Study On Screening And Identification Of Entomopathogenetic Nematodes And Their Virulence To White Grubs

More than 135 soil samples were collected from Cangzhou region of Hebei province and used for isolating Entomopathogenetic Nematodes (EPNs). Preliminary screening of isolated EPNs was taken and one strain (ZZ) was obtained which has strong virulence to white grubs. The mortality of 1st instar larvae of Holotrichia oblita, Holotrichia parallela and Anomala corpulenta which were infected by ZZ were 48.33%,100% and 71.67% respectively, and were significantly higher than the other EPNs .ZZ population was classified and identified based on the morphological characteristics and ITS-rDNA sequence analysis. Through morphological observation and respond measurements of infective juveniles and males, the population was found closely related to Heterorhabditis bacteriophora, at the same time, the results of ITS1-rDNA sequences alignment and phylogenetic analysis showed the population had the closest genetic relationship with H. bacteriophora. Therefore, the population was identified as H. bacteriophora Cangzhou strain.Virulence of different EPNs to H. parallela larvae indicated that the mortality of 1st and 2nd instar larvae infected by H. bacteriophora Cangzhou strain was higher than others infected by Steinernema feltiae and Steinernema longicaudum after infection of 72 h and 96 h respectively , which suggested that the pathogenicity of Cangzhou strain was significantly higher than the two other species.Virulence of H. bacteriophora Cangzhou strain to larvae of H. oblita, H. parallela and A. corpulenta were determinated. For 1st instar larvae,virulence to H. parallela was significantly higher than the others after infection of 72 h; For 2nd instar larvae,the mortality of H. parallela and A. corpulenta had no significant difference, but they were significantly higher than H. oblita after infection of 96 h; After infection of 12 d, the mortality of A. corpulenta 3rd instar larvae was 80% and higher than the others. Thus, the nematode has strong virulence against H. parallela and A. corpulenta. Virulence of H. bacteriophora Cangzhou strain to H. parallela 1st instar larvae was determined at different infective dosage, temperature and soil moisture in lab. It was confirmed that the mortality of larvae increased with dosage and with time dependent. The mortality at the doses of 400 Ijs/mL, 250 Ijs/mL and 200 Ijs/mL all reached to 100% after infection of 72 h, 96 h and 120 h respectively. The temperature treated at 25℃and 30℃were most in favour of infection, all of larvae died after infected 48 h and 96 h. The mortality of treated larvae under soil moisture of 14% and 17% were 100% after infected 96 h and 120 h. There was no significant difference between soil moisture of 14%, 17%, 11%, 20% and 23%, but when the soil moisture below 8% was detrimental to infection.The results of energy substances variation in A. corpulenta hemolymph infected by H. bacteriophora Cangzhou strain showed that the protein content increased greatly and was significantly higher than the control after treatment of 20 h , it had a dramatic decrease subsequently and was significantly lower than the control after treatment of 28 h. The trehalose content decreased greatly than prophase after infection of 16 h; With the infection time, the trehalose content had reached the highest and lowest point after treatment of 20 h and 28 h respectively, and the latter had a significant difference against the control .