In-Vitro Efficacy of Commercial Fungicides against Bipolaris Sorokiniana : Induced Spot Blotch Disease of Wheat

— Spot blotch caused by Bipolaris sorokiniana is an important disease of wheat. A laboratory experiment was conducted at Plant Pathology Division, Nepal Agriculture Research Council, Khumaltar, Nepal to evaluate the efficacy of commercially available fungicides viz, Saaf ( Carbendazim 12%+ Mancozeb63%), Sectin (Fenamidone 10%+ Mancozeb 50%), Angel (Metalaxyl 8%+ Mancozeb 64%), Diathane M-45 (Mancozeb 75%), G-MIL( Cymoxanil 8%+ Mancozeb 64%) tested in two different concentration i.e. 50ppm and 100 ppm except Curex (Copper oxychloride 50%) tested in the concentration of 100ppm and 200ppm and Tilt (propiconazole 25%) tested in 15ppm and 30 ppm concentration with a control test by employing poisoned food technique against spot blotch pathogen of wheat Bipolaris sorokiniana. The experiment was laid out in Completely Randomized Design (CRD). The result revealed that all concentrations of different fungicides successfully inhibited the radial mycelial growth of the pathogen under in vitro condition. Based on the measurement of fungal radial growth, fungicide Tilt of 30ppm and 15 ppm concentration were the most effective followed by Sectin 100ppm, Diathane M-45 100ppm, Angel 100 ppm and Saaf 100ppm. G-MIL 50 ppm in poisoned food technique was the least effective. Use of safer and economical chemical fungicides can provide an effective and long-term solution against plant diseases in agricultural farming.


INTRODUCTION
After rice and maize, wheat (Triticum aestivum) is Nepal's third most significant cereal crop. Wheat is produced from the Terai to the high alpine regions, and wheat consumption is on the rise. However, the yield per unit is much lower than anticipated. The disparity between the maximum observed yield and the national average yield, as well as the declining yield trend, necessitate immediate research. The low yield is due to a number of factors, including the scarcity of improved varieties, the occurrence of various wheat diseases, a lack of reliable irrigation, inclement weather, a lack of improved technology, and biotic and abiotic stress factors, all of which result in significant yield losses each year (Joshi et al., 2007). Fungal diseases in wheat cultivation are more important among biotic stress factors because they produce a significant drop in yield as well as deterioration in grain quality. In the humid subtropics of South Asia, where the irrigated rice-wheat rotation covers more than 12 million ha, there is growing evidence that stress conditions are increasing the severity of foliar diseases (Dubin et al., 1994) because rice serves as a host for the spot blotch fungi and rice stubble plays its role as a substrate for the fungi after rice harvest (Saari, 1998). Therefore, the fungal pathogen, Bipolaris sorokiniana (Sacc.) Shoemaker (teleomorph Cochliobolus sativus) induced spot blotch disease of wheat has emerged as one of the prime diseases for profitable wheat production in different zones of Nepal. Spot Blotch is caused by Bipolaris sorokiniana in Mega Environment 5A (ME5A), characterized by high temperature (coolest month greater than 17ºC) and high relative humidity (RH) (Dubin et al., 1991;Rajaram et al., 1993). It is a seed borne fungal. Spot blotch (oval to round brown blotch encircled by yellow halo) is the pathogen's principal symptom. The pathogen's continuing growth and  (Duveiller et al., 2005).
Bipolaris sorokiniana was also identified as a main contributing factor in a study undertaken in Nepal in 1996 to investigate and identity the national issue of "low germination in wheat' (Shrestha et al., 1997). The losses due to spot blotch in warm regions of Nepal ranged from 23 -40% depending on the genotypes and other environmental factors (Tripathi et al., 2005;Sharma et al., 2006).
To lessen the losses caused by the disease, a range of syste mic fungicides with various modes of action and targets ha ve been developed (Pasquer et al., 2005). However, in vitro evaluation against Bipolaris sorokiniana are barely sufficient and even accurate information on determining efficacy, sensitivity of different fungicides with minimum inhibitory concentrations have yet to be defined against Bipolaris sorokiniana (Iqbal, 2010). It is important to examine the effect of different concentration of fungicides commonly available in market to control this disease. Stock solution of each fungicide was prepared in distilled water and incorporated into Potato dextrose agar medium and mixed thoroughly before autoclaving. After autoclaving the medium was poured aseptically in sterilized petri-plates of 9 cm size under laminar air flow and allowed to cool. Five mm mycelial circular discs of pathogen excised with sterile cork borer from a seven days old culture of test pathogen was placed at the centre of each petri-plate and incubated at 25ºC. PDA with water or without chemical served as control. The experiment was arranged in complete randomized design (CRD) and there were four replications for each (15) treatments. The plates were incubated at 25 ºC inside BOD incubator. Measurement of the colony diameter of pathogens was taken after 48 hours for 10 days ( i.e 2 nd , 4 th , 6 th , 8 th day and 10 th day) after inoculation with the help of vernier caliper. Percent growth inhibition of the pathogen was calculated by using the following formula of Vincent (1947).

II. MATERIALS AND METHOD
Where,

I= inhibition percentage C= Colony diameter in control and T=Colony diameter in treatment
The data collection started from 48 hours after poisoned food technique i.e. 8th to 18 th March 2021. The data were recorded by measuring the growth of the test pathogen after each 24 hours by using Vernier caliper in mm. The data obtained from the experiment were analyzed using the software Gen Stat for the analysis of variance (ANOVA) to test the significance of treatments effect on mycelial growth of Bipolaris sorokiniana. Means of significant treatments at 5% level of significance were compared following Duncan's Multiple Range Test (DMRT) and Microsoft Excels. Means with the different letters are significantly different at 5% level of significance using LSD. In (Table 2), different letters in a column signifies that the treatment means are significantly different with each other at P-value <0.001.  Irrespective of the low concentration, at the end of tenth day maximum inhibition of mycelial growth for the control of Bipolaris sorokinina was recorded in Tilt 30ppm (100%) and Tilt 15ppm (100%) which was significantly superior than other treatments followed by Mancozeb alone highly inhibited the growth of Bipolaris sorokiniana. However, it showed variations when used as combination fungicides. Mancozeb with Fenamidone inhibited the mycelial growth by 88.13 % at 100 ppm. Mancozeb with carbendazim at 100ppm also showed better results i.e. 77.48% mycelial inhibition. Likewise Mancozeb with metalalxyl at 100 ppm showed 79.72% mycelial growth inhibition. But Mancozeb with Cymoxanil at 100 ppm (64.4%) and 50 ppm (59.6%) was lower as compared to other combinations.
It was reported that copper oxychloride showed 75.1% mycelial growth inhibition at 200 ppm and 65.7% at 100ppm. The mode-of-action of copper fungicides is the nonspecific denaturation of cellular proteins. It disrupts the function of proteins and enzymes after absorption and results in cell damage and membrane leakage (Husak, 2015). Samia et al. (2015) also reported 70-80% mycelial growth inhibition in isolates of Bipolaris sorokiniana, collected from different region of Bangladesh, at 300 ppm concentration of copper oxychloride.