Increase In Chitinase Activity Research Articles

Dehydrated and live black soldier fly larvae as environmental enrichment in indigenous slow-growing chickens: performance, gut health, and chitinolytic enzyme activity

The demand for sustainable and ethically farmed animal products is on the rise as consumers become more environmentally and animal welfare conscious. The need to diminish the consumption of soybean meal is urgent, and companies are looking for ways to respond to this necessity by looking for alternatives to soybean meal. This study assessed the impact of introducing whole dehydrated and live black soldier fly larvae (BSFL) into the diet of an indigenous chicken breed as environmental enrichment. A total of 144 39-day-old male Bianca di Saluzzo chickens were distributed among 18 pens and assigned to three different experimental groups. The control group received a diet where soybean meal was entirely replaced by alternative ingredients. The two experimental groups were given the same diet supplemented with 5% of the expected daily feed intake of whole dehydrated BSFL or whole live BSFL. Throughout the trial period (from the bird age of 39–174 days of age), live weight was recorded every 21 days, and the average daily gain, daily feed intake, and feed conversion ratio were calculated. The time required for the birds to consume the larvae was recorded three times a week. At age 147 and 174 days, 12 birds per treatment were selected based on mean live weight and slaughtered. Measurements included hot and chilled carcass weights, organ weights (spleen, liver, heart, stomach), breast and thigh muscle weights, and the corresponding yields were calculated. Acid protease activity was measured in proventriculus extract, and chitinase and chitosanase activity was calculated based on the release of reducing sugars from chitin or chitosan. The results showed little improvement in final live weights and daily feed intakes of the animals fed the insect larvae compared with control birds. Larva supplementation had no negative impact on the overall well-being of the animals assessed by blood analysis and histopathological assessment of the intestinal tract, spleen, and liver. No differences were found between the dehydrated vs live insect larvae consumption times, with all larvae being eaten up very rapidly (< 3 min). The birds fed BSFL showed an increase in chitinase activity. These findings support the potential use of whole BSFL as a form of environmental enrichment, particularly in their dehydrated form, being more convenient to use and store, which would also encourage the uptake of this practice by farmers.

Chemical induction of DNA demethylation by 5-Azacytidine enhances tomato fruit defense against gray mold through dicer-like protein DCL2c.

Postharvest decay, primarily caused by pathogenic fungi in ripening fruits and fresh vegetables, poses a challenge to agricultural sustainability and results in significant economic losses. The regulation of the fruit ripening by DNA methylation has been well demonstrated, while defense response of fruit underlying epigenetic regulation against postharvest decay remains uncertain. In the present study, treatment of tomato fruits with the DNA methyltransferase inhibitor 5-Azacytidine (5-Aza) notably decreased their susceptibility to gray mold. Following 5-Aza treatment, we observed a substantial increase in activities of chitinase (CHI) and glucanase (GLU) in tomato fruits, as well as an increase in the expression of the dicer-like SlDCL2 gene family. Suppression of SlDCL2c through double-stranded RNA-induced RNA interference (RNAi) resulted in a decrease in the expression of chitinases CHI3, CHI9, Class V chitinase, and endochitinase 4 by 71%, 29%, 55%, 64%, as well as glucanases Cel1, Cel2, and GluB by 19%, 93%, and 87%, respectively. This was accompanied by decreased activities of resistance-related enzymes, including CHI and GLU. The expression levels of genes phenylalanine ammonia-lyase PAL2, peroxidase POD 12, POD P7, CCR1, CYP84A2, and COMT in phenylpropanoid biosynthesis pathway also decreased by 33%, 53%, 18%, 50%, 30%, and 24% in SlDCL2c-RNAi fruit, resulting in decreased activities of PAL and POD. Consequently, the lesion diameter of gray mold in SlDCL2c-RNAi fruit increased by 55% compared to the control group. Overall, the present study indicated that DNA methyltransferase inhibitor 5-Aza reduces susceptibility of tomato fruit to gray mold through regulation of DCL2c-mediated inducible defense response.

