A Primary Study On Optimization Of Rearing Conditions For Thitarodes Sp.and Infections Of The Insect By Ophiocordyceps Sinensis

The conditions of artificial rearing of Thitarodes sp.(Lepidoptera:Hepialidae),one of the hosts of the valuable fungus the Chinese caterpillar fungus(Ophiocordyceps sinensis),including temperature,illumination,diet and the ways to handle,etc were investigated by single facor test.The life cycle of Thitaraodes sp.has been completed in the laboratory that consists of pupa,adult,egg and larva.For its extremely long life cycle(usually 3?5 year in natural habitats and more than one year in laboratory),the insect is now in its larval stage and expected to pupate recently.52 kinds of medium were adopted to culture the mycelia of O.sinensis in order to induce the blastospores of the fungus,out of which 11 medium could be used to induce blastospore production.Measurement of pH values and osmotic pressures of 7 medium out of 52 indicated that the parameters were not important in blastospore production.The blatospores of O.sinensis are liable to degrade after a ralatively long term incubation(>30 d).Sufficient mycelia of O.sinensis were harvested by liquid fermentation for later use in this studis.Cellular immune responses of nonhost insects,including the larvae of Spodoptera litura(6th instar),Ostrinia furnacalis(5th instar),Helicoverpa armigera(5th instar),Bombyx mori(prepupal stage),and the pupae of Antheraea pernyi,to the blastospores of O.sinensis were observed under optical microscope by injection of the blastospores into the hemocoele of the insects.The observation showed that almost all the blastospores injected into the hemocoele of the nonhosts were encapsulated rapidly by the insect hemocytes.Therefore,attempt to use the insects other than Thitarodes as the host of O.sinensis is unpractical.5 means to inoculate Thitaraodes sp.with O.sinensis were tested,out of which 2 could be used as the ways of inoculation.The blastospores of O.sinensis proliferate in the larval hemocoele of Thitarodes sp.by budding and can reach very high concentration which makes the hemolymph cloudy.Fluorescence staining showed that the blastospore was unicellular with single nucleus,and teeming with lipid.The lethal causations Thitaraodes sp.larvae and pupae reared in the laboratory were investigated.The results showed that fungi,bacteria and nametodes were main pathogens causing the insect mortality,among which entomogenous fungi was the most important lethal factor.The entomogenous' fungi were isolated from 142 cadavers of Thitaraodes sp.and analysed by ITS sequencing.The analysis indicated that approximate 90%isolates were Paecilomyces farinosus.In brief,the optimization of larvae rearing,the ways to induce O.sinensis blastospores production,inoculation experiments,and analysis of pathogens isolated from Thitaroads sp.larvae and pupae may provide instructive advice to mass rearing Thitaraodes sp.,and further to produce the Chinese caterpillar fungus artificially.