In-vitro evaluation of different fungicides and bioagents against Fusarium oxysporum F. sp. capsici

Introduction

Chilli (Capsicum annuum) is the major economic crop commercially cultivated in India. It is grown in all the regions of country. Production of chilli comes in 400 dissimilar species from all over the world. Chilli crop comes under solanaceae family which is a significant vegetable category. The origin is said to be in tropical America specifically from Brazil (Thompson and Kelly, 1957) [13] . Chilli is important spice crop of Indian diet consumed as fresh or processed. Chilli is a tropical and subtropical plant that requires a mix of warm, humid, and dry temperatures to grow. Warm, humid conditions is required during the growth period. India is the largest producer and exporter of chilli, which is grown over an area of 721.15 ha, with production of 1689.52 T and productivity of 2343 kg/ha. (Anonymous, 2019) [1] . The major states growing chilli are Andhra Pradesh, Orisha, Maharashtra, West Bengal, Karnataka, Rajasthan and Tamil Nadu. Andhra Pradesh stands first in chilli production, with maximum acreage under chilli cultivation in the country (Anonymous, 2019) [1] . During the Kharif and Rabi seasons Fusarium wilt disease was observed to inflict substantial loss in several Chilli fields. The damage due to Fusarium wilt disease are particularly serious in tropical region with symptoms including rolling of leaves, stunting of plants, twig death and withering of upper leaves followed by injury to the vascular system of plant particularly at lower stem and root. The sick plant were found in patches and overall crop production was lowered. The yield loss due to the disease reported were 10-80 percent, worldwide (Loganathan et al., 2013) [4] , depending on the prevailing climatic conditions. The pathogen attack on immature root as they grow and stem vessels blocks the water and nutrient transmission as a result the crop plant vascular system is destroyed resulting in plant death. (Suryanto et al. 2010) [12] . With these aspects in mind as well as economic importance of the Fusarium wilt disease of Chilli present investigation were carried out.

Materials and Methods

The present investigation on "In-vitro Evaluation of different fungicides and bioagents against Fusarium oxysporum F. sp. capsici "was studied during 2023-2024 at Department of Plant Pathology, College of Agriculture, Nagpur. The details of the material and methods used are as follows.

Isolation

Based on visual observation, the plants showing typical symptoms (discolored vascular system) were collected and carried in the laboratory for isolation. Infected roots samples were cut into pieces with sterilized blade and disinfected with sodium hypochlorite solution for two minutes. Pieces were then washed with three changes of distilled water and bits were air dried and placed on sterilized solidified PDA medium. The four bits were placed in each plate. The plates were incubated at 28±2 °C temperature. The plates were examined regularly. When fungal growth starts to appear, the culture was purified by single hyphal tip technique. Identification of fungus was done with the help of compound microscope based on morphological features.

In -vitro evaluation of fungicides

Poisoned food technique was used to evaluate the efficiency of fungicides against pathogens. Potato dextrose agar medium was prepared and distributed at the rate of 100 ml in 250 ml conical flask, autoclaved 1.05 kg/cm 2 for 15 min. Then before solidification of media different fungicides with desired concentration were incorporated aseptically in different flasks. These flasks shaken thoroughly and poured in Petri plates 20 ml/plate likewise three plates for each treatment were poured. One set of three plates was poured without any fungicides to serve as a control. After solidification of medium, the plates inoculated with eight days old pathogens separately. five mm diameter mycelial disc selected from peripheral growth of the plate by sterilized cork borer were used for inoculating the plates by keeping one disc per plate in the centre in inverted position, so as to make the mycelia growth touch the surface of medium. The inoculated plates were incubated at room temperature for seven days. The colony diameter of the fungal pathogens on medium was recorded and percent inhibition in each treatment was calculated (Vincent, 1947) [15] by using following formula.

C -T Percent inhibition (I) =

x 100 T Where, I = Percent inhibition of mycelial growth C = Growth of mycelium in control (mm) T = Growth of mycelium in treatment (mm)

In vitro evaluation of bio-agents by dual culture technique

Efficacy of four bioagents viz, Trichoderma viride, Trichoderma harzianum, Psuedomonas flurescens, Bacillus subtilis were evaluated by dual culture technique. 15 ml of autoclaved and sterilized PDA was poured in sterilized petri plates and were allowed to solidify. Seven days old cultures of the antagonist and the pathogen were used for the study. Five mm diameter discs of culture of the test fungus and bioagents were cut out with sterilized cork borer. Then the two growth culture discs, test fungus and bioagents were placed at equidistance and exactly opposite with each other on sterilized solidified PDA medium in petri plates. Petri dishes with no antagonist served as control for the pathogen. These plates were incubated at 27±2 °C for seven days. For each treatment four replications were maintained. The extent antagonistic activity by bioagents were recorded after incubation period of 7 days by measuring the growth of the test pathogen in dual culture and in control plates. In case of bacterial bioagents pathogen was kept in the middle of bacterial antagonist. Percent inhibition over control was worked out according to formula given by Vincent (1947) [15] .

