Browsing by Author "Mabena, Nokuthula Busisiwe"

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    Characterization of EPNs and their bacterial symbionts
    (2024) Mabena, Nokuthula Busisiwe
    Many synthetic pesticides have been associated with health and environmental issues. Not only are they toxic to non-target organisms but can also pollute the soil, water and vegetation. Hence organic farming has been suggested as an alternative approach of biological pest management. Entomopathogenic nematodes (EPNs), which are obligate parasites to insects and carry pathogenic bacterial symbionts and are effective in managing and controlling populations of a variety of economically important insect pests. These insect pests pose a threat to food security as crop yield losses are primarily caused by them. Identifying natural enemies to insects is crucial as they have no harmful consequences to non-target organism and the environment and can be exploited as biological control agents. The first focus of this research was to isolate, identify and characterize a new nematode species. The nematodes were isolated from soil samples originally collected in Mahobong Leribe, Lesotho. To isolate the nematodes, the soil samples were baited with Tenebrio molitor larvae. The methods used for the identification and phylogenetic analysis of the isolated nematodes involved genomic DNA extraction, PCR amplification and Sanger sequencing of the 18S rDNA gene. The nematode that was isolated was a new, previously uncharacterized Cruznema species. The phylogenetic analysis and comparative morphometric characterization confirmed it was a new nematode. The results showed that the average body length of the female and male were 1006.45 µm and 868.31 µm, with a standard deviation of 210.95 µm and 162.75 µm. The species was also closely related to Oscheius spp., which have been reported to be entomopathogenic. Furthermore, the isolated nematode was able to cause mortality to T. molitor larvae within 3 to 6 days. Another focus of this research was to isolate and identify the bacterial symbionts of the isolated nematode. The methods that were used for the isolation involved the homogenization of sterile nematodes. The methods used for the identification and phylogenetic analysis involved genomic DNA extraction, PCR and Sanger sequencing of the 16S rDNA gene as well as culturing on selective and differential media. The isolated bacteria were identified as Alcaligenes spp., Elizabethkingia spp. and Enterobacter spp. They were also tested using lipid agar media to confirm if they were associated with the isolated nematode species. The results showed they were able to feed on and reproduce on the media. 4 The overall results of this study show that the newly identified Cruznema species has the potential of being used as an EPNs