Combine Biological Control Using Bacillus Thuringiensis And Habrobracon Hebetor (Say) For The Management Of Plodia Interpunctella (Hübner)

Many developing countries are confronted with acute food shortage,and this could probably be linked to their inability to protect and preserve their food produce from quality and quantity deterioration that result from microbial,vertebrate and insect pest infestation.The annual losses incurred in storage due to insect attack are estimated as 5%worldwide.Plodia interpunctella Hübner(Lepidoptera:Pyralidae) is a cosmopolitan insect infesting a broad range of commodities,including raw or processed cereal.The key objectives of this research were to:a.screen and evaluate some Bacillus thuringiensis isolates against P.interpunctella and determine the LC₅₀ of highly toxic strains,b.to evaluate the role of host-instar and refuge on the parasitization behavior of Habrobracon hebetor Say(Hymenoptera:Braconidae),c. Investigate the effect of combining B.thuringiensis with H.hebetor for management of P.interpunctella infestation and to assess the influence of B.thuringiensis on H. hebetor during this combination treatment,d.investigate the host-finding behavior in H.hebetor,by extracting and identifying kairomone from P.interpunctella responsible for eliciting parasitization behavior in the parasitoid.A study was conducted to examine the interactions between trophic levels during parasitism and host location by H.hebetor within a closed environment,by carrying out multiple tests to evaluate the role of refuge and host-instar on the mortality of P.interpunctella and on the emergence of H.hebetor.Results showed that H.hebetor was able to parasitize all(first-fourth) instars of P.interpunctella,but significantly fewer early(first and second) instars were parasitized.Interaction among instar x refuge had no significant effect on parasitism.In no-choice condition,only instar factor was observed to have any significant effect on parasitism at both 24 h and 48 h post treatment.While under choice condition:instar,refuge and not interaction abetween them,significantly affected parasitism by the wasp at both 24 h and 48 h post treatment.Generally,the percentage of early(first and second) instars parasitized was higher under no-choice condition than parasitism observed for the same larvae stages in choice condition.Parasitism appeared to increase in conjunction with the increase in exposure time.Significantly more fourth instar larvae were parasitized than other larvae from the other stages irrespective of the duration of exposure.Fewer hosts were parasitized in the presence of refuge as opposed to the higher parasitism observed in the absence of refuge.The non significant interaction between instar x refuge explained that,irrespective of the presence of refuge or not,percentage of host parasitized by H.hebetor also increased with increase in larval stage.Differences between male and female emergences appeared non significant.Results also revealed that only host-instar significantly affected parasitoid yield.It was however noted that parasitoid emergence was higher under no-choice condition than in choice.There was no significant difference between the parasitoid yield of third and fourth instar larvae. Based on the results from this investigation,H.hebetor could therefore be described as a weak competitor with an opportunistic and specialized behavior,thereby giving more insight to its ecology and biology.Through preliminary bioassays,the toxicity of a collection of 122 isolates of B. thuringiensis were evaluated against P.interpunctella.Afterwards,3 strains that demonstrated significantly greater potency than the standard strain(B.thuringiensis serovar kurstaki HD-1) against Lepidoptera were selected and analyzed in detail. Results indicated that 58.2%of the entire B.thuringiensis isolates evoked≥60% mortality to P.interpunctella and were thus considered toxic.Among these,13 isolates were distributed within the 90%to 100%mortality groups.The results also showed that different isolates belonging to the same serotype had different toxic level to P.interpunctella.The highest frequency of toxicity(causing 100%mortality of Indianmeal moth) was observed in isolates belonging to H₇,while the lowest frequency of toxicity was recorded in isolates belonging to H₈.Among the 122 B. thuringiensis isolates evaluated,3 three isolates exhibited a