MSc

Metarhizium anisopliae Sorokin, is a widely used entomopathogenic fungus for the biocontrol of insect pests. However, vulnerability to various abiotic stresses limits its performance as a biological control agent. Identification and evaluation of isolates tolerant to abiotic stresses can help overcome this limitation. To this end, soil samples were collected through purposive sampling from areas in central Kerala with a history of abiotic stresses. Metarhizium anisopliae isolates were obtained from soil samples collected from Moncompu, Alappuzha among the areas surveyed. Three isolates viz., MC 2, MC 4 and MC 7 were evaluated for tolerance to acidity, salinity, high temperature, drought, insecticides and fungicides.
Highest acidity tolerance was recorded by the isolate MC 7 with the highest mycelial biomass of 1.15 g at the lowest pH of 2.5. In the salinity tolerance trials, isolate MC 2 exhibited highest growth and sporulation at 0.5 M NaCl with only 6.33 per cent growth inhibition while the other two isolates recorded more than 50 per cent growth inhibition. At the higher salt concentration of 1.5 M, more than 89 per cent growth inhibition was recorded in case of isolates MC 4 and MC 7, while MC 2 recorded the least inhibition of 58.84 per cent indicating high salinity tolerance.
Screening for temperature tolerance at temperatures ranging from 25 to 40°C revealed that the isolate MC 7 recorded highest mycelial biomass (1.28 g) at 37°C. Similarly, studies on drought tolerance showed that the isolate MC 2 survived at highest concentration of 35 per cent Polyethylene glycol (PEG) with mycelial weight of 0.93 g suggesting drought tolerance as well. The isolate MC 2 recorded multiple stress tolerance and was found to be suited to both saline and drought prone areas. Similarly, the isolate MC 7 was found to be highly suitable to acidic soils as well as high temperature areas.
Compatibility of M. anisopliae isolates with popular insecticides like spinosad, cypermethrin, imidacloprid and chlorantraniliprole as well as fungicides like copper oxychloride, carbendazim and hexaconazole was also assessed. All the isolates showed less than 22 per cent growth inhibition in PDA medium amended with the highest doses of pesticides except
cypermethrin, carbendazim and hexaconazole. Based on the radial growth and sporulation, isolates MC 7 and MC 2 were found to be relatively more compatible to both insecticides and fungicides.
Biochemical analysis of the stress tolerant isolates, MC 7 and MC 2 revealed that the defensive enzymes like peroxidase and catalase were elevated both at highest temperature (37°C) and highest concentration of PEG (35 %). Both the isolates exhibited peroxidase activity of 0.058 and 0.030 EU/min/g tissue and catalase activity of 0.140 and 0.294 EU/min/mg protein respectively when compared to control. Quantity of trehalose was also maximum at highest temperature and at highest concentration of PEG with a value of 1.380 (MC 7) and 1.580 mg/ml/g mycelia (MC 2) respectively. Esterase, an enzyme essential for fungal infection process was also estimated and its activity was low in the isolate MC 7 at 37°C as well as in the isolate MC 2 at 35 per cent PEG when compared to respective control treatments.
Biocontrol efficacy of M. anisopliae isolates was evaluated against both adults and grubs of the storage pest, Tribolium castaneum at different doses of 1 × 105 to 1 × 109 spores/ml under laboratory conditions. The lowest LT50 values of 9.43 days and 4.14 days were recorded by the isolate MC 2 at a concentration of 1 × 109 spores/ml on adult and grub respectively. However, LT50 values were on par with that of positive check, NBAIR strain as well as MC 7.
The study was successful in identifying isolates of M. anisopliae with potential for biocontrol in agroecosystems vulnerable to abiotic stresses. It also helped to highlight the importance of screening potential stress tolerant isolates of microbial bioagents.