Hemimetabolous genomes reveal molecular basis of termite eusociality

Hemimetabolous genomes reveal molecular basis of termite eusociality Mark C Harrison, 1∗ Evelien Jongepier, 1∗ Hugh M. Robertson, 2∗ Nicolas Arning, 1 Tristan Bitard-Feildel,1 Hsu Chao, 3 Christopher P. Childers, 4 Huyen Dinh, 3 Harshavardhan Doddapaneni, 3 Shannon Dugan, 3 Johannes Gowin, 5,6 Carolin Greiner,5,6 Yi Han, 3 Haofu Hu, 7 Daniel S.T. Hughes, 3 Ann-Kathrin Huylmans, 8 Carsten Kemena, 1 Lukas P.M. Kremer, 1 Sandra L. Lee, 3 Alberto Lopez-Ezquerra, 1 Ludovic Mallet, 1 Jose M. Monroy-Kuhn, 5 Annabell Moser, 5 Shwetha C. Murali, 3 Donna M. Muzny, 3 Saria Otani, 7 Maria-Dolors Piulachs, 9 Monica Poelchau, 4 Jiaxin Qu, 3 Florentine Schaub, 5 Ayako Wada-Katsumata,10 Kim C. Worley, 3 Qiaolin Xie, 11 Guillem Ylla, 9 Michael Poulsen, 7 Richard A. Gibbs, 3 Coby Schal, 10 Stephen Richards, 3 Xavier Belles, 9† Judith Korb, 5,6† Erich Bornberg-Bauer 1† 1Institute for Evolution and Biodiversity, University of M¨ unster, M¨ unster, Germany. 2Department of Entomology, University of Illinois at Urbana-Champaign, Urbana IL, USA. 3Human Genome Sequencing Center, Department of Human and Molecular Genetics, Baylor College of Medicine, Houston, TX, USA. 4USDA-ARS, National Agricultural Library, Beltsville, MD, USA. 5Evolutionary Biology & Ecology, University of Freiburg, Freiburg, Germany. 6Behavioral Biology, University of Osnabr¨ uck, Osnabr¨ uck, Germany. 7Ecology and Evolution, University of Copenhagen, Universitetsparken 15, 2100 Copenhagen, Denmark. 8Instititute of Science and Technology Austria, Klosterneuburg, Austria. 9Institut de Biologia Evolutiva, CSIC-University Pompeu Fabra, Barcelona, Spain. 10Department of Entomology and Plant Pathology, North Carolina State University, Raleigh, NC, USA. 11China National GeneBank, Beijing Genomics Institute(BGI)-Shenzhen,Shenzhen,518083,China †Corresponding authors. E-mail: [email protected] (XB); [email protected] (JK); [email protected] (EBB) ∗These authors contributed equally to this work. 1 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint Around 150 million years ago, eusocial termites evolved from within the cockroaches, 50 million years before eusocial Hymenoptera, such as bees and ants, appeared. Here, we report the first, 2GB genome of a cockroach, Blattella germanica, and the 1.3GB genome of the drywood termite, Cryptotermes secundus. We show evolutionary signatures of termite eusociality by comparing the genomes and transcriptomes of three termites and the cockroach against the background of 16 other eusocial and non-eusocial insects. Dramatic adaptive changes in genes underlying the production and perception of pheromones confirm the importance of chemical communication in the termites. These are ac- companied by major changes in gene regulation and the molecular evolution of caste determination. Many of these results parallel molecular mechanisms of eusocial evolution in Hymenoptera. However, the specific solutions are remarkably different, thus revealing a striking case of convergence in one of the major evolutionary transitions in biological complexity. Eusociality, the reproductive division of labour with overlapping generations and cooperative brood care,1 is one of the major evolutionary transitions in biology. 1 Although rare, eusociality has been observed2 in a diverse range of organisms, including shrimps, mole-rats and several insect lineages. 2, 3, 4 A particu-3 larly striking case of convergent evolution occurred within the holometabolous Hymenoptera and in the4 hemimetabolous termites (Isoptera), which are separated by over 350 my of evolution. 5 Termites evolved5 within the cockroaches around 150 mya, towards the end of the Jurassic, 6, 7 about 50 my before the first6 bees and ants appeared. 5 Therefore, identifying the molecular mechanisms common to both origins of7 eusociality is crucial to understanding the fundamental signatures of these rare evolutionary transitions.8 While the availability of genomes from many eusocial and non-eusocial hymenopteran species8 has allowed9 extensive research into the origins of eusociality within ants and bees, 9, 10, 11 a paucity of genomic data10 from cockroaches and termites has precluded large-scale investigations into the evolution of eusociality11 in this hemimetabolous clade.12 The conditions under which eusociality arose differ greatly between the two groups. Termites and13 cockroaches are hemimetabolous and so show a direct development, while holometabolous hymenopter-14 ans complete the adult body plan during metamorphosis. In termites, workers are immatures and only15 reproductive castes are adults, 12 while in Hymenoptera, adult workers and queens represent the primary16 division of labour. Moreover, termites are diploid and their colonies consist of both male and female17 workers, and usually a queen and king dominate reproduction. This is in contrast to the haplodiploid18 system found in Hymenoptera, in which all workers and dominant reproductives are female. It is therefore19 intriguing that strong similarities have evolved convergently within the termites and the hymenopterans,20 such as differentiated castes and a nest life with reproductive division of labour. The termites can be sub-21 divided into wood-dwelling and foraging termites. The former belong to the lower termites and produce22 simple, small colonies with totipotent workers that can become reproductives. Foraging termites (some23 2 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint lower and all higher termites) form large, complex societies, in which worker castes can be irreversible. 1224 For this reason, higher, but not lower, termites can be classed as superorganismal. 13 Similarly, within25 Hymenoptera, varying levels of eusociality exist.26 Here we provide insights into the molecular signatures of eusociality within the termites. We analysed27 the genomes of two lower and one higher termite species and compared them to the first cockroach genome,28 as a closely-related non-eusocial outgroup. Furthermore, differences in expression between nymphs and29 adults of the cockroach were compared to differences in expression between workers and reproductives of30 the three termites, in order to gain insights into how expression patterns changed along with the evolution31 of castes. Using fifteen additional insect genomes to infer background gene family turnover rates, we32 analysed the evolution of gene families along the transition from non-social cockroaches to eusociality33 in the termites. In this study we concentrated particularly on two hallmarks of insect eusociality, as34 previously described for Hymenoptera, with the expectation that similar patterns occurred along with35 the emergence of termites. These are the evolution of a sophisticated chemical communication, which is36 essential for the functioning of a eusocial insect colony 3, 14, 15 and major changes in gene regulation along37 with the evolution of castes. 9, 10 Additionally, we tested if transposable elements spurred the evolution38 of gene families that were essential for the transition to eusociality.39 Evolution of genomes, proteomes and transcriptomes40 We sequenced and assembled the genome of the German cockroach, Blattella germanica (Ectobiidae),41 and of the lower, drywood termite, Cryptotermes secundus (Kalotermitidae; for assembly statistics see42 Supplementary Table 1). The cockroach genome (2.0 Gb) is considerably larger than all three termite43 genomes. The genome size of C. secundus (1.30 Gb) is comparable to the higher, fungus-growing termite,44 Macrotermes natalensis , (1.31 Gb, Termitidae) 16 but more than twice as large as the lower, dampwood45 termite, Zootermopsis nevadensis (562 Mb, Termopsidae).17 The smaller genomes of termites compared46 to the cockroach are in line with previous size estimations based on C-values. 18 The proteome of B. ger-47 manica (29,216 proteins) is also much larger than in the termites, where we find the proteome size in48 C. secundus (18,162) to be similar to the other two termites (M. natalensis: 16,140; Z. nevadensis: 15,459;49 Fig. 1). In fact, the B. germanica proteome was the largest among all 21 arthropod species analysed here50 (Fig. 1). Strong evidential support for over 80% of these proteins in B. germanica (see Supplementary51 Material) and large expansions in many manually annotated gene families offer high confidence in the52 accuracy of this proteome size.53 We also compared gene expression between the four species. When comparing worker expression54 with queen expression in the termites and nymph expression (5th and 6th instars) with adult female55 3 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint expression in B. germanica , we found shifts in specificity of expression for termites compared to the56 cockroach in several gene families (Fig. 2). It has been previously reported for the primitively eusocial57 paper wasp, Polistes canadensis, that the majority of caste-biased genes, especially those upregulated in58 workers, are novel genes. 