cells were set as control.The reproducibility study of spore fatty acids revealed that,the fatty acids components of spores were stable enough for research purpose,provided under standardized experimentas procedure.The dendrograms obtained by cluster analysis provided some meaningful taxonomic information of the experimental strains.The fatty acids analysis of spores seemed to be a promising supplementary tool for the chemotaxonomic research of aerobic endospore-forming bacilli.