High Chitinolytic Activity Research Articles

Complete genome sequence of Serratia plymuthica SWSY-3.47.

Serratia plymuthica strain SWSY-3.47 is a Gram-negative, rod-shaped bacteria isolated for its high chitinolytic activity. Here, we report the complete genome of this strain comprised of a single circular chromosome of 5,636,345 bp with a G + C content of 56.0%.

Partitioning recombinant chitinase from Nicotiana benthamiana by an aqueous two-phase system based on polyethylene glycol and phosphate salts

The objectives of this study were to purify 42 kDa chitinase derived from Trichoderma asperellum SH16 produced in Nicotiana benthamiana by a polyethylene glycol (PEG)/salt aqueous two-phase system (ATPS). The specific activities of the crude chitinase and the partially purified chitinase from N. benthamiana were about 251 unit/mg and 386 unit/mg, respectively. The study found the 300 g/L PEG 6000 + 200 g/L potassium phosphate (PP) and 300 g/L PEG 6000 + 150 g/L sodium phosphate (SP) systems had the highest partitioning efficiency for each salt in primary extraction. However, among the two types of salt, PP displayed higher efficiency than SP, with a partitioning coefficient K of 4.85 vs. 3.89, a volume ratio V of 2.94 vs. 2.68, and a partitioning yield Y of approximately 95 % vs. 83 %. After back extraction, the enzymatic activity of purified chitinase was up to 834 unit/mg (PP) and 492 unit/mg (SP). The purification factors reached 3.32 (PP) and 1.96 (SP), with recovery yields of about 59 % and 61 %, respectively. SDS-PAGE and zymogram analysis showed that the recombinant chitinase was significantly purified by using ATPS. The purified enzyme exhibited high chitinolytic activity, with the hydrolysis zone's diameter being around 2.5 cm–3 cm. It also dramatically reduced the growth of Sclerotium rolfsii; the colony diameter after treatment with 60 unit of enzyme for 104 spores was only about 1 cm, compared to 3.5 cm in the control. The antifungal effect of chitinase suggests that this enzyme has great potential for applications in agricultural production as well as postharvest fruit and vegetable preservation.

Nematode egg parasitic fungus, Purpureocillium lilacinum: efficacy of indigenous strains for the management of Meloidogyne incognita in chickpea

BackgroundNematode egg parasitic fungus, Purpureocillium lilacinum is the most effective biocontrol agent and has been widely used commercially in many countries for the management of root-knot nematode, Meloidogyne incognita. Availability of indigenous potential strains specific to an agro ecosystem is very crucial for their successful commercial exploitation for suppression of nematode population. Hence, an attempt was made to isolate, characterize, evaluate and identify potential indigenous strains of P. lilacinum from pulse ecosystem for root-knot nematode management in chickpea.ResultsThe fungal colony was initially white and when spore was formed it turned into pink colour in 72 to 96 h. Hyphae was hyaline and septate, conidiophore was blunt, and phialides were with wide base and long neck bearing round to oval conidia in chains. Molecular identification of the species, P. lilacinum was carried out based on ITS1-5.8S-ITS2 region of the genomic DNA. In vitro bioassay of cultural filtrates on juvenile mortality revealed that maximum percentage of mortality was observed in IIPR-Pl-11 (88.36%). Spectrophotometric assay on chitinolytic activity showed that the strain IIPR-Pl-11 produced significantly high chitinolytic activity, chitinase enzyme and total protein content (0.139, 51.1 and173.75 µg/ml at 5days); (0.245, 90.1 and 272.67 µg/ml at 10 days) and (0.273, 100.4 and 306.25 µg/ml), respectively, at 15 days of culturing in colloidal chitin-enriched medium C. 2D gel electrophoresis of the crude chitinase suspension showed the presence of chitinase (32, 46 kDa size) in the sample from chitinase-induced medium C. In vitro bioassay of the cultural filtrates of the fungus grown in chitin-enriched medium C on inhibition on egg hatching revealed that the highest percent inhibition on egg hatching showed by IIPR-Pl-8 strain at 5 days of inoculation (42.6%) and IIPR-Pl-11 at 10 and 20 days of inoculation (62.80 and 93.50%), respectively. In vivo pot experiment revealed that among all strains, IIPR-Pl-11 was efficient in promoting plant growth very effectively by reducing gall number (41.3 per plant), egg mass (28.3 per plant) and soil population (284.3 per 200cc of soil) compared to control.ConclusionP. lilacinum strain IIPR-Pl-11 was the highest potential strain from pulse rhizosphere for the management of root-knot nematode, M. incognita in chickpea.