19 The authors suggested that this may be a feature of early eusociality. We59 did not find the same pattern for the termites. Species-specific genes (those without ortholog) were not60 enriched for differentially expressed genes in any of the termites, with slight peaks among Blattodea- and61 Isoptera-specific genes (Supplementary Figure 1).62 Gene family expansions assisted by TEs in termites63 The transitions to eusociality in ants 10 and bees9 have been linked to major changes in gene family sizes.64 Similarly, we detected significant gene family changes on the branch leading to the termites (7 expansions65 and 10 contractions; Supplementary Figure 2, Supplementary Table 2). The numbers of species-specific,66 significant expansions and contractions of gene families varied within termites (Z. nevadensis: 15/5; C. se-67 cundus: 27/3; M. natalensis: 24/20; Supplementary Figure 2 & Supplementary Tables 3-5). Interestingly,68 in B. germanica we measured 93 significant gene family expansions but no contractions (Supplementary69 Table 6), which contributed to the large proteome.70 The termite and cockroach genomes contain a higher level of repetitive DNA compared to the hy-71 menopterans we analysed (Fig. 1). C. secundus and B. germanica genomes both contain 55% repetitive72 content (Supplementary Table S7), which is higher than in both Z. nevadensis (28%) and the higher73 termite, M. natalensis (46%; Fig. 1). 20 As also found in Z. nevadensis and M. natalensis ,20 LINEs and74 especially the subfamily BovB were the most abundant transposable elements (TEs) in the B. germanica75 and C. secundus genomes, indicating that a proliferation of LINEs may have occurred in the ancestors76 of Blattodea (cockroaches and termites).77 We hypothesised that these high levels of TEs may be driving the high turnover in gene family sizes78 within the termites and B. germanica .21 Expanded gene families indeed had more repetitive content79 within 10 kb flanking regions in all three termites (p< 1.3x10−8; Wald t-test; Supplementary Tables 8-9),80 in particular in the higher termite M. natalensis . In contrast, gene family expansions were not correlated81 with TE content in flanking regions for B. germanica. These results suggest a major expansion of LINEs82 at the root of the Blattodea clade contributed to the evolution of gene families within termites, likely via83 unequal crossing-over; 21 however, the expansions in B. germanica were not facilitated by TEs. It can84 therefore be speculated that the large expansion of LINEs within Blattodea allowed the evolution of gene85 families which ultimately facilitated the transition to eusociality.86 4 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint Massive expansion and positive selection of Ionotropic Receptors87 Insects perceive chemical cues from toxins, pathogens, food and pheromones with three major families88 of chemoreceptors, the Odorant (ORs), Gustatory (GRs) and Ionotropic (IRs) Receptors. 22 Especially89 ORs have been linked to colony communication in eusocial Hymenoptera, where they abound. 14, 15, 2390 Interestingly, as previously detected for Z. nevadensis ,17 the OR repertoire is substantially smaller in91 B. germanica and all three termites compared to hymenopterans. IRs, on the other hand, which are92 less frequent in hymenopterans, are strongly expanded in the cockroach and termite genomes (Fig. 393 & Supplementary Figure 3). Intronless IRs, which are known to be particularly divergent, 24 show the94 greatest cockroach- and Blattodea-specific expansions (Fig. 3a, Blattodea-, Cockroach- and Group D-95 IRs). By far the most IRs among all investigated species were found in B. germanica (455 complete gene96 models), underlining that the capacity for detecting many different kinds of chemosensory cues is crucial97 for this generalist that thrives in challenging, human environments. In line with a specialisation in diet98 and habitat, the total number of IRs is lower within the termites ( Z. nevadensis : 141; C. secundus: 135;99 M. natalensis : 75). Nevertheless, IRs are more numerous in termites than in all other analysed species100 (except Nasonia vitripennis : 111). This is strikingly similar to the pattern for ORs in Hymenoptera,101 which are also highly numerous in non-eusocial outgroups as well as in eusocial species. 14, 23, 25102 We scanned each IR group for signs of species-specific positive selection. Within the Blattodea-specific103 intronless IRs, we found several codon positions under significant positive selection for the higher termite,104 M. natalensis (codeml site models 7 & 8; p < 1.7x10−10). The positively evolving codons are situated105 within the two ligand-binding lobes of the receptors (Fig. 3c), showing that a diversification of ligand106 specificity has occurred along with the transition to higher eusociality and a change from wood-feeding to107 fungus-farming in M. natalensis . Only two IRs were differentially expressed between nymphs and adult108 females in B. germanica. Underlining a change in expression along with the evolution of castes, we found109 35 IRs to be differentially expressed between workers and queens in Z. nevadensis , 11 in C. secundus110 and 10 in M. natalensis (Fig. 2, Supplementary Table 10). The possible role of IRs in pheromonal111 communication has been highlighted both in the cockroach Periplaneta americana 26 and in Drosophila112 melanogaster,27 where several IRs show sex-biased expression.113 One group of ORs (orange clade in Fig. 3b) is evolving under significant positive selection at codon114 positions within the second transmembrane domain in M. natalensis (codeml site model; p = 1.1x10−11)115 and C. secundus (p = 5.6x10−16; Fig. 3d). Such a variation in the transmembrane domain can be related116 to ligand binding specificity, as has been shown for a polymorphism in the third transmembrane domain for117 an OR in D. melanogaster,28, 29 adding further support for an adaptive evolution of chemoreceptors, in line118 with the greater need for a sophisticated colony communication in the termites. Similar to IRs, a higher119 5 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint proportion of ORs were differentially expressed between workers and queens in the three termites than120 between nymphs and adults in the cockroach (Fig. 2; Supplementary Table 11), highlighting a change121 in expression and function along with the transition to eusociality. The evolution of chemoreceptors122 along with the emergence of the termites can also be related to adaptation processes and changes in diet123 compared to the cockroach. Experimental verification will help pinpoint which receptors are particularly124 important for communication.125 CHC producing enzymes have evolved caste-specificity126 Despite their different ancestry, both termites and eusocial hymenopterans are characterised by the pro-127 duction of caste-specific cuticular hydrocarbons (CHCs), 30, 31, 32 which are often crucial for regulating128 reproductive division of labour and chemical communication. Accordingly, we find changes in the ter-129 mites in three groups of proteins involved in the synthesis of CHCs: desaturases (introduction of double130 bonds33), elongases (extension of C-chain length 34) and CYP4G1 (last step of CHC biosynthesis 35).131 Desaturases are thought to be important for division of labour and social communication in ants. 36132 As previously described for ants, 36 Desat B genes are the most abundant desaturase family in the ter-133 mites and the cockroach (Supplementary Table 12), especially in M. natalensis where we found ten gene134 copies (significant expansion; p = 0.0003; Supplementary Table 5; Supplementary Figure 4). As in ants,135 especially the First Desaturases (Desat A - Desat E) vary greatly in their expression between castes and136 species in the three termites (Fig. 2; Supplementary Table 13). 