| As a new biological control factor,entomopathogenic nematode has been increasingly used in the pest control.However,as a living pathogenic microorganism,entomopathogenic nematode is greatly affected by natural conditions.Therefore,it is of great significance to screen excellent resistant nematode strains for local pest control and to carry out the research on the functional analysis of the resistance genes for better promotion and application of this special control factor.In this experiment,the above two aspects are systematically studied and the results are as follows:1.The purpose of this experiment was to screen out the nematode strains with better resistance against high temperature,low temperature and dryness.The results showed that the mortality of Steinernema ceratophorum HQA-87 and S.carpocapsae ALL was the lowest after 6 hours of treatment at 40? which was 4.19%and 1.89%,respectively.So,the result showed that the two nematode strains had the best resistance to high temperature.The mortality of S.carpocapsae ALL and S.sp ZLS-3 were the lowest of 1.93%and 0.3%,respectively.So they have the better resistance to low temperature.In the experiment of desiccation resistance,S.carpocapsae ALL,were the better resistant strains,with the mortality of 1.34%.S.carpocapsae ALL,S.ceratophorum HQA-87,H.beicherriana LJ-24,S.tielingense LFS-65,H.bacteriophora HQ-4 and S.hebeiense JY-20 were screened out as the better resistant nematode strains.The pathogenicity of the six selected nematode strains to Galleria mellonella was determined again after treating with the adversity conditions.Finally,S.carpocapsae ALL and S.ceratophorum HQA-87 were determined to be the best resistant nematode strains.2.The pathogenicity of 26 kinds of nematodes were tested against the neonates,1st instars and 2nd instars larva of three kinds of Scarabaeoidea(Holotrichia diomphalia,Holotrichia parallela and Anomala corpulenta).The results showed that H.bacteriophora HQ-4,H.bacteriophora KF-63,H.beicherriana LJ-24 and H.bacteriophora NT-1 had the best pathogenicity on the neonates of H.diomphalia Bates.The mortality of neonates was 100%,100%,94.44%and 92.86%,respectively.H.indica HNN-2,H.beicherriana LJ-24 and H.bacteriophora KF-63 had the best lethal effect on the 1st instars larva with the mortality of 100%;and H.bacteriophora HQ-4 and H.sp NCWZ-1 had the best lethal effect on the 2nd instars larva with the mortality of 60%and 58.33%,respectively.H.beicherriana HDT-54,H.bacteriophora NT-1,H.sp NCWZ-1,H.beicherriana LJ-24 and H.bacteriophora HQ-4 had the better pathogenicity of on the neonates of H.parallela with the mortality of 100%;H.beicherriana LJ-24,H.bacteriophora HQ-4,H.beicherriana HDT-54,H.bacteriophora NT-1 and H.sp NCWZ-1 had the best lethal effect on the 1st instars larva with the mortality of 100%.H.beicherriana LJ-24,H.bacteriophora NT-1,H.beicherriana HDT-54 and H.bacteriophora HQ-4 had the best lethal effect on the 2nd instars larva with the mortality of 100%.H.indica HNPY-2,H.bacteriophora HQ-4 and H.sp NCWZ-1 had the best pathogenicity on the neonates of A.corpulenta with the mortality of 100%,81.82%and 72.73%,respectively;H.bacteriophora NT-1 and H.bacteriophora HQ-4 had the best pathogenicity on the 1st instars larva with the mortality of 100%and 78.57%,respectively;H.beicherriana LJ-24 and H.bacteriophora NT-1 had the best lethal effect on the 2nd instars larva with the mortality of 97.5%and 100%,respectively.We can find that the control effect of Heterorhabditis is better than that of Steinernema on Scarabaeoidea.H.bacteriophora HQ-4,H.sp NCWZ-1,H.beicherriana LJ-24 and H.bacteriophora NT-1 can be used as the excellent nematode strains to control Scarabaeoidea.3.As the research object,the expression differences of 16 resistance genes from S.ceratophorum HQA-87,including HSP20,HSP90,CHI,SC450,SC3602,SC 1309,CL1317,SC4783,SC1476,SC1825,CL5542,CL497,CL3276,SC5348,CL6144 and SC7883,were determined by fluorescence quantitative PCR.The results showed that the relative genes expression of HSP20,HSP90,CHI,SC450,CL497,SC4783,SC1476,CL5542 and SC3602 were up-regulated at high temperature condition;the relative genes expression of SC7883 and SC 1309 were up-regulated at low temperature condition;the relative genes expression of SC3276,CL1317,SC1825,SC5348 and CL6144 were up-regulated at dry condition.4.In order to study the resistance mechanism of entomopathogenic nematodes in vivo,the functional verification of HSP20 and CL6144 related to high temperature and dry condition were confirmed by RNAi technology.When HSP20 gene was silenced,the results showed that the nematodes mortality were 60.21%,62.20%and 67.29%under high temperature,dry and low temperature conditions,respectively.The difference between the nematodes mortality in the high temperature treatment and the control group was significant.It can be concluded that HSP20 gene is related to the resistance to high temperature.After CL6144 gene was silenced,the nematodes mortality was 19.5%,7.9%and 87.3%under high temperature,dry and low temperature conditions,respectively.The results showed that CL6144 gene might be related to the resistance of nematode to high temperature.The results also showed that there was no difference between the pathogenicity of nematodes treated with the genes silence and the control group,indicating that the resistance gene was not related to the pathogenicity of nematode. |
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