C -T Percent inhibition (I) =

x 100 T Where, I = Percent inhibition of mycelial growth C = Growth of test fungus (mm) in control plate, T = Growth of test fungus (mm) in treatment plates

Results and Discussions

The present study was carried out on chilli wilt caused by Fusarium oxysporum f. sp. capsici. The study includes isolation of pathogen, identification of the isolated pathogen, proving Koch's postulates (Pathogenecity test). In-vitro evaluation of fungicides and bioagents against Fusarium oxysporum f. sp. capsici. The results of the experiments were presented here under.

In -vitro evaluation of fungicides

Efficacy of six fungicides at respective concentration was tested in-vitro by following poison food technique for mycelial growth of Fusarium oxysporum f. sp. capsici. The data presented in Table . 1 revealed that among the six fungicides, the most promising fungicides which restricted 100% growth of the test pathogen was Iprodione 50% WP, Tebuconazole 25.9% EC and Carbendazim12% + Mancozeb 63% WP, followed by Azoxystrobin 18.2% + Difenoconazole 11.4% SC and Metiram 55% + Pyraclostrobin 5% WG which recorded 77.12 and 66.72 percent inhibition respectively. Whereas the least control of the pathogen or minimum percent inhibition was observed in Copper oxychloride 50% WP having 35.70 percent inhibition.

In the present investigstion, the data revealed that Iprodione 50% WP, Tebuconazole 25.9% EC and Carbendazim 12% + Mancozeb 63% WP completely inhibited the mycelial growth at all the concentrations of 0.2%, 0.1% and 0.25% respectively. These fungicides were highly effective even at low concentration. In a similar way Pimpale et al., (2023) [7] reported that Carbendazim 12% + Mancozeb 63% WP @ 0.20% concentration were found most effective against Fusarium oxysporum F. sp. capsici. The results are also in conformity with the findings of several other workers in case of F. oxysporum i.e. Shah et al., (2006) [10] ; Madhavi and Bhattiprolu (2011) [5] ; Tewari et al., (2012) [14] ; Wani and Nazar (2012) [17] and Bashar and Chakma (2014) [3] and Rao et al., (2021) [9] . [8] reported T. harzianum as more effective than P. fluorescens against F. solani and F. oxysporum. Similarly Trichoderma spp. were reported as efficient antagonist against several Fusarium spp. Wani et al., (2014) [17] ; Nagendran et al. (2016) [6] .

References

1. Barari H. Biocontrol of tomato Fusarium wilt by Trichoderma species under in vitro and in vivo conditions. Cercetari Agronomice in Moldova. 2016;1(165):91-98.
2. Bashar MA, Chakma M. In vitro control of Fusarium solani and Fusarium oxysporum, the causative agents of brinjal wilt. Dhaka Univ J Biol Sci. 2014;23(1):53-60.
3. Madhavi GB, Bhattiprolu SL. Evaluation of fungicides, soil amendment practices and bioagents against Fusarium solani causal agent of wilt disease in chilli. J Hort Sci. 2011;6(2):141-144.
4. Nagendran K, Loganath M, Saha S, Mishra A, Pandey KK, Rai RM, et al. Biological Management of wilt disease on chilli caused by Fusarium solani. Veg Sci. 2016;43(2):253-256.
5. Pimpale KB, Suryawanshi JS, Deokar CD, Bansode KA. In vitro evaluation of different fungicides against Fusarium oxysporum f. sp. capsici. The Pharma Innovation Journal. 2023;12(2):2935-2938.
6. Rini CR, Sulochana KK. Usefulness of Trichoderma and Pseudomonas against Rhizoctonia solani and Fusarium oxysporum infecting tomato. J Trop Agric. 2007;45:21-8.
7. Rao V, Dhutraj DN, Navgire KD, Ambadkar CV, Apet KT. In vitro efficacy of fungicides against Fusarium oxysporum f.sp. melongenae causing eggplant wilt. Chem Sci Rev Lett. 2020;9(36):1074-81.
8. Shah MI, Sultan P, Nasier A, Williams P, Jan A, Sajad M, et al. In-vitro study on effect of some fungicides viz., Carbendazim, Mancozeb, Conjoint Carbendazim Mancozeb and Sulphur against F. oxysporum. Res J Microbiol. 2006;1(4):360-5.
9. Singh F, Hooda I, Sindhan GS. Biological control of tomato wilt caused by Fusarium oxysporum f. sp. lycopersici. J Mycol Plant Pathol. 2004;34:568-70.
10. Suryanto D, Patonah S, Munir E. Control of Fusarium wilt of chilli with chitinolytic bacteria. Hayati J Biosci. 2010;5-8.
11. Tewari AK, Shanaz E, Bhat ZA, Hassan MG. Integrated management of wilt complex disease in bell pepper (Capsicum annuum L.). J Agric Sci. 2012;4(7):141.
12. Vincent JM. Distortion of fungal hyphae in the presence of certain inhibitors. Nature. 1947;159(4051):850.
13. Wani SA, Mohiddin FA, Hamid B, Rizvi G, Bhat KA, Hamid A. Incidence of Fusarium wilt of chilli Capsicum annuum L. in Kashmir valley and its management by Trichoderma spp. Mycopath. 2014;12(1):1-8.
14. Wani FA, Najar AG. Integrated management of Fusarium wilt [Fusarium pallidoroseum (Cooke) Sacc.] in chilli Capsicum annuum L. SKUAST J Res. 2012;14(1-2):45.