36 In contrast to ants, where these genes137 are under strong purifying selection, 36 we found significant positive selection within the Desat B genes138 for the highly eusocial termite, M. natalensis , (codeml site models 7 & 8; p = 1.1x10 −16), indicating a139 diversification in function, possibly related to their greater diversification of worker castes (major and mi-140 nor workers, major and minor soldiers). Although desaturases are often discussed in the context of CHC141 production and chemical communication, their biochemical roles are quite diverse, 36 and the positive142 selection we observe for M. natalensis may, at least in part, be related to their rather different ecology of143 foraging and fungus farming rather than nest mate recognition. Future experimental verification of the144 function of these genes will help better understand these observed genomic and transcriptomic patterns.145 Underlining an increased importance of CHC communication in termites, the expression patterns of146 elongases (extension of C-chain length) differ considerably in the termites compared to the cockroach147 (Fig. 2; Supplementary Table 14). In contrast to B. germanica , in which elongases are both nymph-148 (5 genes) and adult-biased (4 genes), only one or two elongase genes in each termite are queen-biased149 in their expression, while many are worker-biased. As with the desaturases, a group of M. natalensis150 elongases also reveal significant signals of positive selection (codeml branch-site test; p = 4x10−4), further151 6 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint indicating a greater diversification of CHC production in this higher termite.152 The last step of CHC biosynthesis, the production of hydrocarbons from long-chain fatty aldehy-153 des, is catalyzed by a P450 gene, CYP4G1, in D. melanogaster .35 We found one copy of CYP4G1 in154 B. germanica, Z. nevadensis and C. secundus , but three copies in M. natalensis, reinforcing the greater155 importance of CHC synthesis in this higher termite. Corroborating the known importance of maternal156 CHCs in B. germanica,37 CYP4G1 is over-expressed in female adults compared to nymphs (Fig. 2; Sup-157 plementary Table 15). In each of the termites, however, CYP4G1 is more highly expressed in workers158 (or kings in C. secundus ) compared to queens (Fig. 2; Supplementary Table 15), adding support that,159 compared to cockroach nymphs, a change in the dynamics and turnover of CHCs in termite workers has160 taken place.161 Changes in gene regulation in termites162 The development of distinct castes underlying division of labour is achieved via differential gene expres-163 sion. Major changes in gene regulation have been reported as being central to the transition to eusociality164 in bees 9 and ants.10 Accordingly, we found major changes in putative DNA methylation patterns (levels165 per 1-to-1 ortholog) among the termites compared to four other hemimetabolous insect species (Fig.166 4a). This is revealed by CpG depletion patterns (CpG o/e), a reliable predictor of DNA methylation, 38, 39167 correlating more strongly between the termites than among any of the other analysed hemimetabolous168 insects (Fig. 4). In other words, within orthologous genes, predicted DNA methylation levels differ169 greatly between termites and other hemimetabolous species but remain conserved among termite species.170 Predicted levels of DNA methylation correlated negatively with caste-specificity of expression for171 each of the termites. This is confirmed by a positive correlation between CpG o/e (negative association172 with level of DNA methylation) and absolute log 2-fold change of expression between queens and workers173 (Pearson’s r = 0.32 to 0.36; p < 2.2x10−16). The caste-specific expression of putatively unmethylated174 genes in termites is reflected in the enrichment of GO terms related to sensory perception, regulation175 of transcription, signalling and development, whereas methylated genes are mainly related to general176 metabolic processes (Fig. 4b, Supplementary Table 16). These results show strong parallels to findings177 for eusocial Hymenoptera.40, 41, 42, 43 This is in stark contrast to the non-eusocial cockroach,B. germanica,178 where there was only a very weak relationship between CpGo/e and differential expression between nymphs179 and adult females (r = 0.14), nor were any large differences apparent in enriched GO terms between180 putatively methylated and non-methylated genes (Fig. 4b).181 Our results argue in favour of a diminished role of DNA methylation in caste-specific expression182 within eusocial insects, as recently shown. 38, 44 In fact, DNA methylation appears to be important for183 7 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint the regulation of house-keeping genes because predicted methylated genes are related to general biological184 processes (further supported by lower CpG o/e within 1-to-1 orthologs than in non-conserved genes), 45185 while caste-specific genes are ’released’ from this type of gene regulation. However, a recent study linked186 caste-specific DNA methylation to alternative splicing in Z. nevadensis .46187 Major biological transitions are often accompanied by expansions of transcription factor (TF) fam-188 ilies, such as genes containing zinc-finger (ZF) domains. 47 We also observed large differences in ZF189 families within the termites compared to B. germanica . Many ZF families were reduced or absent in190 termites, while different, unrelated ZF gene families were significantly expanded (Supplementary Tables191 2-6). Queen-biased genes were significantly over-represented among ZF genes for each of the termites (p192 < 2 x 10−10;χ2 test; Supplementary Table 17), indicating an important role of ZF genes in the regulation193 of genes related to caste-specific tasks and colony organisation in the termites. This is in contrast to the194 significant under-representation of differentially expressed ZF genes within B. germanica (p = 4.8 x 10−5;195 χ2-test). Interestingly, two other important TF families (bHLH and bZIP),47 which were not expanded in196 the termites, showed no caste-specific expression pattern (p > 0.05), except bZIP genes, in which queen-197 biased genes were marginally over-represented for M. natalensis (p = 0.049). These major upheavals in198 ZF gene families and their caste-specific expression show that major changes in TFs accompanied the199 evolution of termites, strikingly similar to the evolution of ants. 10200 Evolution of genes related to molting and metamorphosis201 Hemimetabolous eusociality is characterised by differentiated castes, which represent different develop-202 mental stages. This is in contrast to eusocial Hymenoptera, in which workers and reproductives are adults.203 While cockroaches develop directly through several nymphal stages before becoming reproductive adults,204 termite development is more phenotypically plastic, and workers are essentially immatures (Fig. 2).205 In wood-dwelling termites, such as C. secundus and Z. nevadensis , worker castes are non-reproductive206 immatures that are totipotent to develop into other castes, while in the higher termite, M. natalensis ,207 workers can be irreversibly defined instars. It is therefore clear that a major change during the evolution208 of termites occurred within developmental pathways. Accordingly, we found changes in expression and209 gene family size of several genes related both to molting and metamorphosis.210 In the synthesis of the molting hormone, 20-hydroxyecdysone, the six Halloween genes (5 Cytochrome211 P450s and a Rieske-domain oxygenase) play a key role. 48, 49 Only one Halloween gene, Shade (Shd;212 CYP314A1), which mediates the final step of 20-hydroxyecdysone synthesis, is differentially expressed213 between the final nymphal stages and adults females in B. germanica (Fig. 2; Supplementary Table 18),214 consistent with its role in the nymphal or imaginal molt. In the three termites, the Halloween genes215 8 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint show varying caste-specific expression (Fig. 2; Supplementary Table 18), showing that ecdysone plays216 a significant role in the regulation of caste differences. Ecdysteroid kinase genes (EcK), which convert217 the insect molting hormone into its inactive state, ecdysone 22-phosphate, for storage, 50 are only over-218 expressed in female adults compared to nymphs in B. germanica (16/51 genes, Fig.2, Supplementary219 Table 19). In termites, however, where the gene copy number is reduced (18 to 20 per species), these220 important molting genes appear to have evolved worker-specific functions (Fig. 2; Supplementary Table221 19).222 Whereas 20-hydroxyecdysone promotes molting, juvenile hormone (JH) represses imaginal develop-223 ment in pre-adult instars. 