| Phototaxis behavior of insect is one of the hotspots of behavioral researches.Pest-trapping lamp is an important means of physical control by phototaxis behavior.The species Mythimna separata and Helicoverpa armigera all belong to the Lepidoptera and are very different in trapping effect.However,there is no research report on the difference in phototaxis behavior between these two insects.In this study,we systematically compared the differences of phototaxis behavior between the two species,studied the similarities and differences between the external morphology and internal structure of the compound eye;constructed a comparative head transcriptome,identified and analyzed the types and quantities of opsin genes of M.separata and H.armigera;RT-QPCR and in situ hybridization techniques were used to compare the abundance,location and pattern of opsin gene expression.The main results were as follows:1.The comparison of phototaxis behavior between M.separata and H.armigera(1)The phototaxis rate of M.separata and H.armigera was different.Among the 14 wavelenghts(365 ~ 660 nm),the phototactic rate of female M.separata(19% ~ 45%)was significantly lower than that of female H.armigera(29% ~ 59%),while the phototactic rate of male M.separata(14% ~ 34%)was significantly lower than of male H.armigera(24% ~ 57%);the photophobism rate of female M.separata(10% ~ 34%)was significantly higher than that of female H.armigera(0% ~ 10%),while the photophobism rate of male M.separata(11% ~ 22%)was significantly higher than of male H.armigera(1% ~ 7%).At wavelength of 365 nm,total activity rate in female M.separata(53%)was significantly higher than of female H.armigera(35%).At wavelength of 520 nm,total activity rate in female M.separata(46%)was significantly lower than of female H.armigera(64%).At wavelength of 480 and 500 nm,total activity rate in male M.separata(34%,31%)was significantly lower than of male H.armigera(52%,48%).(2)The photocatalytic sensitive peaks were different.The phototaxis peaks of the female M.separata were 420 nm,480 nm,600 nm and 660 nm,the phototaxis peaks of the female H.armigera were 400 nm,480 nm and 520 nm;male M.separata were sensitive to the light wave of wavelength 380 nm,455 nm,570 nm,and 660 nm,and male H.armigerawere sensitive to the light wave of wavelength 400 nm,445 nm,480 nm,570 nm,600 nm,and 660 nm,and the M.separata had a high photophobism rate in the ultraviolet and red regions.(3)The rhythm of phototaxis behavior were different at the same wavelength between M.separata and H.armigera.In the female,the phototaxis of the Mythimna separata at the wavelength of 400 and 590 nm showed upward trend and increased then decreased trend,while the phototaxis of the H.armigera did not change at scotophase;at 480 and 500 nm wavelengths,the phototaxis rate of M.separata showed down trend,while the phototaxis rate of H.armigera increased then decreased.In the males,the phototaxis of the M.separata at 570,590,600,630,and 660 nm showed down trend,while the phototaxis of the H.armigera at the corresponding wavelength increased and then decreased.In addition,the photophobism rate of M.separata had a gradual upward trend in scotophase.2.Comparison of the morphology and microstructure of compound eyes of the M.separata and H.armigeraThe number and length of ommatrichia on the compound eye of the M.separata and H.armigera were significantly different.The M.separata eye was covered with long and dense ommatrichia(length: 132.26 ?m;diameter: 2.98 ?m),and the corneal surface of H.armigera appears glabrous with inconspicuous ommatrichia(length: 6.41 ?m;diameter: 0.63 ?m).The number of the ommatidia of the M.separata(6704)was greater than that of the H.armigera(5532),and the ratio of eye width/body length(0.088)was significantly smaller than that of the H.armigera(0.092);The M.separata eye radius curvature(720.09 ?m)was significantly larger than the radius of curvature of the H.armigera(649.79 ?m).A remarkable difference was that the number of cornea lamellae in H.armigera(25 layers)was much greater than in M.separata(19 layers);The slightly difference was observed in terms of the dioptric apparatus length,cornea radius curvature,cornea thickness,rhabdom length and Clear zone length.The distribution area of pigment particles was narrower in the H.armigera than M.separate,and closer to the cornea in scotophase in H.armigera.3.Comparison of head transcriptomes between M.separata and H.armigeraTranscriptome analysis indicated that 156254 and 124960 Unigenes were obtained from M.separata and H.armigera with average unigene length of 416 bp and 426 bp,respectively.Among all these unigenes,34457(22.05%)and 36000(28.80%)were annotated in M.separata and H.armigera,respectively.GO fucnction analysis indicated that 22949 and 22949 Unigenes were annotated in M.separata and H.armigera,accounting for 14.69% and 18.37% in total number of unigenes.KOG fucnction analysis indicated that 9794 unigene(6.27%)and 10437 unigene(8.35%)belonged to 26 fucntion classifications in M.separata and H.armigera,respectively?KEGG fucnction analysis indicated that 7976(5.10%)and 8996(7.20%)were belonged to 277 and 283 pathways,respectively.The comparative transcriptome of the heads of M.separata and H.armigera was constructed,and 8 and 7 opsin genes were obtained from M.separata and H.armigera respectively.Among these opsin genes,3 and 1 UV opsin gene were identified in M.separata and H.armigera,respectively;a specific opsin pteropsin was identified in H.armigera.Protein sequence homology were high in these two species(>89%)while certain amino acids were different in some locations.FPKM analysis indicated that the ratio of LW2/LW1 in M.separata(1/176)was significantly lower than that of H.armigera(1/87).4.The comparison of expression patterns of major opsin genes between M.separata and H.armigeraThe expression of head opsin showed signicantly diurnal rhythm in M.separata and showed diurnal rhythm in H.armigera without significance.The proportion of expression of lw,bl and uv showed differences in these two species.In head of female M.separata,MS-lw is 98.31 and 39.68 times in expression of the MS-uv and MS-bl,respectively;in head of female H.armigera,HA-lw is 8.37 and 10.77 times in expression of the HA-uv and HA-bl,respectively;in head of male M.separata,MS-lw is 67.73 and 35.36 times in expression of the MS-uv and MS-bl,respectively;in head of male H.armigera,HA-lw is 14.20 and 15.46 times in expression of the HA-uv and HA-bl,respectively.Among these three opsin genes,MS-uv and HA-bl was minimally expressed in head of M.separata and H.armigera,respectively.5.The comparison of tissue localization of visual protein genes in compound eyesThe expression of three opsin gene(HA-lw,HA-bl and HA-uv)in the compound eye of H.armigera was obviously higher in female than in male during photophase,while which was obviously higher in male than in female during scotophase.However,the expression of three gene(MS-lw,MS-bl and MS-uv)in the compound eye of M.separata showed no sexual difference in photophase and scotophase.In photophase,all three gene(lw,bl and uv)were expressed uniformly in rod cells,while in scotophase they were expressed intensively in distal end of rod cells.In present study,behavioral experiments showed that the phototaxis behaviors of M.separata and H.armigera were different in phototaxis rate,sensitive wavelength,and phototaxis rhythm.The morphology and microstructure of compound eyes of M.separata and H.armigera showed difference in the physiological structure.Difference in the number,type,expression,expression pattern and expression site of the opsin gene at the molecular level were revealed.These results systematically compared the differences in phototaxis behavior between M.separata and H.armigera,revealing the physiological structure and molecular basis of the difference in phototaxis behavior,laying the foundation for revealing the diversity of phototaxis behavior of insects and guiding role in establishing an efficient,specific,eco-friendly light trapping technique at the same time. |
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