Novel Chitin Deacetylase from Thalassiosira weissflogii Highlights the Potential for Chitin Derivative Production.

β-Chitin is an important carbon fixation product of diatoms, and is the most abundant nitrogen-containing polysaccharide in the ocean. It has potential for widespread application, but the characterization of chitin-related enzymes from β-chitin producers has rarely been reported. In this study, a chitin deacetylase (TwCDA) was retrieved from the Marine Microbial Eukaryote Transcriptome Sequencing Project (MMETSP) database and was heterologously expressed in vitro for functional analysis. The results showed that both the full-length sequence (TwCDA) and the N-terminal truncated sequence (TwCDA-S) had chitin deacetylase and chitinolytic activities after expression in Escherichia coli. High-performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS) indicated that TwCDA and TwCDA-S could catalyze the deacetylation of oligosaccharide (GlcNAc)5. TwCDA had higher deacetylase activity, and also catalyzed the deacetylation of the β-chitin polymer. A dinitrosalicylic acid (DNS) assay showed that TwCDA-S had high chitinolytic activity for (GlcNAc)5, and the optimal reaction temperature was 35 °C. Liquid chromatography combined with time-of-flight mass spectrometry (LC-coTOF-MS) detected the formation of a N-acetylglucosamine monomer (C8H15NO6) in the reaction mixture. Altogether, we isolated a chitin deacetylase from a marine diatom, which can catalyze the deacetylation and degradation of chitin and chitin oligosaccharides. The relevant results lay a foundation for the internal regulation mechanism of chitin metabolism in diatoms and provide a candidate enzyme for the green industrial preparation of chitosan and chitin oligosaccharides.

The Effect of Isolated Bacteria against Adult Stages of Periplaneta americana (Blattodea: Blattidae) and Aedes aegypti (Diptera: Culicidae) using Spraying Methods as a Biological Control

Medical entomology involves the study of medically important insects, such as cockroaches and mosquitoes, which have a dangerous role as transmitters for deadly diseases, such as Malaria, Leishmaniasis, and Dengue fever, which are responsible for many deaths among humans. Huge concern about the use of chemicals insecticides encourages the development of alternative methods for insect control, and due to the harmful effects of these chemicals, new strategies are being developed to replace or reduce the use of synthesized insecticides. Therefore, chitinolytic enzymes produced by microorganisms have a significant effect as biocontrol agents and will be more critical than synthetic pesticides for control. This study was primarily aimed to study the impact of various isolated bacteria using chitinolytic and spraying assays against adult stages of Periplaneta americana and Aedes aegypti as biological controls. Eight species of bacteria were isolated, and only Chryseomonas luteola was used against adult insects because of its high chitinolytic activity by spraying assay. Our results showed that the LC50 values of C. luteola against P. americana were 22.04% and 17.21% after 24 and 48 h, respectively. For A. aegypti adult stages, LC50 values of C. luteola were 2.78% and 2.12% after 24 and 48 h, respectively. Based on the results of this investigation, it is reasonable to say that using microbial insecticides may be an effective strategy to control the adult stages of P. americana and A. aegypti.

Noninsect-Based Diet Leads to Structural and Functional Changes of Acidic Chitinase in Carnivora.

