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Antagonistic Activities of Trichoderma Strains Against Chilli Anthracnose Pathogen

Mohammad Ahsanur Rahman
Plant Biotechnology and Microbiology Laboratory, Department of Botany, University of Rajshahi, Rajshahi 6205, Bangladesh.

Mohammad Anowar Razvy
Department Agronomy and Agricultural Extension, University of Rajshahi, Rajshahi 6205, Bangladesh.

Mohammad Firoz Alam
Plant Biotechnology and Microbiology Laboratory, Department of Botany, University of Rajshahi, Rajshahi 6205, Bangladesh.

Abstract/ Executive Summary:

Antagonist activities of different Trichoderma strains were evaluated in vitro against Colletotrichum capsici, a causal agent of anthracnose fruit rot of chili. Dual culture test showed that Trichoderma strains effectively inhibited mycelia growth of the pathogen. T. harzianum IMI-392433 showed the highest inhibition (81.96 %) and mycelial overgrowth (78.98%). Also, metabolites having 80% concentration extracted from 30-day-old T. harzianum IMI-392433 revealed the highest PIRG (percentage inhibition of radial growth) value of 85.16 and 87.18% by using normal poison and modified bilayer poison agar technique, respectively. Further, metabolites extracted from 30-day-old T. harzianum IMI-392433 at a concentration of 2000 mgL ^-1 completely inhibited spore germination and germ tube elongation of the pathogen. Also, severity of anthracnose was significantly decreased as compared with the control (2% methanol) when chili fruits were soaked in 2000 mgL ^-1 of 30-day-old metabolites from T. harzianum strains. Importantly, metabolites extracted from T. virens IMI-392430 (71.09) and T. pseudokoningii IMI-392431 (69.52) were found to be most efficient in the inhibition of disease severity. Taken together, our data suggest that Trichoderma strains especially T. harzianum IMI-392433 is a potential antagonistic organism and can be used to control the anthracnose disease caused by C. capsici.

Keywords: Antifungal metabolites, Trichoderma, Biological control, Anthracnose of chili, Colletotrichum capsici

Location: Biotechnology and Microbiology Laboratory, Department of Botany, Rajshahi University, Bangladesh.

Start Date:

End Date:

Program Area: Pest Management

Commodity: Chilli

Objectives:

(i) To observe the effect of five Trichoderma strains on mycelial growth inhibition and overgrow mycelia of Colletotrichum capsici.

(ii) To evaluate the efficacy of antifungal metabolites extracted from selected five Trichoderma strains on mecelial, spore germination and germ-tube growth inhibition of Colletotrichum capsici.

Methodology:

Five Trichoderma strains namely; T. virens IMI-392430, T. pseudokoningii IMI-392431 and T. harzianum IMI-392432, T. harzianum IMI-392433 and T. harzianum IMI-392434 were used in this study which was collected from Biotechnology and Microbiology Laboratory, Department of Botany, Rajshahi University, Bangladesh.These strains were isolated from decomposed garbage and soil and were verified by CABI Bioscience, Surrey, U.K. Isolation and pathogenicity test of the pathogen C. capsici, a causal agent of anthracnose was isolated from nine anthracnose infected chili fruits obtained, from the different chili planting areas in Pabna and Rajshahi districts, Bangladesh. Anthracnose fungus was isolated by tissue transplanting method described by Agrios (2005). For pathogenicity test, each of nine isolates of C. capsici was cultured on PDA for 3 days. Then 0.7 cm agar plug contained with mycelia of C. capsici was placed on pierced area on chili fruit (Capsicum annum L. var. annum) obtained from chili plantations area at Agriculture field, Khorkhori, Rajshahi district. All inoculated fruits were incubated in moist plastic chamber, kept at room temperature (27±°C). Disease severity of anthracnose infection was recorded at 5 days after incubation by measuring size of diseased lesion on chili fruit. The percentage of disease severity was calculated by using the formula; ((R_T-R_C)/R_T) ×100, when R_T was the mean of diseased lesion radius on chili in the tested treatment and RC was the mean of diseased lesion radius on chili in the control (placed with an agar plug without C. capsici). For each treatment, there were four replicates and 5 chili fruits were used in each replicate. Then a most pathogenic isolate was used for further.For mycelial growth inhibition and overgrowth test, dual culture test was performed. Trichoderma strains and C. capsici was sub cultured onto PDA for 4 days. The margin of colony of C. capsici was cut with sterile cork borer (0.7 cm diameter) and placed on agar surface at 1.5 cm from a margin of 9 cm diameter Petri dish. At 4 days after placing the plug of C. capsici, a plug of the Trichoderma strains was inoculated at the opposite direction, 6 cm apart from the C. capsici plug.The dishes were incubated for 5 days at room temperature, and then mycelial growth inhibition and the ability of Trichoderma strains to overgrow the colony of C. capsici were observed and compared with the control treatment (C. capsici grown on PDA without a Trichoderma). The inhibition levels were calculated by using the formula; ((GC-GT)/GC) ×100, when GC was the mean of colony radius of C. capsici in the control dish and GT was the mean of colony radius of C. capsici in Petri-dish of dual culture test. Each treatment was performed with four replicates, one dish per a replicate. The overgrowth rates of Trichoderma strains were calculated by using the formula; ((D ₁-D₂)/T_d) ×100, when D₁ was the mean of colony radius of the Trichoderma strains on the day of recording, D₂ was the mean of colony radius of the Trichoderma strains on the day before recording and Td was the time (d) between before and after recording. Each treatment was performed with four replicates.Two hundred milliliters of Richard’s solution (KNO₃: 1.0g, KH₂PO₄: 0.5g, MgSO₄·7H₂O: 0.25g, glucose: 34g, and trace amounts of FeCl₃ in 1 liter distilled water, pH 6.5) was prepared and poured into 500 ml conical flasks and autoclaved for 15 minute at 121 ºC/1.05kg/cm² pressure. Six pieces of agar discs (6mm) were kept in a flask (with media) for each strain of Trichoderma with four replicates. The flasks were incubated on a Gallenkamp orbital incubator at 100rpm at 28 ºC. The culture filtrates were collected after 10, 20, and 30 days of incubation. These were then concentrated to about 50% using a vacuum evaporator at 38~40 ºC and filtered by sterilized membrane filter.

Source of Information: International Journal of Microbiology and Mycology, Vol. 1, No. 1, p.7-22, 2013, p ISSN: 2309-4796

Article Link (Online Journal): http://www.innspub.net/wp-content/uploads/2013/05/IJMM-V1No1-p7-22.pdf

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

In vitro results obtained using different techniques suggest that T. harzianum IMI 392433 was the best for inhibition of the mycelial growth, conidial germination, germ tube elongation and disease severity of C. capsicci. This strain can be used as potential biological control agent to control the anthracnose disease of chilli.

Knowledge Management: Journal