Morpho-Molecular Characterization of Trichoderma Isolates from Rhizospheric Soils of Vegetables in Pakistan

Trichoderma, a major fungal genus attaining importance due to its diverse application in biological control programs and is considered a substitute for chemical pesticides. This research was conducted to characterize various Trichoderma species isolated from rhizospheric soil samples morphologically followed by its confirmation using molecular tools. A systematic survey of Trichoderma populations associated with soils of different vegetable hosts would enable a clear picture of the distribution of species in the region. Samples were collected from the rhizospheres of a variety of vegetable hosts and obtained numerous Trichoderma isolates (T. harzianum, T. viride, T. hamatum, T. longibrachiatum, T. asperellum, T. koningii and T. longipile). Morphological characteristics revealed that T. harzianum resembles T. viride but is more pigmented with confined rings than T. viride and other associated species. T. viride sporulation was more rapid than other species, producing a soft mat on PDA media. T. viride produces a sweet smell of coconut; T. asperellum produces a misty odour while T. longibrachiatum produces a yellow pigmentation in the media. Fifty out of 200 morphologically identified species were genetically characterized using universal primers (ITS-1 and ITS-4). ITS-based sequencing resulted in a product of 650 bp in all the isolates. The sequencing of these isolates showed five different species. As per rDNA, the species identified are: T. harzianum, T. hamatum, T. longibrachiatum, T. asperellum and T. viride with 98-100% sequence similarities to other related Trichoderma isolates reported from China, India, Mexico, USA, Portugal, Germany, Spain and Brazil. Bioinformatics analysis was conducted using maximum parsimony (MP) that supports the resemblance of the present study Trichoderma species with species reported from other countries. It is concluded that Trichoderma strains with biocontrol activity are genetically different compared to the pathogenic ones. The findings of this study help in providing an opportunity to test these isolates against different plant pathogens and ultimately leads to the development of bio-pesticides that could be eco-friendly and cost-effective with no chance of resistance development