Microorganism Of Oviposition Slit And Their Pathogenicity On Anoplophora Glabripennis Larva

Anoplophora glabripennis,Asian longhorned beetle(ALB),is one of most important boring pest in China.According to natural population life table of ALB and other longhorned beetles, the mortalities of eggs and low instar larvae are high,and have significant influence on population dynamics.But the reasons remain unknown.This study reports the water content and pH of ALB oviposition slits in two climatic regions, Beijing and Ningxia,and microorganism community structure and diversity in oviposition slits. Two microorganism strains,Serratia marcescens and Beauveria bassiana,were isolated from oviposition slit directly or indirectly.Pathogenicity,invasion behavior and correlative enzyme activity of these two strains were studied.The main research conclusions were listed as follows:1.The water content of oviposition slits or imitated slits of poplar and willow in Beijing was similar to that in Ningxia.As for imitated slits,water contents of middle stage and late stage were high.The difference in water content between imitated slits and healthy bark was not apparent.Similarly,the water contents of middle stage and late stage of oviposition slits were high,which were higher than healthy barks.The trend of average water content from high to low was oviposition slits,healthy bark,imitated slits.2.pH of oviposition slits was the highest than that of other bark types.The average pH was 7.88～9.97.pH of healthy bark was neutral or little acidic(6.06～7.01),while pH of imitated slits(6.20～8.20) was higher than that of healthy bark.Therefore,the trend of average pH was oviposition slits＞imitated slits＞healthy bark.3.There were 1449 fungi strains isolated from oviposition slits,imitated slits and healthy bark from 5 tree species in Beijing and Ningxia.The isolated fungi were designated as 18 genera. Among them,2 genera were identified as Zygomycotina,16 genera as Deuteromycotina. Fusarium and Alternaria were dominated genera in three bark types of two areas.Fusarium was dominate genus in oviposition slits,whereas Alternaria was dominate genus in imitated slits.As for diversity index and species evenness,fungi population of oviposition slits and imitated slits in Beijing were higher than that in Ningxia.Fungi population of healthy bark in Beijing was lower than that in Ningxia.Xanthomonas Dowson and another unidentified strain were the main isolates in three types of bark.Serratia marcescens was found in three types of bark of Salix matsudana.Only one kind of actinomycete was isolated from three types of bark,and identified as Streptomyces spp.ALB larvae were found to dead when incubated with oviposition slit in aseptic condition. The pathogen was identified as Beauveria bassiana,suggesting that originated from oviposition slit.In addition,there were some insects living inside or outside of oviposition slits,such as potosia brevitarsis(Lewis),and insect adults of Nymphalidae,Formicidae,Apidae,attracted by large amount of sap flowing out of slits.Inside oviposition slits,saprophytic larvae of Nitidulidae, Sarcophagidae and Syrphidae were found.4.Laboratory virulence of B.bassiana and S.marcescens on low instar ALB larvae were tested.The results showed that the two pathogenic strains were pathogenic to ALB larvae.Trunk tests showed that B.bassiana had a good control effect(76.8%),but no control effect on S. marcescens.5.The infected larvae of ALB by B.bassiana showed that the part of epithelial tissue was disintegrated and subcutaneous fat was separated from epithelial tissue.Fat cell became atrophic and dead.Fat globule also became unclear.Muscle was disintegrated and some large vacuoles appeared with low coloration effect.In midgut,circular muscle was disintegrated,arranged in disorder and rumpled inward.Vacuoles appeared in blood corpuscle and blood corpuscle became distorted.6.Larvae infected by S.marcescens were found that part of cuticle was destroyed and degraded.Part of muscle was separated and filled with some vacuoles.Profile of midgut muscle became unclear.Circular and midgut circle muscle became a piece of lamina.One day later,all viscera became chaotic and tissues were indistinguishable.7.B.bassiana could invade circular directly by germ-tube.The strain produced few appresoriums.Germ-tube could invade circular with mean length 3.08μm.Silk in between hypha,silk between spores and hypha,and net among hypha were found in late infection.These phenomena have not been reported to date.8.B.bassiana could produce two main extracellular enzymes:extracellular protease and extracellular chitinase.The activity of extracellular protease induced in gelatin medium and larva powder medium was higher than that in yeast medium.High enzyme activity in gelatin medium appeared at 26～28℃,and in larva powder medium appeared at 35℃.Three mediums induced extracellular chitinase having higher enzyme activity at 26～30℃.Enzyme activity was low at 33～35℃.Colloidal chitin medium induced highest enzyme activity at 26℃,and highest enzyme activity at 30℃was larva powder medium.Extracellular chitinase of S.marcescens had a good induced effect in colloidal chitin medium at 30～37℃,and highest enzyme activity appeared at 33℃.Larva powder medium induced effect was lower than colloidal chitin.The difference in activity of enzymes induced by the two mediums was very significant.Chitin powder medium induced very low enzyme activity at 5 temperatures.Induced enzyme activities in three mediums were all low at 40℃.