Pakistan Journal of Biological Sciences1028-88801812-5735Asian Network for Scientific Information10.3923/pjbs.2018.95.100WidiyaningrumPriyantini FauziyahLaily Rini IndriyantiDyah 22018212Background and Objective: Biological activity such as reproduction rate, viability and pathogenicity of local entomopathogenic nematodes (EPNs) are influenced by ecological factors, one of them is temperature. In order to prepare the biopesticides product, the study aimed to compare their production rate, viability and pathogenicity of EPNs from Semarang isolate and commercial biopesticide under various temperatures. Materials and Methods: This study was designed by Completely Randomized Design one-way classification. Both EPNs were cultured in vivo condition with Tenebrio molitor larvae by white trap method. The reproduction rate of nematodes was calculated after 8 days of incubation and its viability test at six storage temperatures. The EPNs with the best viability was further tested for its pathogenicity on Macrotermes sp. The reproduction rate of nematodes were analyzed by using students t-test, while one-way ANOVA was used to analyze the viability of EPNs, and pathogenicity of lethal dose value was calculated using Probit analysis. Results: The findings showed that the reproduction rate on EPNs from Semarang isolate was significantly higher (p<0.05) than EPNs from commercial biopesticide. Various temperatures significantly affected the viability on both nematodes, but under LSD test (p<0.05) revealed that viability at storage of 21, 24 and 27°C were not different of each other. Conclusion: The reproduction rate of EPNs from Semarang isolate was 35% higher than those originated from commercial biopesticide. The optimum viability of both EPNs was obtained at temperature ranged from 21-27°C. 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