Enhanced antifungal activity of sugarcane cv. NCo310 expressing chimeric chitinase 42

The development of sustainable and environmentally safe strategies for crop protection against fungal pathogens is a significant challenge in agriculture. This paper discusses the successful transfer of a chimeric chitinase 42 gene into sugarcane using the biolistic method, resulting in an enhanced antifungal activity. Chitinases possess the enzymatic capability to degrade chitin, a key component of the cell walls in various fungal phytopathogens. In this research, we employed a chimeric chitinase 42 that combines the 42 kDa chitinase from Trichoderma atroviridae with a chitin binding domain from Serratia marcescens chitinase B. This chimeric chitinase exhibits higher antifungal activity against phytopathogenic fungi. PCR analysis confirmed the integration of the transgene into the genome of transformants, while the expression of the transgene was assessed using reverse transcription-PCR. Transgenic lines showed to have different chitinase activity and a high increase in chitinase activity was detected in lines T3, T5 and T6 with about 18 U/mg chitinase activity compared to the control plants (2.8 U/mg). Antifungal activity of transgenic lines T3, T5 and T6 on sugarcane fungal pathogens was shown in radial diffusion and detached leaf assays. The enhanced antifungal activity observed in the transformed sugarcane lines indicates the potential of chimeric chitinase 42 as an effective antifungal agent.

Chitinase induction in Trichoderma harzianum: a solid-state fermentation approach using shrimp waste and wheat bran/commercial chitin for chitooligosaccharides synthesis

This study innovatively employed solid-state fermentation (SSF) to evaluate chitinase induction in Trichoderma harzianum. Solid-state fermentation minimizes water usage, a crucial global resource, and was applied using shrimp waste chitin and a mixture of commercial chitin with wheat bran as substrates. Shrimp waste and wheat bran were pretreated and characterized for SSF, and the fungus’s utilization of the substrates was assessed using spectrophotometric and microscopic methods. The resulting enzymes’ ability to produce chitooligosaccharides (COS) mixtures was studied. Wheat bran/commercial chitin demonstrated superior performance, with a 1.8-fold increase in chitinase activity (76.3 U/mg protein) compared to shrimp waste chitin (41.8 U/mg protein). Additionally, the COS mixture obtained from wheat bran/commercial chitin showed a higher concentration of reducing sugars, reaching 87.85 mM, compared to shrimp waste chitin (14.87 mM). The COS profile from wheat bran/commercial chitin included monomers to heptamers, while the profile from shrimp waste chitin was predominantly composed of monomers. These results highlight the advantages of SSF for chitinase induction and COS production in T. harzianum, offering potential applications as dietary fiber, antioxidants, and antimicrobial agents. The findings contribute to by-product valorization, waste reduction, and the sustainable generation of valuable products through SSF-based enzyme production.

Combining in vitro and in silico studies to unravel the antifungal potential of chitinase from a marine bacterial isolate Cellulosimicrobium cellulans RS7

The main aim of this research is to identify, screen and optimize physio-chemical characteristics for the production of extracellular chitinase from a marine bacterial isolate RS7. The bacterium isolated from Pamban Island, Rameshwaram was characterized as Gram-positive, pleomorphic and was identified as Cellulosimicrobium cellulans RS7 based on 16S rRNA profiling. The growth profile analysis showed maximum chitinase activity at 48 h of inoculum age. The optimization of physio-chemical parameters was performed using OFAT and statistical approach via RSM which resulted in 3.23-fold increase in chitinase activity from 20.89 to 67.6 U/ml. This study investigates the anti-fungal activity of chitinase against pathogenic fungus Mucor sp. where 80 μl (5 U) of enzyme resulted in a 22 mm zone of inhibition. Furthermore, molecular docking studies were done to understand the active properties of chitinase against fungal pathogens. The Molecular Operating Environment module was utilized and the docking poses were visually examined for interaction. The results indicated that chitinase exhibited high affinity towards selected ligand/protein (N-acetylglucosamine, β-1, 3 glucanase and β-1, 3 glucan) which participate in synthesis of fungal cell wall. The docking scores and root mean square deviation values for the selected target molecules were determined and specific interactions and residues were identified.