Acidic chitinase (Chia) digests the chitin of insects in the omnivorous stomach and the chitinase activity in carnivorous Chia is significantly lower than that of the omnivorous enzyme. However, mechanistic and evolutionary insights into the functional changes in Chia remain unclear. Here we show that a noninsect-based diet has caused structural and functional changes in Chia during the course of evolution in Carnivora. By creating mouse-dog chimeric Chia proteins and modifying the amino acid sequences, we revealed that F214L and A216G substitutions led to the dog enzyme activation. In 31 Carnivora, Chia was present as a pseudogene with stop codons in the open reading frame (ORF) region. Importantly, the Chia proteins of skunk, meerkat, mongoose, and hyena, which are insect-eating species, showed high chitinolytic activity. The cat Chia pseudogene product was still inactive even after ORF restoration. However, the enzyme was activated by matching the number and position of Cys residues to an active form and by introducing five meerkat Chia residues. Mutations affecting the Chia conformation and activity after pseudogenization have accumulated in the common ancestor of Felidae due to functional constraints. Evolutionary analysis indicates that Chia genes are under relaxed selective constraint in species with noninsect-based diets except for Canidae. These results suggest that there are two types of inactivating processes in Carnivora and that dietary changes affect the structure and activity of Chia.

Robust chitinolytic activity of crab-eating monkey (Macaca fascicularis) acidic chitinase under a broad pH and temperature range

Diet of the crab-eating monkey (Macaca fascicularis) consists of both plants and animals, including chitin-containing organisms such as crabs and insects. This omnivorous monkey has a high expression of acidic chitinase (CHIA) in the stomach and here, we report on its enzymatic properties under different conditions. When we compared with Mus musculus CHIA (Mm-CHIA), Macaca fascicularis CHIA (Mf-CHIA) exhibits higher chitinolytic activity at broad pH (1.0–7.0) and temperature (30–70 ℃) range. Interestingly, at its optimum pH (5.0), Mf-CHIA showed the highest activity at 65 °C while maintaining it at robust levels between 50 and 70 °C. The degradation efficiency of Mf-CHIA was superior to Mm-CHIA toward both polymeric chitin as well as an artificial chromogenic substrate. Our results show that unique features of Mf-CHIA including its thermostability warrant the nomination of this enzyme for potential agricultural and biomedical applications.

Enzymatic activity and virulence of Cordyceps locustiphila (Hypocreales: Cordycipitaceae) on the South American locust Schistocerca cancellata (Orthoptera: Acrididae)

The first barrier, when penetrating an insect host, encountered by microorganisms like fungi is insects cuticle; fungi produce a wide variety of extracellular enzymes involved in the degradation of protein, chitin, and lipids. The main objectives were to assay the enzymatic activity of Cordyceps locustiphila anamorph, in solid and liquid medium at different temperatures and also to evaluate the pathogenicity against the South American locust, Schistocerca cancellata. Conidia of C. locustiphila anamorph were adjusted to 1 × 104, 1 × 106 and 1 × 108 conidia/ml and mortality was recorded for S. cancellata nymphs. The enzymatic activities were determined through a plate test and liquid medium. When assessing the pathogenicity, the fungus caused the highest mortality (84.5 ± 3.5%) at a dose of 1 × 108 conidia/ml. The proteolytic activity showed the highest values (1.56 ± 0.21) (U) at 26 °C. The highest lipolytic activity (1.13 ± 0.36) (U) was observed at 26 °C, while the highest chitinolytic activity was of 0.85 ± 0.1 (U) at 4 °C. Significant differences were observed for enzymatic production on liquid medium, the highest values were recorded for chitinolytic activity (1.63 ± 0.04) (U) and the lowest for caseinolytic activity (0.04 ± 0.001) (U). The results obtained on mortality as well as the quality and variety of enzymes produced by C. locustiphila suggest that this fungus has features that make it a good candidate to be used as a biological agent for the control of the South American locust.