51 JH is important in caste differentiation in eusocial insects, including ter-224 mites.12, 52 Hemolymph juvenile hormone binding proteins (JHBP), which transport JH to its target225 tissues,53 are reduced within the termites (21 to 33 genes) but significantly expanded in B. germanica226 (51 copies; p = 0018; Supplementary Table 6). Thirteen of the JHBP genes are over-expressed in adult227 females and only 8 in nymphs in B. germanica (Fig. 2, Supplementary Table 20). In both Z. nevadensis228 and M. natalensis , on the other hand, JHBPs are significantly more worker-biased (p < 0.01, χ2 test;229 Supplementary Table 20; Fig. 2). In C. secundus , expression is more varied, with 4 worker-biased, 7230 king-biased and 2 queen-biased genes (Fig. 2; Supplementary Table 20).231 These changes in copy number and caste-specific expression of genes involved in molting and meta-232 morphosis within termites compared to the German cockroach demonstrate that changes occurred in the233 control of the developmental pathway along with the evolution of castes. However, this interpretation234 needs to be experimentally verified.235 Conclusions236 These results, considered alongside many studies on eusociality in Hymenoptera, 9, 10, 14, 36 provide evi-237 dence that major changes in gene regulation and the evolution of sophisticated chemical communication238 are fundamental to the transition to eusociality in insects. Strong changes in DNA methylation patterns239 correlated with broad-scale modifications of expression patterns. Many of these modified expression pat-240 terns remained consistent among the three studied termite species and occurred within protein pathways241 essential for eusocial life, such as CHC production, chemoperception, ecdysteroid synthesis and JH trans-242 port. The stronger patterns we observe for M. natalensis , especially within genes linked to chemical243 communication, such as the expansion of Desat B and CYP4G1 genes and significant positive selection244 in desaturases, elongases and in IRs, may be associated with this termite’s higher level of eusociality and245 its status as a superoganism. 13 The analysis of further higher and lower termites would shed light on the246 generality of these patterns and possibly assist in the distinction between the influences of ecological and247 9 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint eusocial traits.248 Many of the mechanisms implicated in the evolution of eusociality in the termites occurred conver-249 gently around 50 my later in the phylogenetically distant Hymenoptera. However, several details are250 unique due to the distinct conditions within which eusociality arose. One important difference is the251 higher TE content within cockroaches and termites, which likely facilitated changes in gene family sizes,252 supporting the transition to eusociality. However, the most striking difference is the apparent importance253 of IRs for chemical communication in the termites, compared to ORs in Hymenoptera. According to our254 results, the non-eusocial ancestors of termites possessed a broad repertoire of IRs, which favoured the255 evolution of important functions for colony communication in these chemoreceptors within the termites,256 whereas in the solitary ancestors of eusocial hymenopterans ORs were most abundant. 14, 25 The parallel257 expansions of different chemoreceptor families in these two independent origins of eusociality indicate that258 convergent selection pressures existed during the evolution of colony communication in both lineages.259 10 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint METHODS260 Genome sequencing and assembly261 Genomic DNA from a single Blattella germanica male from an inbred line (strain: American Cyanamid262 = Orlando Normal) was used to construct two paired-end (180 bp and 500 bp inserts) and one of the two263 mate pair libraries (2 kb inserts). An 8kb mate pair library was constructed from a single female. The264 libraries were sequenced on an Illumina HiSeq2000 sequencing platform. The 413 Gb of raw sequence data265 were assembled with Allpaths LG, 54 then scaffolded and gap-filled using the in-house tools Atlas-Link266 v.1.0 (https://www.hgsc.bcm.edu/software/atlas-link) and Atlas gap-fill v.2.2. For Cryptotermes267 secundus, three paired-end libraries (250 bp, 500 bp and 800 bp inserts) and three mate pair libraries268 (2 kb, 5 kb and 10 kb inserts) were constructed from genomic DNA that was extracted from the head and269 thorax of 1 000 individuals, originating from a single, inbred field colony. The libraries were sequenced on270 an Illumina HiSeq2000 sequencing platform. The C. secundus genome was assembled using SOAPdenovo271 (v.2.04)55 with optimised parameters, followed by gapcloser (v1.10, released with SOAPdenovo) and kgf272 (v1.18, released with SOAPdenovo).273 Transcriptome sequencing and assembly274 For annotation purposes, twenty-two whole body RNAseq samples from various developmental stages275 were obtained for B. germanica . For C. secundus RNAseq libraries were obtained for three workers,276 four queens and four kings, based on degutted, whole body extracts. In addition, we sequenced 10277 M. natalensis RNAseq libraries from three queens, one king and six pools of workers. All libraries were278 constructed using the Illumina (TruSeq) RNA-Seq kit.279 For protein coding gene annotation, B. germanica reads were assembled with de novo Trinity (version280 r2014-04-13).56 The C. secundus reads were assembled using i) Cufflinks on reads mapped with TopHat281 (version2.2.1),57, 58 ii) de novo Trinity;56 and iii) genome-guided Trinity on reads mapped with TopHat.282 Repeat annotation283 A custom C. secundus and B. germanica repeat library was constructed using a combination of homology-284 based and de novo approaches, including RepeatModeler/RepeatClassifier ( http://www.repeatmasker.285 org/RepeatModeler.html), LTRharvest/LTRdigest 59 and TransposonPSI ( http://transposonpsi.286 sourceforge.net/). The ab initio repeat library was complemented with the RepBase (update 29-287 11 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint 08-2016)60 and SINE repeat databases, filtered for redundancy with CD-hit and classified with Repeat-288 Classifier. RepeatMasker (version open-4.0.6, http://www.repeatmasker.org) was used to mask the289 C. secundus and B. germanica genome. Repeat content for the other studied species (Fig. 1) was290 obtained from the literature. 61, 62, 63, 64,65, 66, 67291 Protein-coding gene annotation292 The B. germanica genome was annotated with Maker (version 2.31.8), 68 using (i) the species-specific293 repeat library, (ii) B. germanica transcriptome data (22 whole body RNAseq samples), and (iii) the swis-294 sprot/uniprot database (last accessed: 21-01-2016) plus the C. secundus and Zootermopsis nevadensis295 protein sequences for evidence-based gene model predictions. AUGUSTUS (version 3.2), 69 GeneMark-ES296 Suite (version 4.21) 70 and SNAP 71 were used for ab initio predictions. Cryptotermes secundus protein-297 coding genes were predicted using homology-based, ab initio and expression-based methods, and inte-298 grated into a final gene set (see Supplementary Material). Gene structures were predicted by GeneWise.72299 The ab initio annotations were predicted with AUGUSTUS73 and SNAP,71 retained if supported by both300 methods and integrated with the homology-based predictions using GLEAN.74 Transcriptome-based gene301 models were merged with PASA75 and tested for coding potential with CPC76 and OrfPredictor.77 PASA302 gene models were merged with the homology-based and ab initio gene set, retaining the PASA models in303 case of overlap. Desaturases, elongases, chemosensory receptors, Cytochrome P450’s and genes involved304 in the juvenile hormone pathway were manually curated in Blattodea.305 Differential gene expression306 The C. secundus and M. natalensis RNAseq libraries, were complemented with nine publishedZ. nevaden-307 sis libraries, yielding 2 to 6 libraries from workers, queens and kings for each termite. These were com-308 pared to six of the B. germanica libraries: two from 5 th instar nymphs, two from 6 th instar nymphs309 and two from adult females. Reads were mapped to the genome using HiSat2. 78 Read counts per gene310 where obtained using htseq-count and DESeq2 79 was used for differential expression analysis. Differen-311 tial expression analysis between kings (M), queens (F) and workers (majors and minors combined for312 M. natalensis ) was assessed for the termites. For B. germanica we evaluated the differential expression313 between adults and the two last nymphal stages combined, with the assumption that the final nymphal314 stages are homologous to termite workers and the adult females are homologous to termite queens. Genes315 were considered significantly differentially expressed if p |1| in order to316 account for allometric differences as recommended by Montgomery and Mank. 80317 12 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint Protein orthology318 In addition to B. germanica , C. secundus , Z. nevadensis and M. natalensis , 16 other insect proteomes319 were included in our analyses; L. migratoria, R. prolixus, E. danica, D. melanogaster, A. aegypti, T. cas-320 taneum, N. vitripennis , P. canadensis , A. mellifera , H. saltator , L. humile , C. floridanus , P. barbatus,321 S. invicta , A. echinatior and A. cephalotes ; as well as for the centipede, S. maritima , as an outgroup322 (for sources see Supplementary Table 22). These proteomes were grouped into orthologous clusters with323 OrthoMCL,81 with a granularity of 1.5.324 IR and OR identification, phylogeny and structure325 Ionotropic receptors (IRs) were identified using two custom Hidden Markov Models (HMMs) obtained326 with hmmbuild and hmmpress of the HMMER suite.82 The first HMM comprises the IR’s ion channel and327 ligand-binding domain based on a MAFFT83 protein alignment of 76 IRs from 15 species (Supplementary328 Table 23). The second HMM was built to distinguish IRs from iGluRs, IR8a and IR25a, which have329 an additional amino-terminal domain (ATD). 