Biochemical purification and characterization of a truncated acidic, thermostable chitinase from marine fungus for N-acetylglucosamine production.

N-acetylglucosamine (GlcNAc) is widely used in nutritional supplement and is generally produced from chitin using chitinases. While most GlcNAc is produced from colloidal chitin, it is essential that chitinases be acidic enzymes. Herein, we characterized an acidic, highly salinity tolerance and thermostable chitinase AfChiJ, identified from the marine fungus Aspergillus fumigatus df673. Using AlphaFold2 structural prediction, a truncated Δ30AfChiJ was heterologously expressed in E. coli and successfully purified. It was also found that it is active in colloidal chitin, with an optimal temperature of 45°C, an optimal pH of 4.0, and an optimal salt concentration of 3% NaCl. Below 45°C, it was sound over a wide pH range of 2.0–6.0 and maintained high activity (≥97.96%) in 1–7% NaCl. A notable increase in chitinase activity was observed of Δ30AfChiJ by the addition of Mg2+, Ba2+, urea, and chloroform. AfChiJ first decomposed colloidal chitin to generate mainly N-acetyl chitobioase, which was successively converted to its monomer GlcNAc. This indicated that AfChiJ is a bifunctional enzyme, composed of chitobiosidase and β-N-acetylglucosaminidase. Our result suggested that AfChiJ likely has the potential to convert chitin-containing biomass into high-value added GlcNAc.

Production of &lt;em&gt;Aspergillus quadrilineatus&lt;/em&gt; MT083999 Chitinase, β-1,3-Glucanase and Nano-silver Important for Biocontrol of &lt;em&gt;Fusarium&lt;/em&gt; spp&lt;em&gt;.&lt;/em&gt; Infecting Crops

In our search for an active biocontrol agent against F. solani and F. oxysporum , the major species causing wilt, root rot and damping-off in plants affecting crops. The fungal isolate genetically identified as Aspergillus quadrilineatus MT083999 displayed high antagonistic activities against the tested root-rot fungi, and presented valuable chitinase and β-glucanase activities of 235.484 and 508.953 U/gds, respectively. Solid state fermentation conditions were optimized by applying Three–factor–five–level central composite design (CCD) with twenty experiments and two responses of chitinase R1 and β-glucanase R2 activities, respectively. The optimum activities of chitinase (386.70 U/gds) and β-glucanase (1094.70 U/gds) were obtained in trial 4 and 2 respectively, with about 1.64-fold increase in chitinase activity and 2.15-fold increase in β-glucanase activity when compared with initial fermentation. Biosynthesis of nano-silver from A. quadrilineatus was evaluated in 10 experiments of another design of 2–factors–5–levels. The antifungal effects of each biosynthesized nano-silver against F. solani (R1) and F. oxysporum (R2) were evaluated as Inhibition zone diameter (IZD). The high fungicide nano-silver against both tested phytopathogens was characterized by UV–visible spectrophotometry, transmission electron microscopy (TEM), Zeta dynamic light scatter (DLS) and Fourier transform infrared spectroscopy (FTIR). The results indicated that A. quadrilineatus MT083999 chitinolytic enzymes and nano-silver can be applied as a novel candidate in biocontrol of Fusarium wilt infecting crops worldwide.

A major release of urban untreated wastewaters in the St. Lawrence River (Quebec, Canada) altered growth, reproduction, and redox status in experimentally exposed Daphnia magna

In 2015, five billion liters of untreated urban wastewater (UWW) were released into the St. Lawrence River (Quebec, Canada) over the course of four days in order to repair the Montreal's sewer interceptor network related to the city's primary wastewater treatment plant. The UWW discharge originated mainly from household, industrial, and hospital sources. The objective of this study was to investigate the toxicological effects of this unprecedented punctual UWW release on aquatic invertebrates to gather information that could help understand the potential impacts to the receiving environment of overflow episodes occurring during heavy rain events. Water samples were collected at four impacted and non-impacted sites during and four weeks after the release. The freshwater crustacean Daphnia magna were experimentally exposed to surface water collected from UWW-impacted sites for 13 days and analyzed for life-history endpoints and suitable biomarkers related to oxidative stress (i.e., catalase, superoxide dismutase, lipid peroxidation, and glutathione-s-transferase) and reproduction (chitinase). Results indicated that D. magna growth and reproduction were significantly increased by exposure to UWWs. These effects were correlated with an increase in chitinase activity, which is primarily controlled by reproductive hormones and involved in growth, suggesting potential impacts on these processes. Results also indicated that some UWW samples might have caused oxidative stress during the release but that it was overcome by antioxidant defenses and did not lead to cellular damage. Overall, current results contribute to a better understanding of the biological impacts of UWW to aquatic invertebrates for a better stormwater management.