Isolation and characterization of a thermostable alkaline chitinase-producing Aeromonas strain and its potential in biodegradation of shrimp shell wastes

Chitinases are employed to the conversion of chitin and are produced by a wide range of bacteria. The objective of this study was to isolate chitinase-producing microorganisms with high chitinolytic activity. A thermostable alkaline chitinase producing isolate strain CQNU6-2 was obtained from soil samples and showed potential in biodegradation of shrimp shell wastes. The optimal culturing conditions of isolate CQNU6-2 is at 25°C and pH 7 for 24 h. The chitinase produced by strain CQNU6-2 exhibited maximum activity at pH 6.0 and 40°C and it could tolerate the treatment of high temperature (up to 80°C) and high pH (over 10). Taxonomic study, based on biochemical and morphological analysis and phylogenetic analysis of 16S rDNA, showed that strain CQNU6-2 was belongs to the genus Aeromonas sp. The isolate can effectively hydrolyze colloidal chitin with degradation rate of 100% and also can directly degrade the shrimp shells. Ammonium sulfate precipitation method can be used to preliminary purify the chitinase. In conclusion, strain CQNU6-2 had a promising potential for biodegradation of chitin under harsh pH or temperature conditions and could be employed to the comprehensive utilization of shrimp shell wastes.

Chitinase activity potential and identification of chitinolytic bacteria isolated of swimmer crab’s cell

This study aimed at investigating the chitinase enzyme activity produced by chitinolytic bacteria from the skin of blue swimmer crab (Portunus pelagicus) and identification of the genus isolate. This study consists of two stages: firstly, the qualitative and quantitative activity of the chitinase enzyme; and secondly, biochemical identification of the bacteria. The quantitative chitinase enzyme activity is measured using the UV-Vis spectrophotometer UV-Vis at the wavelength at 660 nm. The chitinase enzyme is obtained from the isolation of chitinolytic bacteria cultured within a media to grow solid chitin, which contains colloidal chitin substrate as chitinase inductor at the temperature of 30°C. The highest chitinolytic activity is obtained from the 24 h supernatant culture, with a value of enzyme activity at 0.149 U/mL. Macroscopic and microscopic identification showed that the chitinolytic bacteria isolate R1, whereas the biochemical cell shows the characteristics of the genus Pseudomonas. Keywords: biodegradable, chitinase, spectrophotometer, Portunus pelagicus, Pseudomonas DOI: 10.25165/j.ijabe.20211403.5273 Citation: Sulistijowati R, Sudin, Harmain R M. Chitinase activity potential and identification of chitinolytic bacteria isolated of swimmer crab’s cell. Int J Agric & Biol Eng, 2021; 14(3): 228–231.

Immobilization of Bacterial Cells and Chitinolytic Activity ofStreptomycessp. (PB 2)

This study was aimed to immobilize the chitinolytic bacteriaStreptomycessp. PB 2 using sodium alginate as immobilization agent.Streptomycessp. PB 2 was reported have a good chitinolytic activity and immobilization is known to increase the stability of bacteria during repeated used. Sodium alginate has been used on several methods of immobilization and sodium alginate are reported as a good agent. Cell immobilization was done by growing the cell on nutrient broth (NB), mixed with 1% sodium alginate to form the beads, then the mixture were put to a syringe and droped into CaCl2 solution. The beads were washed with 0.85% NaCl solution and filtered. To test the bacterial activity, beads were applied into chitin liquid medium and the chitinolytic activity was observed every 24 hours for 5 days. To test its stability, after the 5thday, the beads was filtered and put in to chitin liquid medium and tested the chitinolytic activity. This experiment was repeated twice. The highest chitinolytic activity ofStreptomycessp. PB2 was observed on the 4thday with the value of 0.00014 U/ml and the concentration of NAG of 5.42087 µg/ml with the treatment of immobilized NB medium without the addition of chitin.

A Cold-Adapted Chitinase-Producing Bacterium from Antarctica and Its Potential in Biocontrol of Plant Pathogenic Fungi.

