Effect of pesticides on the growth and secreted chymotrypsin-like activity of a biocontrol strain of bacillus amyloliquefaciens

Biocontrol is an alternative approach to reduce the harmful effects of pathogen species in the agriculture. Pesticide-tolerant biocontrol agents are preferred in the integrated pest management because they can be applied together with different fungicides, herbicides and insecticides. A potent biocontrol agent, Bacillus amyloliquefaciens SZMC 22206 strain was isolated and studied previously. It was revealed, that the extracellular chymotrypsin-like protease and fengycin secretion of the strain resulted its antagonistic effect. The aim of our present study was to analyse the effect of different pesticides on the growth and activity of the extracellular chymotrypsin-like proteases of this Bacillus strain. The tested pesticides were a fungicide (carbendazim), and three frequently used herbicides (linuron, chlortoluron and 2,4-dichlorophenoxyacetic acid). The tested Bacillus strain was able to grow in the presence of the pesticides, but the activities of the extracellular chymotrypsin-like proteases were significantly reduced in some cases

Effect of pesticides on the growth and secreted chymotrypsin-like activity of a biocontrol strain of bacillus amyloliquefaciens

Biocontrol is an alternative approach to reduce the harmful effects of pathogen species in the agriculture. Pesticide-tolerant biocontrol agents are preferred in the integrated pest management because they can be applied together with different fungicides, herbicides and insecticides. A potent biocontrol agent, Bacillus amyloliquefaciens SZMC 22206 strain was isolated and studied previously. It was revealed, that the extracellular chymotrypsin-like protease and fengycin secretion of the strain resulted its antagonistic effect. The aim of our present study was to analyse the effect of different pesticides on the growth and activity of the extracellular chymotrypsin-like proteases of this Bacillus strain. The tested pesticides were a fungicide (carbendazim), and three frequently used herbicides (linuron, chlortoluron and 2,4-dichlorophenoxyacetic acid). The tested Bacillus strain was able to grow in the presence of the pesticides, but the activities of the extracellular chymotrypsin-like proteases were significantly reduced in some cases

Investigation of antimicrobial agents produced by gram-negative bacteria

Bacterial secondary metabolites are low molecular mass compounds, which are not essential for bacterial growth. These secondary metabolites are produced in the stationary phase of bacterial growth and the produced compounds could have a variety of biological functions. One of these functions are the antibiotic or antimicrobial function that is gaining interest in the microbial community recently. In this work Gram-negative bacterial strains were cultivated, and their produced secondary metabolites were extracted and tested in antimicrobial plate assays against Gram-positive and Gram-negative bacterial strains as well. The well-known antibiotic, pyrrolnitrin content of the extracts were also determined by HPLC-Quadrupole-Orbitrap MS

Purification of surfactin compounds produced by a Bacillus subtilis strain

Surfactins are lipopeptide-type biosurfactants produced mainly by Bacillus species containing a peptide loop of seven amino acids and a hydrophobic fatty acid chain. These molecules exhibit various biological activities; therefore, their therapeutic and environmental applications are in the focus worldwide. In our work, a multi-step purifi cation and separation process was developed to isolate surfactins from the ferment broth of B. subtilis SZMC 6179J strain. The process incorporates normal phase fl ash chromatography for pre-purifying the crude extract and two consecutive reverse phase HPLC separations for the isolation of the various surfactin molecules. The determination of the relative amounts of lipopeptides both in the crude extract and in each fraction of every separation step were carried out by HPLC-HESI-MS examinations. The ratio of surfactins in the crude extract was 21.35%, but after the preparative fl ash chromatographic separation the relative amount of surfactins was observed to be 30.44%. The preparative HPLC purifi cation step resulted 85.39% purity of the surfactins. Nine diff erent surfactin variants were isolated and identifi ed from the fractions of this fi nal semi-preparative HPLC purifi cation, out of which three compounds were completely purifi ed, and three others were detected in relative amounts of more than 95% in some fractions

Identifications of Surfactin-Type Biosurfactants Produced by Bacillus Species Isolated from Rhizosphere of Vegetables

Surfactins are cyclic lipopeptides consisting of a β-hydroxy fatty acid of variable chain length and a peptide ring of seven amino acids linked together by a lactone bridge, forming the cyclic structure of the peptide chain. These compounds are produced mainly by Bacillus species and are well regarded for their antibacterial, antifungal, and antiviral activities. For their surfactin production profiling, several Bacillus strains isolated from vegetable rhizospheres were identified by their fatty acid methyl ester profiles and were tested against phytopathogen bacteria and fungi. The isolates showed significant inhibition against of E. amylovora, X. campestris, B. cinerea, and F. culmorum and caused moderate effects on P. syringae, E. carotovora, A. tumefaciens, F. graminearum, F. solani, and C. gloeosporioides. Then, an HPLC-HESI-MS/MS method was applied to simultaneously carry out the quantitative and in-depth qualitative characterisations on the extracted ferment broths. More than half of the examined Bacillus strains produced surfactin, and the MS/MS spectra analyses of their sodiated precursor ions revealed a total of 29 surfactin variants and homologues, some of them with an extremely large number of peaks with different retention times, suggesting a large number of variations in the branching of their fatty acid chains

Antidermatophytic effect of Bacillus mojavensis SZMC 22228 and its secreted chymotrypsin-like protease

The aim of the present study was to investigate the antifungal effect of Bacillus mojavensis SZMC 22228 against different dermatophytes and to isolate the antidermatophytic compound from the bacterial ferment broth. B. mojavensis SZMC 22228 and its cell-free ferment broth effectively inhibited the growth of clinical reference strain of Microsporum canis, Microsporum gypseum, Trichophyton mentagrophytes, Trichophyton rubrum and Trichophyton tonsurans in agar diffusion test in vitro. An antidermatophytic, ~25 kDa protein (B. mojavensis SZMC 22228 antidermatophyitc protein, BMAP) was purified from the antifungally active, cell-free ferment broth using size exclusion and ion-exchange chromatography. BMAP showed antifungal effect against all of the investigated dermatophytes both in agar diffusion and broth microdilution susceptibility tests. M. gypseum proved to be the most susceptible dermatophyte to BMAP (MIC=40 μg/ml), all the other investigated fungi were less susceptible (MIC=80 μg/ml). The enzymatic activity of this protein was investigated in microtiter plate assay using hydrolase specific chromogenic substrates. BMAP showed high proteolytic activity towards N-Succ-Ala-Ala-Pro-Phe-pNA, and proved to be a chymotrypsin-like protease. These results suggest that the antidermatohytic activity of B. mojavensis SZMC 22228 correlates with its chymotrypsin-like protease production. After further investigations, the purified BMAP could be a promising base of a novel antidermatophytic strategy