Study of ChiR function in Serratia marcescens and its application for improving 2,3-butanediol from crystal chitin.

Microbial utilization of chitin, a potential renewable biomass feedstock, is being pursued as a means of developing novel consolidated bioprocessing for the production of chemicals. Serratia marcescens is a gram-negative bacterium that is known for its chitinolytic capability and as a native 2,3-butanediol producer. In S. marcescens, ChiR has been suggested to be a positive regulator of chitinase production. In this study, we aim to understand the effect of ChiR in regulating nine chitinase-related genes in S. marcescens Db11 and demonstrate manipulation of chiR as a useful and efficient genetic target to enhance chitin utilization. First, a chiR overexpression (chiROE) strain and a chiR deletion (ΔchiR) strain were generated and characterized in terms of cellular growth, chitinase activity, and total secreted protein. Compared to the wild-type Db11 strain, the S. marcescens chiROE strain showed an increase in chitinase activity (2.14- to 6.31-fold increase). Increased transcriptional expression of chitinase-related genes was measured using real-time PCR, showing 2.12- to 10.93-fold increases. The S. marcescens ΔchiR strain showed decreases in chitinase activity (4.5- to 25-fold decrease), confirming ChiR's role as a positive regulator of chitinase expression. Finally, chiR overexpression was investigated as a means of increasing biochemical production (2,3-butanediol) from crystal chitin. The chiROE strain produced 1.13±0.08g/L 2,3-butanediol from 2% crystal chitin, a 2.83-fold improvement from the wild-type strain, indicating ChiR is an important and useful genetic engineering target for enhancing chitin utilization in S. marcescens.

EFFICIENCY OF BIOCIDES, FUNGICIDES AND PLANT EXTRACTS AGAINST FUSARIUM SOLANI ON TOMATO PLANTS

Tomato is one of the most important economic vegetable crops which are attacked by several serious diseases such as root-rot. The effect of biocides (Bio-Zeid, Bio-Arc, Plant guard and Clean-Root), fungicides (Rhizolex-T, Vitavax 200 and Maxiam) and plant extracts (Eucalyptus globules, Allium cepa, Datura spp, and Mentha viridis) on growth of Fusarium solani fungus in vitro and on root-rot disease of tomato caused by Fusarium Solani and their effect on biochemical changes were studied under greenhouse conditions. In vitro, all tested biocides were able to inhibit growth of F. solani. Seed treatment with biocides decreased percentage of disease incidence compared with control. Bio-Zeid was the most effective compared with other biocides. All fungicides were able to inhibit linear growth of F. solani compared to the control. Rhizolex-T and Vitavax 200 were the most effective fungicides treatments followed by Maxiam. All water extracts of camphor, onion, Datura and Mint were able to inhibit growth of F. solani compared to the control. Seed of tomato treatment with plant extract decreased disease incidence. Water extract of onion plants was the most effective compared with other plant extracts. Also, all treatments effected on enzymes activity, Bio-Zeid gave the highest increment in peroxidase followed by Bio-Arc and Rhizolex-T. Where, onion gave the highest increase in polyphenloxidase activity followed by Bio-Zeid. Also it gave the highest increase in chitinase activity followed by Rhizolex-T compared to other treatments.

