Safety Evaluation Of Avermectin Nano Sustained-release Material On Harmonia Axyridix

Harmonia axyridix is a widely distributed predatory natural enemy in agricultural and forest ecosystems.It is also one of the most important natural enemies in our country.Currently,the chemical control still plays an important role in agricultural pest integrated management,therefore,natural enemies in habitats may be at risk to directly or indirectly exposure to chemical pesticides.In recent years,with the application and development of nanotechnology in the field of pesticide assistant research and development,nano additives are beneficial to improve the utilization of the active ingredients of pesticides,and the application prospect is broad.However,there is still a lack of research regarding the potential effects of pesticide nano-materials on natural enemy.In addition,previous studies have shown that using nanomaterials（lignin）to encapsulate avermectin can make avermectin nano sustained-release agents.Previous research in our laboratory has shown that the toxicity of avermectin nano sustained-release material to aphids is equivalent to that of the avermectin.Therefore,we select avermectin nano sustained-release material in this paper.The biosafety of avermectin nano sustained-release material to H.axyridis was comprehensively evaluated by carrying out laboratory bioassay,enzyme activity assay,predation function assay,construction of two-sex life table,transcriptome analysis and other research methods,and compared with avermectin,lignin and solvent.The main findings of this study are as follows:1.Toxicity of avermectin nano sustained-release material to H.axyridisThe results of the laboratory bioassay showed that the LC₅₀ of avermectin nano sustained-release material on 2nd instar larvae was 23.158 mg/L,while the LC₅₀ of avermectin on 2nd instar larvae was 8.940 mg/L.Therefore,the acute toxicity of avermectin nano sustained-release material is lower than that of avermectin to H.axyridis.2.Effect of sublethal dose avermectin nano sustained-release material on predation function of H.axyridisPredation function of 2nd instar larvae of the four treatment groups was compared.At five gradients density of Acyrthosiphon pisum（10,20,30,40,50 individuals/dish）,sublethal avermectin treatment significantly inhibited predation function of H.axyridis2nd instar larvae when compared with two control group.However,sublethal avermectin nano sustained-release material treatment significantly inhibited the predation function of H.axyridis 2nd instar larvae at lower prey densities,but not higher prey densities when compared with the two control groups.The results illustrated that both avermectin and avermectin nano sustained-release material treatment had adverse effect on predation function of 2nd instar larvae.However,under the condition of high density of prey,the avermectin nano sustained-release material group showed higher predation function than the avermectin group.3.Effect of sublethal dose of avermectin nano sustained-release material on development and reproduction of H.axyridisStatistical analysis of H.axyridis life history parameters was performed under the four treatments.We found out that treated with sublethal avermectin significantly extended the developmental period of the 2nd,3rd,and 4th instar larvae and reduced the pupation rate and pupal weight when compared with the two control groups.However,treated with sublethal avermectin nano sustained-release material only significantly prolonged the developmental period of the 4th instar larvae,and reduced the pupation rate,but had no significant effect on pupal weight.In addition,both two pesticide treatments had significantly reduced pre-adult survival rate.At the adult stage,treating with sublethal avermectin and avermectin nano sustained-release material significantly prolonged the total pre-oviposition period,shortened the longevity of female adults and reduced the adult fecundity when compared to the two control groups.In terms of population parameters,intrinsic growth rate（R）,finite rate of increase（λ）and net reproductive rate（R₀）was significantly lower in sublethal avermectin group when compared with two control groups,whereas no significant difference was observed between the sublethal avermectin nano sustained-release material treatment and the two control groups.The results indicated that the nano sustained-release material treatment reduced the inhibitory effect of avermectin on the growth and development of H.axyridis larvae and reproduction,and maintained the normal growth of the population.4.Effect of sublethal dose avermectin nano sustained-release material on enzyme activity of H.axyridisThe changes of four enzyme activities including catalase（CAT）,peroxidase（POD）,acetyl cholinesterase（ACh E）and glutathione S-transferase（GST）in 2nd instar larvae after treating with sublethal dose avermectin nano sustained-release material,avermectin,lignin and solvent for 24,48 and 72 h were measured.The results showed that sublethal avermectin treatment significantly induced the elevation of CAT and GST activities after 24 and 48 h,but sublethal avermectin nano sustained-release material significantly induced the enzyme activities of CAT and GST only at 48 h.When treated with sublethal avermectin,no significant effect was observed on POD and Ach E activities at 24,48 and 72 h.However,the sublethal avermectin nano sustained-release material was able to significantly induce the activity of POD at 48 h and had no significant effect on POD and Ach E at other time point.All the above results showed that the avermectin could rapidly and chronically induce the elevation of protective and detoxifying enzyme activity in H.axyridis,while the avermectin nano sustained-release material had delayed and shortened induction time of protective and detoxifying enzyme activity,which indicated that the avermectin nano sustained-release material had posed less abiotic stress to H.axyridis.5.Effect of sublethal dose of avermectin nano sustained-release material on the transcriptome level of H.axyridisTranscriptome sequencing analysis of H.axyridis in four treatments were conducted.And the reasults showed that compared with solvent control,there were 154differential expressed genes up-regulated and 195 genes down-regulated after treated with sublethal avermectin.When treated with sublethal avermectin nano sustained-release material,there were 63 differential expressed genes up-regulated and 101 genes down-regulated compared with lignin control.Compared to the solvent control group,the lignin treated group had 46 up-regulated different genes and 36 down-regulated different genes.Through KEGG enrichment analysis,we found that the differential expressed genes were mainly significantly enriched in the related pathways including detoxification metabolism,insect hormone synthesis and life span regulation.At the same time,we selected 15 differential expressed genes for q PCR validation,result showed that the transcriptome sequencing results were reliable.The above results show that compared with the avermectin,the avermectin nano sustained-release material has less influence on detoxication metabolism,insect hormone synthesis,life span regulation and other pathways,which shows that it has less abiotic stress on the H.axyridis again.In conclusion,this study demonstrated that the nanomaterial lignin encapsulated avermectin reduced its acute toxicity to H.axyridis,alleviated the inhibitory effects of avermectin on larval growth and development,predation function,and reproduction.Reduced abiotic stress imposed by avermectin on the physiological metabolism of H.axyridis.Maintained the population growth of H.axyridis.The research results of this article will provide a theoretical basis and data support for the spreading applications of nano-insecticides,as well as the protection and utilization of natural enemies.