Ultrastructure of in vivo interactions of the antagonist bacteria Bacillus cereus X16 and B. thuringiensis 55T with Fusarium roseum var. sambucinum, the casual agent of potato dry rot

The interaction of Fusarium roseum var. sambucinum, the causal agent of potato dry rot, with two antagonistic bacteria, Bacillus cereus X16 and B. thuringiensis 55T, was studied on wounded potato tubers using light and electron microscopy. Application of B. cereus X16 or B. thuringiensis 55T to potato wounds before challenge with the pathogen suppressed dry rot and restricted fungal growth in plant tissues to the first few cell layers beneath the site of inoculation. Both bacterial antagonists penetrated into potato tissues and established themselves through intercellular and intracellular proliferation. The extent of Fusarium colonization was appreciably reduced in the bacterized tubers, and most fungal cells in these tubers were severely damaged, with appreciable morphological and structural changes. In potato tubers bacterized by B. thuringiensis 55T, Fusarium invasion of the host tissues did not stimulate structural host reactions, and direct parasitism, which operates by degradation of the fungal cell walls and disintegration of the fungal cytoplasm, seemed to play a key role in the antagonism against Fusarium hyphae. In potato tubers inoculated with B. cereus X16 and challenged with the pathogen, on the other hand, a set of defense reactions, were triggered, including modifications of the primary cell walls and the occlusion of some cells and vascular tissues with different types of electron-opaque materials. Fungal hyphae in the vicinity of these barriers, apparently containing higher than usual levels of phenol-like compounds, usually showed advanced stages of disorganization, suggesting the existence of a fungitoxic environment. The results presented here show that the two antagonistic bacilli use different biocontrol strategies to suppress Fusarium dry rot development

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

Marinobacter piscensis sp nov., a moderately halophilic bacterium isolated from salty food in Tunisia

An aerobic, Gram-negative, moderately halophilic bacterium, oxidase, and catalase positive-designated Abdou3(T), was isolated from salted traditional foods (Anchovies) in Tunisia. Cells were rod-shaped, non-spore-forming and motile. Growth occurred at 15-45 A degrees C (optimum, 37 A degrees C), pH 5.5-8.75 (optimum, 7.3), and in the presence of 1-15 % NaCl (optimum, 10 %). Strain Abdou3(T) used glucose, d-arabinose, and sucrose. Strain Abdou3(T) had Q9 as the major respiratory quinone and C-18:1 omega 9c and C-16:0 as predominant fatty acids. The DNA G+C content was 55.2 mol%. Phylogenetic analysis of the small-subunit ribosomal RNA (rRNA) gene sequence indicated that strain Abdou3(T) had as its closest relative Marinobacter maritimus (identity of 96 %). Based on phenotypic, phylogenetic, and taxonomic characteristics, strain Abdou3(T) is proposed as a novel species of the genus Marinobacter within the order Alteromonadales, for which the name M. piscensis sp. nov. is proposed. The type strain is Abdou3(T) (=DSM 26804(T))

Characterization of Halanaerobaculum tunisiense gen. nov., sp nov., a new halophilic fermentative, strictly anaerobic bacterium isolated from a hypersaline lake in Tunisia

A new halophilic anaerobe was isolated from the hypersaline surface sediments of El-Djerid Chott, Tunisia. The isolate, designated as strain 6SANG, grew at NaCl concentrations ranging from 14 to 30%, with an optimum at 20-22%. Strain 6SANG was a non-spore-forming, non-motile, rod-shaped bacterium, appearing singly, in pairs, or occasionally as long chains (0.7-1 x 4-13 mu m) and showed a Gram-negative-like cell wall pattern. It grew optimally at pH values between 7.2 and 7.4, but had a very broad pH range for growth (5.9-8.4). Optimum temperature for growth was 42A degrees C (range 30-50A degrees C). Strain 6SANG required yeast extract for growth on sugars. Glucose, sucrose, galactose, mannose, maltose, cellobiose, pyruvate, and starch were fermented. The end products from glucose fermentation were acetate, butyrate, lactate, H-2, and CO2. The G + C ratio of the DNA was 34.3 mol%. Strain 6SANG exhibited 16S rRNA gene sequence similarity values of 91-92% with members of the genus Halobacteroides, H. halobius being its closest phylogenetic relative. Based on phenotypic and phylogenetic characteristics, we propose that this bacterium be classified as a novel species of a novel genus, Halanaerobaculum tunisiense gen. nov., sp. nov. The type strain is 6SANG(T) (=DSM 19997(T) = JCM 15060(T))

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

MALDI-TOF typing highlights geographical and fluconazole resistance clusters in Candida glabrata2188

Utilizing matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectra for Candida glabrata typing would be a cost-effective and easy-to-use alternative to classical DNA-based typing methods. This study aimed to use MALDI-TOF for the typing of C. glabrata clinical isolates from various geographical origins and test its capacity to differentiate between fluconazole-sensitive and -resistant strains. Both microsatellite length polymorphism (MLP) and MALDI-TOF mass spectra of 58 C. glabrata isolates originating from Marseilles (France) and Tunis (Tunisia) as well as collection strains from diverse geographic origins were analyzed. The same analysis was conducted on a subset of C. glabrata isolates that were either susceptible (MIC &lt;/= 8 mg/l) or resistant (MIC &gt;/= 64 mg/l) to fluconazole. According to the seminal results, both MALDI-TOF and MLP classifications could highlight C. glabrata population structures associated with either geographical dispersal barriers (p &lt; 10-5) or the selection of antifungal drug resistance traits (&lt;10-5). In conclusion, MALDI-TOF geographical clustering was congruent with MPL genotyping and highlighted a significant population genetic structure according to fluconazole susceptibility in C. glabrata. Furthermore, although MALDI-TOF and MLP resulted in distinct classifications, MALDI-TOF also classified the isolates with respect to their fluconazole susceptibility profile. Further prospective studies are required to evaluate the capacity of MALDI-TOF typing to investigate C. glabrata infection outbreaks and predict the antifungal susceptibility profile of clinical laboratory isolates</p

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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