Effect of ozone and antioxidants on wheat and its pathogen — Bipolaris sorokininana

Tropospheric ozone (O3) adversely affects growth and productivity of crops and also influences crop–pathogen interactions. Adverse effects of O3 on crops can be mitigated by antioxidants application. In the present study through lab and field experiments impact of O3 and antioxidants treatment to wheat was assessed on growth of Bipolaris sorokiniana (BS-75 strain) pathogen responsible for Spot blotch disease, pathogenesis related (PR) proteins and chitinase content. Results showed that growth of Bipolaris was significantly higher in elevated ozone (EO3) exposed plants as compared to control plants. Antioxidants – ascorbic acid (AA), tagetes extract (T) and quercetin (Q) application on culture media and wheat plants, respectively, retarded the growth of Bipolaris sorokiniana. Among the three antioxidants minimum growth of Bipolaris was observed in AA-treated plants as compared to control plants. Reduction in chitinase activity and PR proteins content due to EO3 treatment in wheat plants was 18% and 78%, respectively, as compared to control plants. Increase in chitinase activity and PR proteins content due to antioxidants treatment in wheat plants was 45% and 60%, respectively, as compared to control plants.

Enhancement of Exochitinase Production by Bacillus licheniformis AT6 Strain and Improvement of N-Acetylglucosamine Production.

A strain producing chitinase, isolated from potato stem tissue, was identified as Bacillus licheniformis by biochemical properties and 16S RNA sequence analysis. Statistical experimental designs were used to optimize nine independent variables for chitinase production by B. licheniformis AT6 strain in submerged fermentation. Using Plackett-Burman design, (NH4)2SO4, MgSO4.7H2O, colloidal chitin, MnCl2 2H2O, and temperature were found to influence chitinase production significantly. According to Box-Behnken response surface methodology, the optimal fermentation conditions allowing maximum chitinase production were (in gram per liter): (NH4)2SO4, 7; K2HPO4, 1; NaCl, 1; MgSO4.7H2O, 0.1; yeast extract, 0.5; colloidal chitin, 7.5; MnCl2.2H2O, 0.2; temperature 35°C; pH medium 7. The optimization strategy led to a 10-fold increase in chitinase activity (505.26±22.223mU/mL versus 50.35±19.62mU/mL for control basal medium). A major protein band with a molecular weight of 61.9kDa corresponding to chitinase activity was clearly detected under optimized conditions. Chitinase activity produced in optimized medium mainly releases N-acetyl glucosamine (GlcNAc) monomer from colloidal chitin. This enzyme also acts as an exochitinase with β-N-acetylglucosaminidase. These results suggest that B. licheniformis AT6 secreting exochitinase is highly efficient in GlcNAc production which could in turn be envisaged as a therapeutic agent or as a conservator against the alteration of several ailments.

Evaluation of Chitinase Activity in Tall Fescue Cultivars Inoculated with Rhizoctonia solani

Tall fescue has great utility as a low maintenance turfgrass in the northern and transition zone regions of the United States. A factor limiting tall fescue utility is its susceptibility to the pathogen Rhizoctonia solani Kuhn, the causal agent of foliar brown patch. Chitinase activity has been positively correlated with resistance to R. solani in other plant species. A chitinase assay was developed for tall fescue. Three tall fescue cultivars with differing agronomic qualities and brown patch susceptibility as well as a resistant hybrid bluegrass cultivar were inoculated with R. solani in a greenhouse humidity chamber. Chitinase activity 48 hours after inoculation was negatively correlated with percent brown patch severity 10 days after inoculation. ‘Jaguar’ tall fescue was the most tolerant to R. solani and exhibited the highest chitinase activity before and after inoculation. No significant increase in chitinase activity was observed in the other tall fescue cultivars following R. solani inoculation. Identifying tall fescue cultivars expressing high amounts of chitinase activity could be important for developing brown patch-tolerant tall fescue cultivars.

