Abstract
Foot rot caused by Sclerotium rolfsii Sacc. is one of the major diseases of vegetable all over the world. It significantly reduces the vegetable yield. Chemicals used to control the disease are less effective and sometimes impractical considering cost and hazardous impact of chemical to environment and human health. As alternative, attempt was taken to formulate antagonistic fungus Trichoderma which is ecofriendly and recognized as growth enhancer. Rhizosphere soil of different vegetable crops were collected from five different districts of Bangladesh, and a total of 49 isolates of Trichoderma were purified and identified as T. harzianum thereoff.. Isolates of T. harzianum distinctly differed on their cultural and morphological properties. Among the fast growing isolates the highest spore density (85.44x106-80.47x10°/plate) was obtained from the isolates of dark green and green colour. PCR-based Random Amplified Polymorphic DNA (RAPD) technique was used to assess genetic diversity and relationships among selected 35 Trichoderma isolates. Amplification with 3 decamer primers produced 36 scorable bands of which all (100%) were polymorphic. The co-efficient of gene differentiation (Gst) was 1.0000 reflecting the existence of high level of genetic diversity among the isolates. The Unweighted Pair Group Method of Arithmetic Means (UPGMA) dendrogram constructed from Nei's (1972) genetic distance produced 2 main cluster groups (13 isolates in cluster group 1 and 22 isolates in cluster group 2). These isolates may be used in genetic diversity and more developed analysis for the improvement of Trichoderma formulation. Ten isolates of Sclerotium rolfsii causal agent of foot rot of vegetables were obtained from four hosts and four locations of Bangladesh and their morphological characters were studied. The isolate obtained from Indian spinach had the highest linear mycelial growth rate (1.5 cm/day), highly virulent towards foot rot disease and it had significant effect on seed germination, pre-emergence death, damping off and plant stand as compared to other isolates. Selected 3 primers in RAPD analysis of eight isolates of S. rolfsii generated 20 scorable bands of which 9 bands (45%) were polymorphic and 11 bands (55%) were monomorphic. The co-efficient of gene differentiation (Gst) was 1.000 reflecting the existence of high level of genetic variations among the 8 isolates. The UPGMA dendogram produced two clusters (2 isolates in cluster 1 and 6 isolates in cluster 2). Antagonistic potentiality of 49 isolates of T. harzianum against the virulent isolate of S. rolfsii (isolated from spinach) was studied following the dual culture method. Among the isolates significantly the highest percentage of inhibition (99.00% to 99.07%) were observed in isolates 20, 37 and 49 and the maximum growth inhibition due to fungitoxic metabolite was found in isolate-49 (54.81%). Twenty two substrates were used for the formulation of Trichoderma, black gram bran combined with peat soil and water at 1:1:2 (W/W/V) ratio yielded the highest performance in starting sporulation and producing highest Colony Forming Unit (6.1 x108/g) followed by the grasspea bran combined with peat soil and water at 1:1:2 (W/W/V) ratio. Chickpea bran used with peat soil and water (1:1:2-W/W/V) had statistically identical performance with that of grasspea bran. Among the different doses of formulated Trichoderma the highest germination of eggplant (81.67%), tomato (87.33%) and Indian spinach (81.00%) were recorded in trays where formulated Trichoderma was used @ 20g/kg soil. Soil treatment with formulated Trichoderma was found to be most effective method in enhancing seed germination and reducing the disease incidence followed by seed + soil treatment method. Field study indicated more than seven days requirement for Trichoderma to be effective against S. rolfsii. For mass production of Trichoderma in the field poultry litter inoculated with formulated Trichoderma produced the maximum CFU/g (3.46x108) after one month of decomposition followed by cowdung CFU/g (2.23x10). Thus 7. harzianum CP (T49) can be formulated in blackgram-peat soil mixture for effective management of S. rolfsii. For commercial use T. harzianum can be multiplied in poultry litter or cowdung.