To obtain chitinase-producing microorganisms with high chitinolytic activity at low temperature, samples collected from Fildes Peninsula in Antarctica were used as sources for bioprospecting of chitinolytic microorganisms. A cold-adapted strain, designated as GWSMS-1, was isolated from marine sediment and further characterized as Pseudomonas. To improve the chitinase production, one-factor-at-a-time and orthogonal test approaches were adopted to optimize the medium components and culture conditions. The results showed that the highest chitinolytic activity (6.36 times higher than that before optimization) was obtained with 95.41 U L−1 with 15 g L−1 of glucose, 1 g L−1 of peptone, 15 g L−1 of colloid chitin and 0.25 g L−1 of magnesium ions contained in the medium, cultivated under pH 7.0 and a temperature of 20 °C. To better understand the application potential of this strain, the enzymatic properties and the antifungal activity of the crude chitinase secreted by the strain were further investigated. The crude enzyme showed the maximum catalytic activity at 35 °C and pH 4.5, and it also exhibited excellent low-temperature activity, which still displayed more than 50% of its maximal activity at 0 °C. Furthermore, the crude chitinase showed significant inhibition of fungi Verticillium dahlia CICC 2534 and Fusarium oxysporum f. sp. cucumerinum CICC 2532, which can cause cotton wilt and cucumber blight, respectively, suggesting that strain GWSMS-1 could be a competitive candidate for biological control in agriculture, especially at low temperature.

High Production of Chitinolytic Activity in Halophilic Conditions by a New Marine Strain of Clonostachys rosea.

Twenty-eight fungal strains have been isolated from different natural marine substrates and plate screened for their production of chitinolytic activity. The two apparent best producers, Trichoderma lixii IG127 and Clonostachys rosea IG119, were screened in shaken cultures in media containing 1% colloidal chitin, 1% yeast nitrogen base and 38‰ NaCl, for their ability to produce chitinolytic enzymes under halophilic conditions. In addition, they were tested for optimal growth conditions with respect to pH, salinity and temperature. The Trichoderma strain appeared to be a slight halotolerant fungus, while C. rosea IG119 clearly showed to be a halophilic marine fungus, its optimal growth conditions being very coherent for life in the marine environment (i.e., pH 8.0, salinity 38‰). Due to its high and relatively fast activity (258 U/L after 192 h of growth) accompanied by its halophilic behaviour (growth from 0 to 160‰ of salinity), C. rosea was selected for further studies. In view of possible industrial applications, its medium for chitinolytic enzyme production was optimized by Response Surface Methodology using 1% colloidal chitin and different concentrations of corn step liquor and yeast nitrogen base (0–0.5%). Time course of growth under optimized condition showed that maximum activity (394 U/L) was recorded after 120 h on medium containing Corn Steep Liquor 0.47% and Yeast Nitrogen Base 0.37%. Maximum of productivity (3.3 U/Lh) was recorded at the same incubation time. This was the first study that demonstrated high chitinolytic activity in a marine strain of C. rosea.

Entomopathogenic efficacy of the chitinolytic bacteria: Aeromonas hydrophila isolated from Siwa Oasis, Egypt

Thirty-four bacterial isolates were isolated from soil samples collected from the North Western Coast and a water sample collected from brackish water at Siwa Oasis, Matrouh Governorate, Egypt. Only six isolates showed chitinase activity when screened on colloidal chitin agar medium. The highest chitinolytic activity was achieved by a bacterial isolate labeled as A.S. This isolate was identified as Aeromonas hydrophila based on analysis of 16S rRNA gene sequence and morphological, physiological, and biochemical characteristics. Optimization of the cultural conditions for maximum chitinase production by A. hydrophila revealed that the highest level of chitinase was recorded when the bacterium was grown in malt nitrogen-based medium containing 1% colloidal chitin at pH7 for 48 h incubation at 30 °C. Crude chitinase from isolate A. hydrophila was evaluated against first instar larvae of the greater wax moth; Galleria mellonella L. (Lepidoptera: Pyralidae) at different concentrations of 0, 185, 205, 235, 265, 295 U/mg protein. It increased larval and pupal mortality rates in a concentration-dependent manner. The tested crude chitinase significantly induced a decrease in adults’ emergence rate and their fecundity.

