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MORPHOLOGICAL CHARACTERIZATION OF SEED-BORNE FUNGI ASSOCIATED WITH BRRI RICE VARIETIES IN BANGLADESH

Twenty five fungal species, representing 13 genera were found to be associated with seeds of 20 varieties of rice (BRRI dhan 56 to BRRI dhan 75). Fungi were isolated following "Tissue planting" method and "Blotter method". The isolated fungi were Alternaria alternata, A. tenuissima, Aspergillus flavus, A. fumigatus, A. niger, A. ochraceus, A. terreus, Bipolaris multiformis, B. oryzae, B. sorokiniana, Chaetomium globosum, Curvularia lunata, Fusarium equiseti, F. fugikuroi, F. oxysporum, F. proliferatum, Microdochium fisheri, Nigrospora oryzae, Penicillium sp., Pestalotiopsis oxyanthi, Phanerochaete chrysosporium, Rhizopus stolonifer, Sarocladium oryzae, Syncephalastrum racemosum and Trichoderma viride.


Introduction
Rice (Oryza sativa L.) is the second largest crop grown in the world in terms of both area and production. It is the staple food for more than half of world's population. The majority of the rice produced comes from China, India, Indonesia, Pakistan, Bangladesh, Vietnam, Thailand, Myanmar, Philippines and Japan (1) .
Rice suffers from more than 60 different diseases of which 43 are known to occur in Bangladesh. Among these diseases, 27 are seed-borne of which 14 are of major importance (2) . Different seed-borne diseases of rice are bakanae, stackburn, brown spots, black kernel, blast, sheath blight, sheath rot, stem rot and leaf scald and they cause yield reduction, quality deterioration and germination failure (3)(4)(5) . In Bangladesh, approximately 2.5 million tons of rice worth more than Tk.12000 million is lost annually due to diseases caused by seed-borne pathogens (6).
The information about morphological features of seed-borne fungi with newly released BRRI rice varieties is inadequate in Bangladesh. Therefore, the present investigation has been undertaken to investigate morphological characteristics of seed-borne fungi associated with BRRI dhan 56 to BRRI dhan 75.

Materials and Methods
Seeds of 20 BRRI released rice varieties (BRRI dhan56 to BRRI dhan75) were collected from Genetic Resources and Seed Division of Bangladesh Rice Research Institute (BRRI) during January, 2016 to July, 2017. The samples were kept in brown paper bag, labeled properly and stored immediately in a dry safe place in the laboratory until used for the experiments and preserved at 4ºC in refrigerator for further studies. The experiment was conducted in Laboratory of Mycology and Plant Pathology, Department of Botany, University of Dhaka, Bangladesh.
The fungi were isolated from the seed samples following Tissue Planting method on PDA medium (12) and Blotter method (13) . For Tissue Planting method, 400 seeds were washed in sterile water and then surface sterilized by dipping in 10% chlorox for 5 minutes. Then they were transferred into a sterile Petri plate containing sterile blotting paper to remove the excess surface water. The seed inocula were placed in Petri plates containing sterilized PDA medium. Each Petri plate contained 15 ml of PDA medium with an additional of 1 drop (ca. 0.03 ml) of lactic acid.
Then the inoculated plates were incubated at room temperature (25 ± 2°C) for 5 -7 days. For Blotter method, 400 surface sterilized seeds of each sample were placed on two layers of moist blotting paper (Whatman No.1) in sterilized Petri plates and incubated at 25 ± 2°C for 5 -7 days. A total number of 100 inocula were transferred in 10 Petri plates. After 7 days of incubation the fungi were observed and recorded.
Morphological studies of the fungal isolates were done in order to determine their identification. The microscopic structural characters of the isolated fungi were recorded under Nikon D 5000 digital camera. Identification of the isolates were determined based on morphological characteristics observed under a compound microscope following the standard literature (14)(15)(16)(17)(18)(19) . All the specimens, were preserved in Mycology and Plant Pathology Laboratory, Department of Botany, University of Dhaka, Bangladesh.