Naturally produced citral can significantly inhibit normal physiology and induce cytotoxicity on Magnaporthe grisea

Given the importance of finding alternatives to synthetic fungicides, the antifungal effects of natural product citral on six plant pathogenic fungi (Magnaporthe grisea, Gibberella zeae, Fusarium oxysporum, Valsa mali, Botrytis cinerea, and Rhizoctonia solani) were determined. Mycelial growth rate results showed that citral possessed high antifungal activities on those test fungi with EC50 values ranging from 39.52 to 193.00 µg/mL, which had the highest inhibition rates against M. grisea. Further action mechanism of citral on M. grisea was carried out. Citral treatment was found to alter the morphology of M. grisea hyphae by causing a loss of cytoplasm and distortion of mycelia. Moreover, citral was able to induce an increase in chitinase activity in M. grisea, indicating disruption of the cell wall. These results indicate that citral may act by disrupting cell wall integrity and membrane permeability, thus resulting in physiology changes and causing cytotoxicity. Importantly, the inhibitory effect of citral on M. grisea appears to be associated with its effects on mycelia reducing sugar, soluble protein, chitinase activity, pyruvate content, and malondialdehyde content.

The natural product citral can cause significant damage to the hyphal cell walls of Magnaporthe grisea.

In order to find a natural alternative to the synthetic fungicides currently used against the devastating rice blast fungus, Magnaporthe grisea, this study explored the antifungal potential of citral and its mechanism of action. It was found that citral not only inhibited hyphal growth of M. grisea, but also caused a series of marked hyphal morphological and structural alterations. Specifically, citral was tested for antifungal activity against M. grisea in vitro and was found to significantly inhibit colony development and mycelial growth with IC50 and IC90 values of 40.71 and 203.75 μg/mL, respectively. Furthermore, citral reduced spore germination and germ tube length in a concentration-dependent manner. Following exposure to citral, the hyphal cell surface became wrinkled with folds and cell breakage that were observed under scanning electron microscopy (SEM). There was damage to hyphal cell walls and membrane structures, loss of villous-like material outside of the cell wall, thinning of the cell wall, and discontinuities formed in the cell membrane following treatment based on transmission electron microscopy (TEM). This increase in chitinase activity both supports the morphological changes seen in the hyphae, and also suggests a mechanism of action. In conclusion, citral has strong antifungal properties, and treatment with this compound is capable of causing significant damage to the hyphal cell walls of M. grisea.

Enhanced biocontrol activity of Rhodotorula mucilaginosa cultured in media containing chitosan against postharvest diseases in strawberries: possible mechanisms underlying the effect.

The effect of Rhodotorula mucilaginosa cultured in media containing chitosan on its antogonistic activity against postharvest diseases of strawberries and the possible mechanisms involved are discussed. Two-dimensional gel electrophoresis were applied in the analysis of the proteins of R. mucilaginosa in response to chitosan. Compared with the application of R. mucilaginosa alone, the biocontrol efficacy of the yeast combined with 0.5% chitosan was enhanced greatly, with significant increase in chitinase activity of antagonistic yeast, polyphenoloxidase, peroxidase, phenylalanine ammonia lyase, chitinase and β-1,3-glucanase activity, and with an inhibition of lipid peroxidation of strawberries. The population of R. mucilaginosa harvested from NYDB amended with chitosan at 0.5% increased rapidly in strawberry wounds compared with those harvested from NYDB without chitosan. In the cellular proteome, several differentially expressed proteins were identified, most of which were related to basic metabolism.

Enzymatic production of N-acetyl-D-glucosamine by solid state fermentation of chitinase by Penicillium ochrochloron MTCC 517 using agricultural residues

The optimization studies for production of chitinase were carried out by response surface methodology (RSM) based on statistics experimental design using three substrates, which were wheat, rice and red gram bran. 24 full factorial central composite design was applied to evaluate optimal combinations of variables. These variables were chitin concentration, initial moisture content, inoculum level, and incubation time. The results of second order polynomial showed that all four variables had significant effect on chitinase production. Maximum chitinase activity was recorded for wheat bran (2443.23 U g−1) than rice (1216.65 U g−1) and red gram bran (961.32 U g−1). An overall 3-fold increase in chitinase activity was achieved using optimized strategies of RSM. Growth of the fungus on all bran particles have been visualized by scanning electron microscopy. These results indicated the potential of Penicillium ochrochloron for economical production of chitinase using agricultural residues. TLC and HPLC analysis of colloidal chitin hydrolysate with partially purified chitinases revealed that the major reaction product was monomeric GlcNAc indicating the potential of these enzymes for efficient production of GlcNAc.