Isolation and identification of marine strains of Stenotrophomona maltophilia with high chitinolytic activity.

Chitin is the second most abundant organic compound in nature and represents a rich carbon and nitrogen source that is primarily transformed by bacterial communities. Bacteria capable of gradually hydrolyzing chitin into N-acetylglucosamine monomers can have applications in the transformation of residues from shrimp and other crustaceans. The objective of the present study was to isolate, characterize and identify microorganisms with high chitinolytic activity. These microorganisms were isolated and characterized based on macro- and microscopic morphological traits. Strains were selected on colloidal chitin agar medium primarily based on a hydrolysis halo larger than 2 mm and a growing phase no longer than 6 days. Secondary selection consisted of semi-quantitative evaluation of chitinolytic activity with a drop dilution assay. From the above, ten strains were selected. Then, strain-specific activity was evaluated. The B4 strain showed the highest specific activity, which was 6,677.07 U/mg protein. Molecular identification indicated that the isolated strains belong to the species Stenotrophomonas maltophilia.

Biocontrol of Root-Knot Nematode Meloidogyne incognita by Chitinolytic Trichoderma spp.

The inhibitory potential of chitinolytic Trichoderma spp. was assayed against Meloidogyne incognita in vitro and in vivo. Sixteen Trichoderma isolates were assayed for chitinase activity using basal solid medium supplemented with colloidal chitin. Isolates recorded high chitinolytic activity were identified morphologically as Trichoderma asperelloides, T. hamatum, T. harzianum and T. viride. The four species were quantitatively assayed for total and specific chitinase activity. T. asperelloides showed maximum chitinase activity (1.736 U/mL). In vitro studies revealed that egg hatching and second juveniles vitality of root-knot nematodes were significantly inhibited by chitinase-containing growth culture of the four selected chitinolytic Trichoderma species. However, the reduction effect was directly correlated with the concentration of fungal growth suspension. Under greenhouse conditions, three sequentially treatments of soil with homogenized culture suspension were studied. Trichoderma species significantly reduced the final nematode population by 97.81% and root galling by 92.81%. Moreover, all tested Trichoderma species obviously promoted tomato growth compared to untreated control. Significant positive correlation between the biocontrol efficacy of Trichoderma species against root-knot nematodes and their chitinolytic activity was confirmed. The most potential chitinolytic species, T. asperelloides, also recorded the highest biocontrol activity against M. incognita.

Occurrence of chitinolytic bacteria in shrimp rusip and measurement of their chitin-degrading enzyme activities

Puspita ID, Wardani A, Puspitasari ROA, Nugraheni PS, Putra MMP, Pudjiraharti S, Ustadi. 2017. Occurrence of chitinolytic bacteria in shrimp rusip and measurement of their chitin-degrading enzyme activities. Biodiversitas 18: 1275-1281. The objectives of this research were to isolate chitinolytic bacteria from shrimp rusip (an Indonesian traditional fermented shrimp product), identify bacterial isolates showing high chitinolytic activity, and determine the chitinolytic activity of these isolates. There were 44 chitinolytic bacteria isolated from shrimp rusip: 39 isolates of Gram-positive bacteria and 5 isolates of Gram-negative bacteria. The quantitative method we used to evaluate chitin-degrading enzyme activity measured the amount of N-acetylglucosamine produced from the reaction of crude enzyme and colloidal chitin. Seven isolates showing highest chitinolytic activity were Bacillus cereus (KKT 1, KKT 14, and KKT 19), Bacillus thuringiensis (KKT 6), Enterobacter cloacae (LCK 20), Pseudomonas stutzeri (LCK 17), and Stenotrophomonas maltophilia (THK1). The highest chitinolytic activity showed by KKT 1, KKT 6, KKT 14, KKT 19, and THK 1 were 0.0016 U/mL (at 72 hours); 0.0018 U/mL (at 48 hours); 0.0019 U/mL (at 48 hours); 0.0033 U/mL (at 48 hours); and 0.0032 U/mL (at 24 hours), respectively.