significantly(df=3,f= 48.35,P＜0.001) greater potency against P.interpunctella,with their LC₅₀ lower than the standard strain HD-1.Treatments with spore-crystal mixture caused higher larval mortality than soluble protein.The LC₅₀ s for these 3 isolates IMM-408,IMM-130 and IMM-368 are 1.24,2.86 and 4.62μg/ml respectively while that of isolate HD-1 was 8.48μg/ml.Of the 3 highly toxic strains,IMM-408 was found to be most toxic. SDS-PAGE of 3 isolates that yielded 100%mortality against P.interpunctella indicated that both isolates IMM-130 and IMM-368 contained～135 kDa and 65 kDa crystal proteins,similar to standard B.thuringiensis strain HD-1,while isolate IMM-408 contained～135 kDa crystal protein,and contained cry1Ab9,cry1Ca1,cry1Da1 and cry2 gene.The serological tests conducted showed that isolates IMM-130,IMM-368 and IMM-408 belong to serotypes H₃,H₄ and H₇ respectivelyFurthermore,the suitability of combining microbial pesticides and natural enemies for integrated pest management(IPM) of stored cereal in China was evaluated using laboratory assays.This was achieved by studying the interactions between B.thuringiensis,B.thuringiensis-intoxicated host larvae and H.hebetor during control of P.interpunctella.B.thuringiensis or H.hebetor alone caused 41.67%and 35.35%P.interpunctella larval mortality respectively.The B. thuringiensis-parasitoid combination treatment significantly evoked more P. interpunctella mortality(86%) than other treatments.Progeny development of H. hebetor was dependent upon its susceptibility to B.thuringiensis contaminated host. However,more eggs were significantly laid by H.hebetor on hosts placed in control (257 eggs) than on hosts in B.thuringiensis contaminated diets(93 eggs).Hatchability of H.hebetor eggs were not affected in all treatments,data between treatment and control did not vary significantly.Fewer wasps emerged from B.thuringiensis-parasitoid combine treatment than in none B.thuringiensis treatments.H.hebetor larval mortality was significantly higher in combine treatments than in control. Although the size of adult parasitoids that emerged from combine treatments appeared smaller than those in the control,their developmental period was not significantly different.Similarly,no significant difference was observed in the longevity of adult wasps emerging from both treatments.The sex ratio of H.hebetor progeny in control experiment was female biased in the ratio of 1:2.27 male:female and this trend is similar to the observations in combine treatment Therefore,since B.thuringiensis did not prevent parasitoid development,this investigations is suggestive that a combine treatment with B.thuringiensis and parasitoid release,would produce better protection against P.interpunctella.Lastly,the response of gravid female H.hebetor to extracts from clean diet, infested diet,larvae and frass of P.interpunctella was investigated.Result revealed that acetone and hexane extracts elicited searching behavior in H.hebetor.Extracts of frass significantly stimulated more parasitization behavior in the parasitoid than extracts of other P.interpunctella-related materials tested.It was also noted that experience plays a huge role in parasitoid foraging efficiency.Females that had previous contact with host produced stronger responses towards treated patch,than their inexperienced counterparts.The active components in P.interpunctella-related materials were more soluble in hexane,than in acetone,ethyl ether or methanol. Using silica gel flash column chromatography,crude extracts of larva and frass were fractionated,then the response of H.hebetor to each fraction was evaluated and most active fractions were analyzed by gas chromatography-mass spectrometry(GC-MS). Three main peaks were observed on the GC of both fractions.Result showed that extracts contained hydrocarbons.The hydrocarbons which are mainly saturated long chain alkanes(1,3-dimethyl benzene,2-acyl-1,3-cyclohexanediones) are present in both extracts.Other chemicals extracted from the active fractions are toluene, squalane,ethyl cyclohexane,nonacosane and hexadecane.These results showed that P.interpunctella-related materials that were tested contained infochemicals which elicits searching/ parasitization behavior in the parasitoid H.hebetor.And these compounds(kairomone),may be synthesized and applied to surface of stored produce in other to enhance H.hebetor parasitization performance.