Production and partial characterization of chitinase from a halotolerant Planococcus rifitoensis strain M2-26

peer reviewedThis paper is the first to investigate the production and partial characterization of the chitinase enzyme from a moderately halophilic bacterium Planococcus rifitoensis strain M2-26, earlier isolated from a shallow salt lake in Tunisia. The impact of salt, salinity concentration, pH, carbon and nitrogen sources on chitinase production and activity have been determined. This is the first report on a high salt-tolerant chitinase from P. rifitoensis, since it was active at high salinity (from 5 to 30% NaCl) as well as in the absence of salt. This enzyme showed optimal activity at 70 C and retained up to 82 and 66% of its original activity at 80 or 90 C, respectively. The activity of the enzyme was also shown over a wide pH range (from 5 to 11). For characterization of the enzyme activity, the chitinase secreted in the culture supernatant was partially purified. The preliminary study of the concentrated dialysed supernatant on native PAGE showed at least three chitinases produced by strain M2-26, with highest activity approximately at 65 kDa. Thus, the thermo-tolerant and high salt-tolerant chitinases produced by P. rifitoensis strain M2-26 could be useful for application in diverse areas such as biotechnology and agro-industry

Phenotypic characterization and 16S rDNA identification of culturable non-obligate halophilic bacterial communities from a hypersaline lake, La Sal del Rey, in extreme South Texas (USA)

Background: La Sal del Rey ( the King’s Salt”) is one of several naturally-occurring salt lakes in Hidalgo County, Texas and is part of the Lower Rio Grande Valley National Wildlife Refuge. The research objective was to isolate and characterize halophilic microorganisms from La Sal del Rey. Water samples were collected from the lake and a small creek that feeds into the lake. Soil samples were collected from land adjacent to the water sample locations. Sample salinity was determined using a refractometer. Samples were diluted and cultured on a synthetic saline medium to grow halophilic bacteria. The density of halophiles was estimated by viable plate counts. A collection of isolates was selected, gram-stained, tested for catalase, and characterized using API 20E® test strips. Isolates were putatively identified by sequencing the 16S rDNA. Carbon source utilization by the microbial community from each sample site was examined using EcoPlate™ assays and the carbon utilization total activity of the community was determined. Results: Results showed that salinity ranged from 4 parts per thousand (ppt) at the lake water source to 420 ppt in water samples taken just along the lake shore. The density of halophilic bacteria in water samples ranged from 1.2 × 102 - 5.2 × 103 colony forming units per ml (cfu ml-1) whereas the density in soil samples ranged from 4.0 × 105 - 2.5 × 106 colony forming units per gram (cfu g-1). In general, as salinity increased the density of the bacterial community decreased. Microbial communities from water and soil samples were able to utilize 12 - 31 carbon substrates. The greatest number of substrates utilized was by water-borne communities compared to soil-based communities, especially at lower salinities. The majority of bacteria isolated were gram-negative, catalase-positive, rods. Biochemical profiles constructed from API 20E® test strips showed that bacterial isolates from low-salinity water samples (4 ppt) showed the greatest phenotypic diversity with regards to the types and number of positive tests from the strip. Isolates taken from water samples at the highest salinity (420 ppt) tended to be less diverse and have only a limited number of positive tests. Sequencing of 16S DNA displayed the presence of members of bacterial genera Bacillus, Halomonas, Pseudomonas, Exiguobacterium and others. The genus Bacillus was most commonly identified. None of the isolates were members of the Archaea probably due to dilution of salts in the samples. Conclusions: The La Sal del Rey ecosystem supports a robust and diverse bacterial community despite the high salinity of the lake and soil. However, salinity does appear to a limiting factor with

High salt-tolerant protease from a potential biocontrol agent bacillus pumilus M3-16

In this paper, we investigate the characterization and evaluation of the antifungal protease activity from a halotolerant strain M3-16 of Bacillus pumilus, earlier isolated from a shallow salt lake in Tunisia. Protease enzyme was highly induced by the pathogen tested in vitro (27.4 U/ml). This is the first report on high salt-tolerant protease from B. pumilus, since it was active at high salinity (from 5 to 30% NaCl, w/v) as well as in the absence of salinity. This enzyme showed optimal activity at 60 °C and pH 8. At 80 °C and 30 min, the enzyme retained up to 91% and it showed stability over a wide pH range (from pH 5 to 11). The enzyme was found to be monomer with an estimated molecular mass of 31 kDa. The amino acid sequence showed high similarity (94%) to ATP-dependent protease from B. pumilus strain ATCC 7061. Thus, our alkaline thermostable and high salt-tolerant protease induced by a phytopathogenic fungus, could be useful for application in diverse areas such as biotechnology alimentary and agronomy industries

Study of Lasidiodiplodia pseudotheobromae, Neofusicoccum parvum and Schizophyllum commune, three pathogenic fungi associated with the Grapevine Trunk Diseases in the North of Tunisia

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Characterization of a novel chitinase from a moderately halophilic bacterium, Virgibacillus marismortui strain M3-23

A new chitinase produced by the moderately halophilic bacterium Virgibacillus marismortui strain M3- 23 was identified and characterized. Distinguishable characteristics of high activity and stability at different pH, temperatures and salinity of M3-23 chitinase are reported. Analysis of the catalytic domain sequence from the enzyme highlighted its relationship to glycosyl hydrolase family 18. Comparison of the deduced chitinase sequence from strain M3-23 to known chitinases from Bacillus species showed low similarity (82%), suggested its novelty. This is the first report of the characterization of chitinase from the species V. marismortui. The halo- and thermo-tolerant nature of the chitinolytic enzyme allows its potential use in agricultural and industrial applications