24 For this we built an HMM from 48 protein sequences330 (Supplementary Table 23). The proteomes were scanned with pfam scan and the two custom HMMs,331 where proteins that matched the IR HMM, but not the ATD HMM were annotated as IRs. ORs were332 identified based on the Pfam domain PF02949 (7tm Odorant receptor).333 Multiple sequence alignments of IRs and ORs were obtained with hmmalign,82 using the Pfam OR334 HMM PF02949 and custom IR HMM to guide the alignment. Gene trees were computed with FastTree 84335 (options: -pseudo -spr 4 -mlacc 2 -slownni) and visualised with iTOL v3. 85 Putative IR ligand-336 binding residues and structural regions were identified based on the alignments with D. melanogaster IRs337 and iGluRs of known structure. 86338 Gene family expansions and contractions339 For the analyses of gene family expansions and contractions, the hierarchical clustering algorithm340 MC-UPGMA87 was used, with a ProtoLevel cutoff of 80. 88 Protein families were further divided into341 sub-families if they contained more than 100 proteins in a single species, or more than an average of 35342 proteins per species. Proteins were blasted against the RepeatMasker TE database (E-value 50% of the proteins were identified as transposable elements were discarded. Clade- and344 species-specific protein family expansions and contractions, were identified with CAFE v3.0 89 using the345 same protocol as 9, 10 (see also Supplementary Material).346 13 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint TE-facilitated expansions347 The repeat content in the 10 kb flanking regions of B. germanica , C. secundus, Z. nevadensis and348 M. natalensis genes was calculated using bedtools. 90 CDS’ from neighbouring genes were removed and349 the repeat content was analysed using Generalized Linear Mixed Models (glmmPQL implemented in the350 R91 package MASS92) with binomial error distribution. Fixed predictors included gene family expansion,351 species ID and their interaction. Cluster ID was fitted as random factor to avoid pseudo-replication.352 Significance was assessed based on the Wald-t test (R package aod93) at α <0.05. Main and interaction353 effects for each of the genomic regions are listed in table S8. Model parameters are listed in table S8.354 Tests for positive selection355 To test for positive selection within gene families of interest, (i) site model tests 7 and 8 were performed356 (model = 0; NSsites = 7 8) on species-specific CDS alignments or ii) branch-site test (model = 2; NSsites357 = 2; fix omega = 1 for null model and 0 for alternative model) on multi-species alignments. Protein358 sequences were aligned using MAFFT 83 with the E-INS-i strategy, and CDS alignments were created359 using pal2nal.pl. 94 Phylogenetic trees were created with FastTree. 84 Alignments were trimmed using360 Gblocks (settings: -b2 = 21; -b3 = 20; -b4 = 5; -b5 = a). Models were compared using LR test and361 where p < 0.05, Bayes Empirical Bayes (BEB) results were consulted for codon positions under positive362 selection.363 CpG depletion patterns and GO enrichment364 To estimate DNA methylation we compared observed to expected CpG counts within CDS sequences.38, 39365 A low CpG o/e indicates a high level of DNA methylation, as the cytosine of methylated CpGs often366 mutate to thymines. Expected CpG counts were calculated by dividing the product of cytosine and367 guanine counts by the sequence length. The PCA in figure 4 was created using the R function prcomp368 on log transformed CpG o/e values for all 1-to-1 orthologs for the seven hemimetabolous species. These369 orthologs were extracted from the OrthoMCL results. The 3D plot was created with the plot3d command370 from the R package rgl.371 CpG depleted (first quartile) and enriched genes (fourth quartile) were tested for enrichment of Gene372 Ontology terms. Pfam protein domains were obtained for B. germanica , Z. nevadensis , C. secundus373 and M. natalensis protein sequences using PfamScan. 95 Corresponding GO terms were obtained with374 Pfam2GO. GO-term over-representation was assessed using TopGO96 package in R. Enrichment analysis375 14 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint was performed using the weight algorithm selecting nodesize=10 to remove terms with less than 10376 annotated GO terms. 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Alexa, A. & Rahnenfuhrer, J. topgo: enrichment analysis for gene ontology. R package version 2586 (2010).587 97. Bell, W. J., Roth, L. M. & Nalepa, C. A. Cockroaches: ecology, behavior, and natural history (JHU588 Press, Baltimore, Maryland, 2007).589 Acknowledgements590 We thank Oliver Niehuis for allowing use of the unpublished E. danica genome, J¨ urgen Gadau and591 Chris Smith for comments and advice on the manuscript, Jonathan Schmitz for assistance with analyses592 and proof-reading the manuscript. JK thanks Charles Darwin University (Australia), especially593 Prof. Stephen Garnett and the Horticulture and Aquaculture team for providing logistic support to594 collect C. secundus . The Parks and Wildlife Commission, Northern Territory, the Department of the595 Environment, Water, Heritage and the Arts gave permission to collect (Permit number 36401) and596 export (Permit WT2010-6997) the termites. USDA is an equal opportunity provider and employer.597 MCH and EJ supported by DFG grant BO2544/11-1 to EBB. JK by University of Osnabr¨ uck and DFG598 grant KO1895/16-1. XB and MDP supported by Spanish Ministerio de Econom´ ıa y Competitividad599 (CGL2012-36251 and CGL2015-64727-P to XB, and CGL2016-76011-R to MDP), including FEDER600 funds, and by Catalan Government (2014 SGR 619). CS: grants from US Department of Housing and601 Urban Development (NCHHU-0017-13), National Science Foundation (IOS-1557864), Alfred P. Sloan602 Foundation (2013-5-35 MBE), National Institute of Environmental Health Sciences (P30ES025128) to603 Center for Human Health and the Environment, and Blanton J. Whitmire Endowment. MP is supported604 by a Villum Kann Rasmussen Young Investigator Fellowship (VKR10101).605 606 Author contributions607 E.B-B. conceived, managed and coordinated the project; M.C.H., E.J. and H.M.R. are joint first authors.608 J.K. conceived and managed C. secundus sequencing project, coordinated termite-related analyses; S.R.609 conceived and managed B. germanica sequencing project; S.R., S.D., S.L.L., H.C., H.V.D., H.D., Y.H.,610 J.Q., S.C.M., D.S.T.H., K.C.W., D.M.M. and R.A.G. carried out B. germanica library construction,611 genome sequencing and assembly; C.S., A.W.K. provided biological material through full-sib mating for612 B. germanica; X.B. and C.S. co-managed the B. germanica analysis; M.P. and C.P.C. implemented Web613 Apollo data traces; S.O. and M.P. provided biological material for M. natalensis ; C.G., J.G., J.M.M.-K.,614 A.M., F.S., H.H. & J.K. coordinated and carried out DNA and RNA sequencing for C. secundus; M-D.P.,615 23 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint X.B. and G.Y. coordinated transcriptome sequencing of B. germanica ; L.M. performed automated616 gene prediction on C. secundus ; E.J. and N.A. improved assembly and annotation for B. germanica &617 C. secundus , compared and analysed genome sizes and quality. E.J., N.A. and L.P.M.K. analysed TEs;618 M.C.H. analysed CpG patterns and signatures of selection; T.B-F., E.J., C.K., L.P.M.K. and A.L-E.619 performed orthology and phylogenetic analyses; L.P.M.K., E.J., H.M.R. and M.C.H. analysed gene620 family evolution; A.L-E., E.J. and M.C.H. analysed transcriptomes and performed DE analyses; T.B.