COMPARATIVES STUDY OF BLOTTER, AGAR-PLATE, AND DEEP-FREEZING METHODS FOR THE DETECTION OF SEED-BORNE FUNGAL PATHOGENS IN WHEAT (TRITICUM AESTIVUM L.) CULTIVARS OF THE DISTRICT MANSEHRA REGION, KHYBER PAKHTUNKHWA, PAKISTAN
DOI:
https://doi.org/10.64013/bbasrjlifess.v2026i1.64Keywords:
Seed-borne fungi, Triticum aestivum, blotter test, agar-plate method, deep-freezing test, Alternaria alternata, Tilletia indica, seed health testing, MansehraAbstract
Seed-borne fungal pathogens are major issues limiting wheat (Triticum aestivum L.) productivity, affecting germination potential, vigour of seedlings, grain quality, and also serving as a primary source of fungal inoculum for field epidemics. This study was conducted to compare the efficacy of three most popular methods, such as the blotter test, deep-freezing blotter method, and agar-plate method for detection of seed-borne fungal pathogens in Wheat (Triticum aestivum L.) seeds. Fifty wheat seed samples from 14 different areas in District Mansehra of Khyber Pakhtunkhwa, Pakistan, were collected and belonged to 9 commercial cultivars. A total of 9 species of fungi were isolated from 50 wheat seed samples of nine wheat varieties, including Alternaria alternata, Ustilago tritici, Fusarium oxysporum, Penicillium chrysogenum, Curvularia lunatus, Aspergillus niger, Tilletia indica, Mucor mucedo, and Rhizopus stolonifera. All nine species were isolated by the agar-plate method compared to the other two methods, which also had the highest overall isolation frequency (96%), blotter test (84%), and the deep-freezing method (80%), respectively; three species (A. niger, P. chrysogenum, and R. stolonifera) were only isolated by the agar-plate method. In all methods, the most common species was A. alternata, detected in 90% of the blotter test and 94% of the deep-freezing test. A supplementary wash test found T. indica in 90% of samples, and the deep-freezing method showed the highest recovery of this quarantine-significant pathogen in the comparative tests. In location-wise analysis, the highest fungal growth was found in Dhodial, while the cultivar-wise analysis found that the cultivar with the highest fungal load was Pirsabak-2013 and the cultivar with the highest germination percentage was Paseena-2017, while Pirsabak-2013 had the lowest. A. alternata, Curvularia lunatus, Tilletia indica, Mucor mucedo and Fusarium oxysporum were symptomatically transmitted from infected seed to emerging seedlings of cultivars Zincol and Paseena-2017 in a seed-to-seedling transmission assay. The results indicated that the agar-plate method is the most reliable single test to use for routine phytosanitary screening in this agro-ecological zone, and that this test, when used in combination with the blotter, freezing and wash test, can be most effective in detection of saprophytic and quarantine pathogens.
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