Skrining dan Evaluasi Aktivitas Kitinase dari Sembilan Isolat Bakteri Lokal

Chitinase is an enzyme that degrades chitin on ?-1,4-acetamido-2-deoxy-D-glycoside bound to produce Chitin oligosaccharide or its soluble monomer, N-acetylglucosamine (NAG). Chitinase are usually produced naturally by chitinolytic microorganism that are found in soil or water environment. Chitinase have been widely used for biocontrol agents of plant pests. The product from chitinase hydrolysis can be used as antiinflammatory agent and anti hyperpigmentation. This research was aimed to screen and evaluate the chitinolytic ability of 9 isolate. The best isolate was the one which produced high chitinolytic activity and hydrolyzed chitin into NAG. 9 isolate were examined and BPPTCC-2 was the best isolate which produced highest level of chitinolytic activity 0.1780 U/ml. The chitinase was able to hydrolyzed chitin resulting high yields of NAG 11.5% as the only product.

Characterization of the structure of the prokaryotic complex of Antarctic permafrost by molecular genetic techniques

A prokaryotic mesophilic organotrophic community responsible for 10% of the total microbial number determined by epifluorescence microscopy was reactivated in the samples ofAntarctic permafrost retrieved from the environment favoring long-term preservation of microbial communities (7500 years). No culturable forms were obtained without resuscitation procedures (CFU = 0). Proteobacteria, Actinobacteria, and Firmicutes were the dominant microbial groups in the complex. Initiation of the reactivated microbial complex by addition of chitin (0.1% wt/vol) resulted in an increased share of metabolically active biomass (up to 50%) due to the functional domination of chitinolytics caused by the target resource. Thus, sequential application of resuscitation procedures and initiation of a specific physiological group (in this case, chitinolytics) to a permafrost-preserved microbial community made it possible to reveal a prokaryotic complex capable of reversion of metabolic activity (FISH data), to determine its phylogenetic structure by metagenomic anal-ysis, and to isolate a pure culture of the dominant microorganism with high chitinolytic activity.

MYCOLYTIC ACTIVITY OF PHOSPHATE MOBILIZING SOILBA CTERIA OF GENUS BACILLUS COHN

The aim of this research was study of the chitinolytic activity of phosphateobilizing soil bacteria of genus Bacillus and their antagonistic properties towards microscopic fungi. Methods. In this research 16 strains of phosphate-mobilizing soil bacteria of genus Bacillus and saprophytic and plant pathogenic strains of micrtomycetes were investigated. The antagonistic activity of bacilli against micromycetes was determined by the method of blocks and holes. Bacilli on agar medium and also their supernatants were used. Inhibition of fungal growth under co-cultivation with bacilli under static conditions in the liquid culture medium was determined gravimetrically by the difference of dry weight mycelium in control and in experiment. Bacilli chitinolytic activity was determined photocolorimetrically by the formation of reducing sugars by hydrolysis of colloidal chitin using 3,5-dinitrosalicylic acid. Student test for 5% significance level was used for statistical processing of the obtained data. Results. The presence of chitinolytic activity in all tested bacilli in colloidal chitin medium was established. B. megaterium 16, B. pumilus 7, B. cereus v mycoides 10 strains showed high chitinolytic activity, they were also characterized by high antagonistic activity against plant pathogenic fungi (Fusarium oxysporum 54 and Trichothecium roseum 658). Bacilli were capable to using autoclaved mycelium as the sole of carbon and nitrogen source. Cultivation of bacteria and microscopic fungi under static conditions showed growth inhibition and changes in morphological features – blistering and areas of lysis of mycelium. Conclusions. It was established that phosphate-mobilizing strains of soil bacteria of the genus Bacillus showed chitinolytic activity and growth inhibition of plant pathogens and caused their morphological changes in co-culture. The direct relationship between the antagonistic and chitinolytic activity of bacilli was demonstrated. The bacilli ability to use autoclaved fungal mycelium as a sole source of carbon and nitrogen was showed on the example of B. cereus v. mycoides 10.