-F.621 and A.L-E. carried out orthoMCL clustering; H.M.R. corrected gene models for chemoreceptors; C.K.622 and E.J. for desaturases and elongases; A-K.H. and M.C.H. of Cytochrome p450s; E.B-B and M.C.H623 drafted and wrote the manuscript; X.B., M-D.P., J.K. contributed to sections of the manuscript; E.J.,624 L.P.M.K., A.L-E., C.K., M.C.H. wrote and organized Supplementary Materials; L.P.M.K., N.A., A.L-E.,625 M.C.H. and E.B-B. prepared figures for the manuscript. All authors read, corrected and commented on626 the manuscript.627 628 Competing interests629 The authors declare no competing financial interests.630 24 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint Figures Figure 1 Phylogenetic, genomic and proteomic comparisons of 20 insect species. From left to right: Phylogenetic tree of 20 insect species withStrigamia maritima (centipede) as outgroup; level of eusociality (one red insect: simple eusociality; two red insects: advanced eusociality; black fly: non-eusocial); fractions of repetitive content (yellow) within genomes of selected species (for sources see supplementary material); proportions of species-specific gene family expansions (green), contractions (red) and stable gene families (black), size of pies represents relative size of gene family change (based on total numbers). Bar chart showing protein orthology across taxonomic groups within each genome. Figure 2 Comparison of developmental pathways between B. germanica , the lower termites, Z. nevadensis and C. secundus , and the higher termite, M. natalensis . Shown from left to right are: a simple phylogeny 97 describing important novelties along the evolution- ary trajectory to termites (numbers in brackets are genome sizes); life cycles; differential expression (log2FoldChange> 1 & p < 0.05) between workers and queens (between nymphs and adult females in B. germanica ) of selected gene families (Desat = desaturases, Elong = elongases, H’ween = Halloween genes) and total numbers within all genes; numbers denote total numbers of genes in each gene family. Figure 3 Expansions, contractions and positive selection within IRs and ORs in termites. a. IR and b. OR gene trees of 13 insect species. In each tree only well supported clades (support values> 85) that include B. germanica or termite genes are highlighted within the gene trees. Lengths of coloured bars represent number of genes per species within each of these clades. Red asterisk in a b. denotes putative root of intronless IRs. c. The upper cartoon depicts the 2D structure of an IR, containing ligand binding lobes (S1 & S2), transmembrane regions (TM1-3) and the pore domain (P). Below, the sequence of the domains along the peptide is represented, showing that the sites, which are under significant positive selection (red bars; codeml site models 7 & 8) within Blattodea-IRs for M. natalensis (p < 1.7x10−10) are all situated within the ligand binding lobes and on or around the putative ligand binding sites (asterisks).86 d. The same representation for ORs, which include 8 transmembrane regions. Positive selection was found for M. natalensis (p = 1.1x10−11) and C. secundus (p = 5.6x10−16) of the orange clade, each at two codon positions within the second transmembrane region and at a third position within 25 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint the C-terminal extra-cellular region for M. natalensis . Figure 4 CpGo/e of seven hemimetabolous insects. a. PCA of predicted DNA methylation patterns among 2664 1-to-1 orthologs, estimated via CpG o/e. Spheres represent positions of species within 3D PCA, with the distance between spheres representing the similarity of CpG o/e between species at each ortholog; curves are distribution of CpG o/e with dotted line showing CpG o/e = 1. b. Tag clouds of enriched (p < 0.05) GO terms (biological processes) among lower (left) and higher quartile (right) of CpG o/e within termites (top) and B. germanica (bottom). For termites, genes were merged from all three species for analysing GO term enrichment. High CpGo/e indicates low level of DNA methylation and vice versa. 26 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint 60.0 lmi aec eda hsa cse rpr ace tca aae nvi ame Pcan cfl d m e bge lhu pba sma sin zne m n a Mnat Csec Znev T cas Aech A cep Dmel Bger Lmig Aaeg Sinv Pbar C flo Hsal Pcan Lhum 0 10000 Number of proteins 5000 Eusociality Repetitive content Gene family expansions Isoptera Blattodea Orthoptera Hymenoptera Insects Diptera Coleoptera Formicidae 0 Mya100200300400500 Aculeata independent origins of eusociality Arthropoda (present in S. maritima) Insecta (present in hemimetabolous insects) Holometabolous (not in hemimetabolous insects) Hymenoptera-specific Isoptera-specific Blattodea-specific Species-specific 15000 20000 25000 30000 Smar: Strigamia maritima; Edan: Ephemera danica; Rpro: Rhodnius prolixus; Nvit: Nasonia vitripennis; Amel: Apis mellifera; Pcan: Polistes canadensis;Hsal: Harpegnathos saltator; Lhum: Linepithema humile; Cflo: Camponotus floridanus; Pbar: Pogonomyrmex barbatus; Sinv: Solenopsis invicta; Aech: Acromyrmex echinatior; Acep: Atta cephalotes; T cas: Tribolium castaneum; Aaeg: Aedes aegypti; Dmel: Drosophila melanogaster; Lmig: Locusta migratoria; Bger: Blattella germanica; Znev: Zootermopsis nevadensis; Csec: Cryptotermes secundus; Mnat: Macrotermes natalensis Estimated divergence times based on (5) and timetree.org Smar Rpro Edan Amel Nvit not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint 15,86618,23616,3207 19 21 6 18 33 6 EcK JHBP 51 51 12 61 ORs 90 B. germanica Z. nevadensis M. natalensis C. secundus reproductive stages reproductive adult 6 nymphal stages primary reproductive dispersing alate soldier larval instars nymphal instars worker nymphal pathway apterous pathway immature stages Termitidae Kalotermitidae Termopsidae Ectobiidae Obligate association with hindgut flagellates, biparental subsociality Eusociality Loss of fat body bacteroids Mantids Cryptocercidae Acquisition of fat body bacteroids primary reproductive dispersing alate neotenic replacement reproductive soldier larval instars nymphal instars egg case egg egg 25% 50% 75% 100% 0% 25% 50% 75% 100% 0% 25% 50% 75% 100% 0% 25% 50% 75% 100% 0% DesatElong CYP4G1IRs ZF-gen es H'ween Differential expression 22 22 1 455 704 7 13 16 1 141 589 20 28 14 14 1 135 819 22 15 3 75 464 nymph- / worker-biased adult female- / queen-biased non differentially expressed 54 C. secundus M. natalensis Z. nevadensis Total 30,130 B. germanica (2.0 Gb) (562 Mb) (1.30 Gb) (1.31 Gb) not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint Csec G13974 T1 Csec G15054 T1 Csec G04826 T1 FBpp0085205 Nasvi2EG001972t1 XM 015983540.1 Znev 18934 Csec G06976 T1 Bger IR 201 Bger IR 657 Csec G07625 T1 Znev 18918 Bger IR 660 XM 015981575.1 Csec G09342 T1 Bger IR 613 Bger IR 589 FBpp0081400 Bger IR 730 Bger IR 145 Mnat 10147 LOCMI01384 Bger IR 355 Csec G05001 T1 Nasvi2EG019393t1 Csec G17293 T1 Nasvi2EG016024t1 FBpp0076857 Bger IR 210 Csec G03269 T1 Bger IR 874 LOCMI15741 XM 015983307.1 Bger IR 594 Csec G12501 T1 Csec G12196 T1 Bger IR 552 Znev 18930 Nasvi2EG016618t1 Bger IR 142 Bger IR 882 Csec G05019 T1 Znev 19067 FBpp0291494 Znev 19050 Bger IR 604 Znev 18904 GB45591-PA Csec G14954 T1 Nasvi2EG010620t1 Bger IR 275 Bger IR 336 Bger IR 255 Bger IR 447 Znev 18808 XM 015979622.1 Bger IR 321 Znev 18962 Mnat 11297 Znev 18999 Bger IR 250 Znev 18977 Bger IR 655 Csec G06137 T1 Bger IR 449 Csec G15113 T1 HSAL22072-PA Bger IR 602 Nasvi2EG016604t1 Bger IR 212 Znev 18809 Bger IR 215 EDAN010593-PA Csec G15053 T1 Nasvi2EG008965t1 Nasvi2EG008880t1 Nasvi2EG008974t1 Bger IR 404 RPRC002248-PA EDAN017398-PA Znev 19017 HSAL22819-PA Csec G04371 T1 Bger IR 939 Nasvi2EG011370t1 Znev 18955 Bger IR 301 Nasvi2EG015274t1 Csec G07053 T1 Znev 18945 Bger IR 209 Bger IR 115 Csec G13366 T1 Bger IR 673 Bger IR 783 Znev 18941 Csec G06519 T1 Nasvi2EG002366t1 FBpp0075333 HSAL22389-PA LOCMI15721 Znev 18989 Bger IR 41a5 Bger IR 615 GB53122-PA FBpp0088541 Mnat 15684 GB49791-PA Bger IR 187 Bger IR 401 Nasvi2EG008635t1 Bger IR 435 RPRC000191-PA Znev 18811 Bger IR 273 Znev 18946 Znev 18968 Bger IR 628 Znev 18884 Mnat 13171 FBpp0076023 PB16964-PA Nasvi2EG005822t1 Mnat 14393 Nasvi2EG016015t1 Bger IR 710 Nasvi2EG010621t1 GB43093-PA Nasvi2EG010587t1 LOCMI15742 GB47387-PA Nasvi2EG008704t1 Mnat 11831 XM 962172.2 Bger IR 141 Bger IR 175 LOCMI16874 Bger IR 249b Csec G04164 T1 Bger IR 678 FBpp0086462 Nasvi2EG005417t1 Nasvi2EG001402t1 Csec G18426 T1 Znev 18826 FBpp0307908 Mnat 14220 Bger IR 372 GB41839-PA Znev 05743 Nasvi2EG002756t1 Nasvi2EG001529t1 Csec G06141 T1 Bger IR 490 Csec G12571 T1 Csec G03444 T1 Bger IR 75o Bger IR 677 Bger IR 535 Mnat 03857 RPRC001816-PA GB49568-PA FBpp0070276 FBpp0288952 Bger IR 359 Mnat 15052 LOCMI17597 Bger IR 583 GB46679-PA Nasvi2EG022737t1 RPRC000247-PA Nasvi2EG001432t1 Znev 19031 Bger IR 116 XM 008199960.2 Znev 18952 RPRC017354-PA Bger IR 197 Znev 18919 Csec G12450 T1 Mnat 13520 HSAL23330-PA Csec G12036 T1 Bger IR 300 FBpp0110195 Nasvi2EG010951t1 Nasvi2EG010457t1 FBpp0307912 Bger IR 702 Bger IR 307 Bger IR 246 Bger IR 617a Bger IR 705 XM 015981165.1 Nasvi2EG004268t1 FBpp0271852 Csec G17653 T1 Csec G01746 T1 Bger IR 41a2 Bger IR 767 Znev 19028 Bger IR 394 Bger IR 75y Nasvi2EG010822t1 Bger IR 666 PB14974-PA GB46886-PA Bger IR 493 Bger IR 517 Bger IR 687 Csec G10299 T1 FBpp0085779 Bger IR 791 HSAL21655-PA Bger IR 421 Mnat 17319 Nasvi2EG015467t1 RPRC000105-PA Bger IR 601 Nasvi2EG008560t1 Csec G04494 T1 Csec G06520 T1 RPRC017346-PA GB49792-PA Csec G10728 T1 Znev 18887 Bger IR 346 Csec G06119 T1 Bger IR 748 RPRC000047-PA Bger IR 780 Csec G16059 T1 Bger IR 322 XM 008195697.2 Bger IR 8a Bger IR 75k FBpp0271713 Znev 05406 Nasvi2EG019411t1 Mnat 09267 Znev 18827 Bger IR 238 Bger IR 236a FBpp0083391 Bger IR 737 LOCMI01052 Znev 01873 Csec G05491 T1 Bger IR 559 Bger IR 525 Znev 05185 Bger IR 749 Csec G12192 T1 Bger IR 720 Znev 18830 Mnat 15587 Csec G13528 T1 Znev 18980 Bger IR 269 Csec G01569 T1 Mnat 01797 Bger IR 877 FBpp0290822 Znev 19058 Bger IR 390 FBpp0076691Csec G00769 T1 Bger