Evidence Supporting a Role for Mammalian Chitinases in Efficacy of Caspofungin against Experimental Aspergillosis in Immunocompromised Rats

ObjectivesCaspofungin, currently used as salvage therapy for invasive pulmonary aspergillosis (IPA), strangely only causes morphological changes in fungal growth in vitro but does not inhibit the growth. In vivo it has good efficacy. Therefore the question arises how this in vivo activity is reached. Caspofungin is known to increase the amount of chitin in the fungal cell wall. Mammals produce two chitinases, chitotriosidase and AMCase, which can hydrolyse chitin. We hypothesized that the mammalian chitinases play a role in the in vivo efficacy of caspofungin.MethodsIn order to determine the role of chitotriosidase and AMCase in IPA, both chitinases were measured in rats which did or did not receive caspofungin treatment. In order to understand the role of each chitinase in the breakdown of the caspofungin-exposed cells, we also exposed caspofungin treated fungi to recombinant enzymes in vitro.ResultsIPA in immunocompromised rats caused a dramatic increase in chitinase activity. This increase in chitinase activity was still noted when rats were treated with caspofungin. In vitro, it was demonstrated that the action of both chitinases were needed to lyse the fungal cell wall upon caspofungin exposure.ConclusionCaspofungin seemed to alter the cell wall in such a way that the two chitinases, when combined, could lyse the fungal cell wall and assisted in clearing the fungal pathogen. We also found that both chitinases combined had a direct effect on the fungus in vitro.

Interaction of Ulocladium atrum, a Potential Biological Control Agent, with Botrytis cinerea and Grapevine Plantlets

The effectiveness of biological control agent, Ulocladium atrum (isolates U13 and U16) in protecting Vitis vinifera L. cv. Chardonnay against gray mold disease caused by Botrytis cinerea, and simulation of the foliar defense responses was investigated. A degraded mycelium structure during cultural assay on potato dextrose agar revealed that U. atrum isolates U13 and U16 were both antagonistic to B. cinerea, mainly when isolates were inoculated two days before Botrytis. Under in vitro conditions, foliar application of U. atrum protected grapevine leaves against gray mold disease. An increase in chitinase activity was induced by the presence of U. atrum isolates indicating that the biological control agents triggered plant defense mechanisms. Moreover, U13 has the potential to colonize the grapevine plantlets and to improve their growth. The ability of U. atrum isolates to exhibit an antagonistic effect against B. cinerea in addition to their aptitude to induce plant resistance and to promote grapevine growth may explain a part of their biological activity. Hence, this study suggests that U. atrum provides a suitable biocontrol agent against gray mold in grapevines.

Determinants of fenhexamid effectiveness against grey mould on grapevine: Respective role of spray timing, fungicide resistance and plant defences

Botrytis bunch rot of grapes is mainly controlled by applying fungicides at three crop growth stages: the end of flowering (BBCH 68), bunch closure (BBCH 77) and the beginning of veraison (BBCH 81). The hydroxyanilide derivative fenhexamid is among the most effective fungicides registered to control Botrytis cinerea. Its effectiveness was examined in relation to spray timing, fungicide resistance and defense responses of grapevine. Overall, the earlier fenhexamid was applied, the more effective it was at controlling B. cinerea. Frequencies of B. cinerea strains which were resistant to fungicides were evaluated at harvest. The frequencies of resistant phenotypes were similar among treatments and years with the exception of a class of multidrug resistant strain (MDR 2) whose frequency appeared to increase after fenhexamid applications. If current spray programs including fenhexamid appear to control bunch rot at the current MDR frequency, a propagation of MDR 2 strains might lead to a decline in disease control. Finally, defense responses were studied in grapevine flowers/berries following fenhexamid application. None of the defense processes tested was induced in flowers/berries at stages 68 and 77. Only an increase in chitinase activity was observed in treated-berries at stage 81, suggesting that fenhexamid effectiveness was not related to a stimulation of defense responses.