IR 549 FBpp0307900 Bger IR 226 RPRC000199-PA HSAL22071-PA Mnat 05832 Bger IR 338 Bger IR 533 Mnat 17378 Znev 18814 Mnat 14299 Mnat 11939 GB46788-PA Bger IR 134 Csec G05697 T1 Bger IR 254 Bger IR 272 RPRC006418-PA Bger IR 167 Mnat 16836 Mnat 03961 FBpp0088259 FBpp0291499 Bger IR 41a16 Bger IR 207 Csec G17413 T1 Nasvi2EG008879t1 Bger IR 41a8 Znev 18937 Bger IR 109 Bger IR 872 Nasvi2EG024926t1 Mnat 17051 Bger IR 715 Bger IR 553 Csec G10366 T1 Znev 18958 Znev 18984 Bger IR 631 PB15237-PA Nasvi2EG009090t1 Bger IR 701 Bger IR 770 Csec G07002 T1 LOCMI15737 Bger IR 524 Znev 18987 RPRC011875-PA Bger IR 199 Znev 18888 Znev 18986 PB19243-PA Bger IR 879 Bger IR 436 Bger IR 106 FBpp0311747 Bger IR 495 Znev 18859 Bger IR 135 Csec G09343 T1 FBpp0071043 Bger IR 567 Bger IR 529 Znev 19007 Bger IR 590 Bger IR 216 Bger IR 531Bger IR 528 FBpp0071044 Bger IR 433 GB40973-PA Nasvi2EG014083t1 Znev 15586 Csec G07394 T1Znev 16401 Csec G16841 T1 FBpp0288947 Znev 19006 Znev 19025 Csec G08155 T1 Znev 05745 Bger IR 580 Bger IR 473 PB15785-PA Mnat 16454 Nasvi2EG010077t1 Bger IR 632 XM 015984212.1 XM 961435.2 Nasvi2EG006240t1 Bger IR 25a Nasvi2EG005798t1 Bger IR 674 Csec G17881 T1 FBpp0304756 Bger IR 766 Bger IR 477 Bger IR 434 XM 963253.3 EDAN014757-PA Csec G08911 T1 RPRC017366-PA Bger IR 308 Nasvi2EG033718t1 Bger IR 41a14 Bger IR 789 Mnat 01740 Bger IR 75v Bger IR 160 Nasvi2EG014499t1 Mnat 17622 XM 015978086.1 Bger IR 489Bger IR 482 Znev 19011 Znev 05107 Bger IR 609 RPRC000021-PA Bger IR 755 Bger IR 146 EDAN005107-PA Mnat 16809 Mnat 06184 LOCMI12015 Bger IR 667 Bger IR 424 Bger IR 513 Nasvi2EG015473t1 RPRC002129-PA FBpp0087185 Nasvi2EG013946t1 Nasvi2EG012213t1 FBpp0087414 HSAL17854-PA Bger IR 218 Csec G12035 T1 Znev 19005 Bger IR 342 Bger IR 727 HSAL23771-PA Csec G17656 T1 Bger IR 883 Csec G10525 T1 Bger IR 271 Bger IR 739 Bger IR 247 Znev 18866 Bger IR 261 Bger IR 516 Nasvi2EG013075t1 XM 015985319.1 EDAN009865-PA Znev 18961 Znev 19045 Csec G09893 T1 Nasvi2EG008461t1 XM 015982078.1 Bger IR 357 Bger IR 349 Csec G04160 T1 Csec G02868 T1 Bger IR 159 RPRC017348-PA Bger IR 408 XM 015978116.1 Csec G01076 T1 Mnat 16142 Bger IR 430 Mnat 10148 Bger IR 591 Bger IR 344 Bger IR 299 Csec G02663 T1 PB17940-PA GB48097-PA Bger IR 214 Bger IRs 313 Bger IR 234 Nasvi2EG001401t1 XM 015980701.1 Csec G13529 T1 Bger IR 627 Bger IR 156 Bger IR 464 Nasvi2EG001379t1 Bger IR 688 LOCMI15728 Mnat 15509 HSAL21454-PA Bger IR 117 Nasvi2EG000707t1 Bger IR 790 FBpp0075758 LOCMI15734 RPRC017345-PA FBpp0307023 Znev 18857 Bger IR 176 Bger IR 661 Nasvi2EG021285t1 Nasvi2EG001380t1 Znev 18833 HSAL16956-PA Znev 19019 Bger IR 488 Nasvi2EG020567t1 Bger IR 460 Bger IR 551 Mnat 15442 XM 015984855.1 Bger IR 445 Bger IR 905 Nasvi2EG008737t1 Bger IR 584 PB19088-PA LOCMI15744 Nasvi2EG011748t1 Bger IR 903 Bger IR 281 Bger IR 398 Bger IR 41a3 EDAN007271-PA Csec G01238 T1 Znev 19057 Csec G10647 T1 FBpp0288956 XM 008197827.2 Bger IR 222 Nasvi2EG001475t1 Bger IR 565 Nasvi2EG012860t1 Csec G05002 T1 Bger IR 140 FBpp0110206 Csec G04165 T1 PB19089-PA Bger IR 318 HSAL13538-PA Nasvi2EG009821t1 Bger IR 41a4 Bger IR 262 Bger IR 508 Csec G11401 T1Znev 18861 Mnat 03466 Bger IR 707 Znev 18957 Bger IR 680 Znev 18854 Znev 18950 Znev 19018 Nasvi2EG020582t1 Mnat 03467 Znev 19020 Znev 18886 Znev 19053 Bger IR 734 Csec G13364 T1 Bger IR 669 XM 015977838.1 LOCMI15743 Nasvi2EG006460t1 Bger IR 463 GB51842-PA Bger IR 462 Bger IR 764 Bger IR 375 HSAL14200-PA Bger IR 907 Bger IR 768 Bger IR 675 Nasvi2EG016023t1 Bger IR 492 Bger IR 603 Nasvi2EG004852t1 Bger IR 75a Bger IR 139 Bger IR 784 XM 015981164.1 EDAN013514-PA Bger IR 558 RPRC017356-PA Bger IR 902 Bger IR 75h Nasvi2EG008454t1 XM 008195137.2 Znev 18933 PB19241-PA GB47549-PA Bger IR 586 Bger IR 451 Bger IR 595 Csec G05021 T1 Bger IR 544 Bger IR 663 RPRC017355-PA Bger IR 227 Csec G11294 T1 Nasvi2EG004269t1 Bger IR 230 Csec G15051 T1 FBpp0076909 Bger IR 402 Bger IR 884 EDAN019029-PA Bger IR 582 EDAN010594-PA Bger IR 871 Nasvi2EG019905t1 Znev 12219 FBpp0087171 RPRC000314-PA Bger IR 205 Csec G16535 T1 Bger IR 331 Bger IR 380 Bger IR 337 XM 008201422.2 Nasvi2EG010177t1 Nasvi2EG012232t1 Bger IR 480 XM 963693.4 Bger IR 75x RPRC000296-PA Csec G06973 T1 RPRC002040-PA Bger IR 440 XM 015981167.1 Bger IR 324 XM 008202574.2 Nasvi2EG000021t1 Bger IR 794 Bger IR 154 Csec G03196 T1 Bger IR 75b Bger IR 868 Csec G00893 T1 Bger IR 129 Bger IR 341 Mnat 16835 Bger IR 557 EDAN002626-PA GB54881-PA Bger IR 268 Bger IR 736 Nasvi2EG028333t1 Bger IR 592 XM 969840.3 HSAL11608-PA Nasvi2EG016492t1 Nasvi2EG013884t1 Znev 18940 Znev 18942 LOCMI15740 Znev 18956 Mnat 05963 Znev 19042 Znev 18967 XM 008192384.2 LOCMI15735 Bger IR 611 Mnat 16742 GB55622-PA LOCMI15724 Bger IR 377 Bger IR 593 Mnat 06952 Bger IR 427 Bger IR 113 HSAL21026-PA Bger IR 511 Nasvi2EG008878t1 Znev 18844 Bger IR 263 Nasvi2EG006241t1 Mnat 01722 LOCMI08219 Bger IR 635 Nasvi2EG009428t1 Csec G07003 T1 FBpp0307907 Bger IR 694 Bger IR 103 Csec G10374 T1 LOCMI17612 Znev 19003 Csec G11400 T1 RPRC003290-PA PB25135-PA Csec G06118 T5 Znev 19054 FBpp0307897 Bger IR 208 LOCMI17590 Bger IR 785 Znev 18917 Znev 18985 Mnat 14395 Bger IR 738 LOCMI17599 Znev 18939 Nasvi2EG006322t1 Mnat 01423 Mnat 11725 Mnat 10662 Mnat 10144 Csec G03316 T1 Mnat 15685 RPRC013097-PA Bger IR 41a10 Csec G08335 T1 Bger IR 880 FBpp0288955 Znev 18960 Bger IR 700 Znev 18976 Bger IR 912 Bger IR 285 Znev 18829 Znev 04642 Bger IR 709 Bger IR 41a12 Znev 18963 Mnat 03468 HSAL22970-PA Mnat 11520 Bger IR 679 Znev 19051 HSAL13539-PA EDAN016924-PA HSAL17231-PA FBpp0307909 Bger IR 683 XM 008203439.2 Bger IR 786 Bger IR 345 Bger IR 143 Bger IR 703 Znev 18889 Bger IR 448 FBpp0271810 Znev 18813 Nasvi2EG016576t1 Csec G18356 T1 Mnat 16065 Bger IR 235 Mnat 16834 Bger IR 225 Nasvi2EG009822t1 Nasvi2EG015304t1 Csec G04078 T1 Bger IR 157 PB26335-PA FBpp0288950 Bger IR 654 Nasvi2EG021930t1 HSAL21707-PA FBpp0085662 Bger IR 361 GB49273-PA Bger IR 388 Csec G03325 T1 PB13586-PA Bger IR 668 Bger IR 442 Znev 19037 Bger IR 132 PB19078-PA Bger IR 664 Bger IR 545 Znev 18923 Nasvi2EG008552t1 Csec G02892 T1 Bger IR 173 Bger IR 876 Bger IR 253 Bger IR 236c Nasvi2EG006175t1 Bger IR 471 Mnat 16347 Bger IR 397 Znev 18806 Bger IR 431 LOCMI15738 Mnat 16301 Bger IR 381 Csec G10648 T1 FBpp0086025 Csec G05038 T1 Bger IR 303 Bger IR 202 LOCMI15723 Csec G01962 T4 Nasvi2EG012080t1 Znev 18819 Bger IR 740 Csec G04166 T1 RPRC007037-PA Nasvi2EG008450t1 Bger IR 600 Mnat 06321 Bger IR 732 LOCMI15727 Bger IR 719 Znev 18817 Bger IR 510 LOCMI15739 HSAL22388-PA Bger IR 548 RPRC007206-PA Bger IR 348 Bger IR 163 Bger IR 670 Mnat 17304 Nasvi2EG008565t1 Mnat 14298 GB49268-PA Bger IR 570 XM 015978316.1 GB50006-PA Csec G10053 T1 Nasvi2EG012014t1 RPRC017347-PA Mnat 10145 Bger IR 507 Bger IR 107 FBpp0083346 Bger IR 118 Bger IR 105 Bger IR 693 Bger IR 878 Nasvi2EG006236t1 Csec G03872 T2 Bger IR 577 Znev 18843 LOCMI17587 PB26334-PA Nasvi2EG001530t1 Bger IR 399 Bger IR 910 Bger IR 578 FBpp0112397 Bger IR 334 Bger IR 746 Nasvi2EG005921t1 Bger IR 566 Znev 05744Csec G06117 T1 HSAL19189-PA Bger IR 177 PB14973-PA FBpp0071032 Bger IR 569 XM 015978324.1 Bger IR 203 Bger IR 481 Csec G10241 T1 Znev 18800 Bger IR 193 FBpp0085686 Znev 18895 Znev 18935 FBpp0307898Nasvi2EG013074t1 RPRC000538-PA Bger IR 353 Csec G06079 T1 Bger IR 41a7 Nasvi2EG007394t1 GB45776-PA Znev 15585 LOCMI15722 Bger IR 295 Bger IR 217 Znev 18944 RPRC017353-PA Bger IR 634 FBpp0307910 Bger IR 123 Nasvi2EG013077t1 Znev 18920 Csec G02869 T1 Bger IR 101 Znev 19066 Bger IR 665 Bger IR 788 GB53118-PA Nasvi2EG004232t1 Nasvi2EG004983t1 RPRC000846-PA Nasvi2EG008394t1 Csec G05000 T1 Znev 18847 FBpp0307896 HSAL13924-PA Bger IR 479 Znev 18862 Csec G04162 T1 Nasvi2EG008455t1 XM 015984531.1 Csec G04606 T1 Csec G15923 T1 Csec G05696 T1 RPRC000589-PA Nasvi2EG018970t1 Znev 18825 PB12162-PA PB19619-PA Bger IR 174 Bger IR 186 Znev 18828 PB15313-PA Bger IR 690 Znev 19060 Nasvi2EG012568t1 Bger IR 423 Bger IR 124 Csec G07528 T1 LOCMI15732 Bger IR 277 Bger IR 378 Bger IR 629 Bger IR 725 Bger IR 396 Bger IR 75d PB17660-PA GB54996-PA Bger IR 429 Nasvi2EG009427t1 Bger IR 607 FBpp0071268 Znev 18820 Bger IR 692Bger IR 684 Bger IR 698 XM 008201710.2 Nasvi2EG016025t1 Nasvi2EG008453t1 Bger IR 717 LOCMI15736 XM 962344.2 Znev 18949 Znev 18883 Mnat 17582 FBpp0271710 Znev 18936 Mnat 08227 Mnat 12825 Nasvi2EG002289t1 Znev 18812 Mnat 15443 HSAL14199-PA Mnat 17415 Csec G07680 T1 Bger IR 148 FBpp0110134 Csec G03135 T1 Mnat 15415 Bger IR 392 Csec G17654 T1 Bger IR 588 PB26185-PA LOCMI15733 Nasvi2EG000002t1 Bger IR 651 Csec G11515 T1 Mnat 17932 Bger IR 617b EDAN009612-PA Bger IR 75j Csec G12191 T1 Znev 18832 FBpp0288951 Bger IR 335 Bger IR 457 Bger IR 741 Csec G03443 T1 EDAN004513-PA RPRC001831-PA XM 015981166.1 Bger IR 485 Bger IR 671 Bger IR 626 Bger IR 556 Bger IR 274 Csec G10365 T1 Mnat 11612 Bger IR 376 Bger IR 41a13 Bger IR 249a FBpp0307916 Mnat 16066 Bger IR 259 Mnat 03858 Bger IR 662 Nasvi2EG001532t1 PB25133-PA Znev 18885 Nasvi2EG001534t1 Csec G05006 T1 FBpp0301551 Bger IR 456 Bger IR 119 Csec G03278 T1 Bger IR 400 Nasvi2EG000567t1 FBpp0076022 Znev 01874 FBpp0083591 Bger IR 108 GB49275-PA Csec G02881 T1 XM 015978130.1XM 015981872.1 FBpp0078739 Bger IR 676 Bger IR 369 Bger IR 200 LOCMI12203 RPRC000489-PA Bger IR 504 Csec G13527 T1 EDAN017002-PA HSAL13086-PA Bger IR 282 Znev 19030 Bger IR 514 Znev 18831 Csec G08686 T1 Bger IR 368 Znev 18974 Bger IR 561 Bger IR 329 Bger IR 546 Znev 18938 Bger IR 41a11 XM 015983852.1 PB21237-PA Csec G16315 T1 XM 015981272.1 FBpp0271858 Csec G04495 T1 Bger IR 735 Bger IR 310 Bger IR 360 Bger IR 221 XM 008200235.2 Nasvi2EG005140t1 Nasvi2EG015255t1 RPRC000154-PA Csec G15055 T1 Bger IR 219 Mnat 17086 Bger IR 223 Znev 18834 Bger IR 432 Bger IR 164 PB26743-PA HSAL12158-PA Bger IR 506 Mnat 17002 Bger IR 656 Csec G08611 T1 Bger IR 260 RPRC010372-PA Nasvi2EG010019t1 Csec G13524 T1 Csec G08612 T1 Bger IR 518 Bger IR 630 Nasvi2EG013887t1 Csec G13188 T1 Bger IR 509 Mnat 16465 Bger IR 579 Nasvi2EG008452t1 Bger IR 599 Nasvi2EG008556t1 Nasvi2EG006009t1 Csec G17212 T1 Csec G12037 T1 Bger IR 104 PB12653-PA Csec G00980 T1 Csec G04163 T1 Csec G04558 T1 PB25136-PA Bger IR 211 FBpp0291498 Mnat 16709 Mnat 06646 Bger IR 873 Bger IR 41a1 Nasvi2EG020725t1 Bger IR 41a9 Znev 18856 Mnat 06322Bger IR 204 Nasvi2EG010011t1 Csec G01810 T1 Csec G02897 T1 Mnat 17418 GB55621-PA LOCMI15725 Bger IR 494 Mnat 05833 Znev 18810 Bger IR 236b EDAN004012-PA Znev 18853 Bger IR 581 Bger IR 379 Znev 18845 Znev 18835 Mnat 09716 Znev 19052 Bger IR 354 RPRC017352-PA Csec G16618 T1 Znev 18943 Bger IR 623 XM 015978094.1 RPRC002124-PA EDAN013654-PA FBpp0086497 Csec G15840 T1 Znev 19014 Bger IR 714 PB17941-PA Bger IR 779 Bger IR 452 Znev 18867 Mnat 03323 FBpp0271900 Csec G14083 T5 Bger IR 351 RPRC000007-PA PB19090-PA EDAN014957-PA Nasvi2EG016575t1 FBpp0307906 Bger IR 41a6 Bger IR 75m Csec G00983 T1 Bger IR 554 Bger IR 75s LOCMI15729 RPRC002112-PA Csec G11113 T1 Csec G09569 T1 Nasvi2EG013073t1 Csec G04254 T1 RPRC002996-PA PB19332-PA Bger IR 875 Bger IR 198 Bger IR 251 RPRC000379-PA Bger IR 298 Mnat 10839 Znev 18990 Nasvi2EG008566t1 Znev 01639 Bger IR 706 Csec G04161 T1 LOCMI15730 FBpp0288732 Bger IR 576 LOCMI01953 Bger IR 450 FBpp0099386 Znev 18922 Znev 18915 Csec G10921 T1 Znev 07227 Bger IR 574 Nasvi2EG008534t1 FBpp0303373 Csec G10051 T1 Znev 07836 Bger IR 585 Nasvi2EG001531t1 Csec G10524 T1 Bger IR 206 Bger IR 689Bger IR 691 PB21526-PA FBpp0078410 Znev 19041 Bger IR 606 Mnat 15440Bger IR 195 Bger IR 352 HSAL17272-PA Znev 19032 Bger IR 652 Znev 19055 Mnat 15414 Bger IR 383 Bger IR 596 Nasvi2EG021904t1 RPRC017357-PA Nasvi2EG013076t1 Znev 19004 Znev 16376 Nasvi2EG012685t1 Bger IR 881 Csec G02996 T1 FBpp0085661 Mnat 16741 Csec (64) Znev (48) Bger (176) Mnat (23) Bger (161) T ermites (21) Bger (13) T cas (12) Dmel (41) Nvit (72) Others (17) T ermites (20) Bger (23) Others (24) Mnat (25) Csec (34) Znev (44) Bger (36) Others (21) Csec (9) Znev (20)Bger (13)Rpro (16)T cas (12) Nvit (14) Others (29) Blattodea IRs Cockroach IRs Mnat (9)Group A Group B Grp C Group D iGluRs, IR8a/IR25a b Nasvi2EG037005t1 RPRC002152-PA XM 015978292.1 Nasvi2EG036934t1 Nasvi2EG036954t1 Csec G13562 T1 RPRC001952-PA Bger OR 40 Bger OR 17 Nasvi2EG036928t1 GB52398-PA HSAL13437-PA Nasvi2EG037237t1 Csec G08666 T1 XM 015985262.1 XM 015985328.1 GB40195-PA LOCMI17440 XM 015982987.1 GB46831-PA Nasvi2EG036984t1 GB43026-PA LOCMI17617 GB50051-PA Csec G06757 T1 PB18679-PA GB52369-PA Bger OR 27 Nasvi2EG037214t1 LOCMI17369 XM 015979619.1 GB50001-PA GB52365-PA GB52414-PA HSAL20024-PA LOCMI17458 Nasvi2EG037240t1 Bger OR 41 XM 008199721.2 Nasvi2EG037242t1 Bger OR 58 GB47987-PA HSAL20021-PA Bger OR 57A LOCMI17603 LOCMI17372 Csec G03246 T1 LOCMI17359 HSAL23785-PA Znev 19027 RPRC000055-PA HSAL13738-PA Nasvi2EG036913t1 PB18756-PA Bger OR 50E LOCMI17619 GB47983-PA HSAL12628-PA Nasvi2EG037292t1 GB52402-PA Nasvi2EG037269t1 Bger OR 31 LOCMI17607 HSAL20059-PA LOCMI17444 Nasvi2EG037186t1 PB27050-PA Nasvi2EG037024t1 Znev 18971 LOCMI17433 PB26537-PA Csec G08627 T1 Nasvi2EG037103t1 PB26324-PA PB26465-PA RPRC000330-PA HSAL16293-PA Nasvi2EG037087t1 Nasvi2EG037167t1 Bger OR 75 HSAL13785-PA RPRC001309-PA LOCMI17380 HSAL16976-PA XM 015977545.1 HSAL13786-PA PB26511-PA RPRC000433-PA Nasvi2EG037229t1 Nasvi2EG037118t1 Znev 18982 GB52404-PA XM 015983215.1 GB44713-PA GB50053-PA Nasvi2EG002474t1 PB14764-PA HSAL13433-PA XM 015983957.1 XM 008200573.2 LOCMI17600 FBpp0083295 GB42602-PA GB50055-PA Nasvi2EG037166t1 Nasvi2EG037187t1 PB13702-PA HSAL13723-PA LOCMI17465 HSAL13726-PA Csec G10644 T1 Bger OR 1 Nasvi2EG037185t1 HSAL10294-PA Nasvi2EG019676t1 Znev 19056 HSAL13436-PA XM 001814810.2 RPRC000619-PA GB42590-PA Nasvi2EG037081t1 XM 015982612.1 Nasvi2EG037213t1 GB52379-PA FBpp0070069 PB15728-PA Nasvi2EG037239t1 HSAL13438-PA HSAL11771-PA Bger OR 71 PB10210-PA RPRC002156-PA PB23952-PA RPRC000530-PA PB26202-PA GB55926-PA HSAL16292-PA Nasvi2EG037021t1 Nasvi2EG001643t1 GB42646-PA Nasvi2EG037172t1 LOCMI17381 XM 015978954.1 Nasvi2EG037043t1 RPRC000555-PA GB47123-PA XM 961697.2 PB26123-PA PB18758-PARPRC002142-PA XM 015982874.1 RPRC000526-PA XM 008202761.2 Nasvi2EG037137t1 GB45832-PA Nasvi2EG002489t1 LOCMI17601 Bger OR 51B RPRC000301-PA LOCMI17363 HSAL21172-PA LOCMI17591 Nasvi2EG015554t1 PB27140-PA LOCMI17462 FBpp0081361 LOCMI17621 Bger OR 52 Bger OR 9 Nasvi2EG037136t1 HSAL13730-PA Nasvi2EG037278t1 Bger OR 63 Nasvi2EG037090t1 PB26275-PA Znev 19033 HSAL13776-PA Bger OR 66 LOCMI17395 FBpp0086894 LOCMI17407 RPRC002122-PA GB52393-PA PB26730-PA LOCMI17429 XM 015977553.1 Bger OR 19 HSAL13736-PA RPRC000547-PA HSAL14207-PA Nasvi2EG037082t1 XM 015980754.1 HSAL10994-PA Bger OR 85 GB40192-PA Nasvi2EG037201t1 Nasvi2EG037117t1XM 008201092.2 GB52370-PA Nasvi2EG037048t1 Nasvi2EG037099t1 XM 015979453.1 PB26835-PA Nasvi2EG008758t1 Nasvi2EG037353t1 PB11614-PA PB26462-PA GB51178-PA GB47988-PA LOCMI17463 HSAL13637-PA GB52376-PA GB53347-PA Csec G09268 T1 Bger OR 61 XM 015983848.1 GB52388-PA Nasvi2EG037077t1 LOCMI17439 XM 015983100.1 Nasvi2EG000535t1 Nasvi2EG004440t1 Nasvi2EG037236t1 LOCMI17446 Bger OR 54 Znev 18871 GB42588-PA RPRC000456-PA RPRC000059-PA HSAL13431-PA Nasvi2EG037275t1 PB26573-PA FBpp0076714 FBpp0071929 PB17573-PA Bger OR 51A HSAL18041-PA HSAL20023-PA HSAL23021-PA PB26469-PA Nasvi2EG037092t1 XM 015982467.1 Znev 18981 HSAL16499-PA Nasvi2EG037215t1 PB26472-PA Nasvi2EG037115t1 FBpp0089408 GB40667-PA Nasvi2EG037039t1 PB10232-PA Nasvi2EG014222t1 HSAL12626-PA HSAL13442-PA Nasvi2EG037197t1 RPRC001799-PA FBpp0075008 XM 015984411.1 Znev 18983 FBpp0085645 Nasvi2EG037183t1 RPRC000249-PA Bger OR 72 Znev 19008 Csec G16524 T1 HSAL23579-PA XM 015984062.1 Znev 18870 PB15434-PA FBpp0087962 GB40189-PA HSAL23577-PA Nasvi2EG037203t1 XM 008196471.2 PB21571-PA Znev 07294 HSAL10466-PA LOCMI17385 PB26616-PA PB14128-PA Nasvi2EG036985t1 Bger OR 97 XM 015977456.1 Bger OR 112 Nasvi2EG037028t1 XM 008201568.2 LOCMI17392 Nasvi2EG004495t1 GB45081-PA Nasvi2EG037116t1 Nasvi2EG037246t1 Nasvi2EG037019t1 LOCMI17414 GB41599-PA GB40198-PA Nasvi2EG036920t1 PB26510-PA XM 015984060.1 HSAL13724-PA XM 015982565.1 Nasvi2EG037223t1 LOCMI17379 PB20696-PA Nasvi2EG001640t1 XM 015982984.1 PB21574-PA FBpp0078441 LOCMI17411 Nasvi2EG037031t1 PB27142-PA PB11549-PA PB16912-PA RPRC001980-PA Csec G09264 T1 PB26617-PA Nasvi2EG036955t1 RPRC000125-PA PB14301-PA PB26836-PA Csec G09265 T1 XM 008202113.2 PB11226-PA HSAL10814-PA LOCMI17391 RPRC000522-PA HSAL10518-PA Csec G03324 T1 LOCMI17375 Nasvi2EG037038t1 XM 015983878.1 HSAL17572-PA Bger OR 5 XM 015980190.1 LOCMI17412 XM 015982429.1 Znev 18972 Bger OR 55D Nasvi2EG037164t1 XM 015982634.1 RPRC000371-PA RPRC000468-PA LOCMI17384 PB26262-PA PB21573-PA Bger OR 35 Nasvi2EG037100t1 HSAL23581-PA RPRC000188-PA Nasvi2EG037003t1 RPRC000507-PA Bger OR 12 XM 015984150.1 Nasvi2EG037042t1 Nasvi2EG036927t1 HSAL11024-PA Bger OR 24 Csec G08633 T1 PB26766-PA Nasvi2EG037280t1 Nasvi2EG037129t1 HSAL13308-PA HSAL18851-PA GB52395-PA GB40664-PA Nasvi2EG037147t1 Znev 05616 Znev 18970 Znev 19012 Znev 18823 LOCMI17629 RPRC000431-PA Nasvi2EG000297t1 Nasvi2EG036923t1 PB27139-PA RPRC000452-PA GB40669-PA GB40202-PA Nasvi2EG037120t1 LOCMI17387 RPRC000601-PA PB11556-PA LOCMI17361 GB46573-PA XM 015982988.1 XM 015981387.1 LOCMI17394 GB52363-PA PB26759-PA Nasvi2EG037222t1 PB10658-PA HSAL13727-PA HSAL12586-PA Znev 19034 GB40196-PA FBpp0085388 XM 015984390.1 XM 015982628.1 XM 015982432.1 LOCMI17457 PB21817-PA LOCMI17390 XM 015981021.1 Csec G10479 T1 XM 015984151.1 Nasvi2EG037097t1 Bger OR 102 GB44714-PA XM 008199722.2 Nasvi2EG015558t1 XM 015984400.1 PB11389-PA Bger OR 119 Znev 18841 Znev 18948 Znev 19036 HSAL16500-PA RPRC000222-PA HSAL15952-PA Csec G01467 T1 XM 015980763.1 Bger OR 56 PB21432-PA Znev 19064 HSAL20025-PA RPRC000132-PA Nasvi2EG036929t1 PB26246-PA GB40186-PA XM 015984379.1 Nasvi2EG037225t1 XM 008192262.2 Csec G08634 T1 GB40191-PA PB18755-PA LOCMI17423 PB16375-PA Bger OR 39 FBpp0077499 HSAL13782-PA Nasvi2EG019678t1 PB14765-PA RPRC000375-PA XM 015984754.1 PB11555-PA Znev 18979 GB47122-PA Nasvi2EG002490t1 LOCMI17430 Znev 18852 HSAL15957-PA FBpp0071396 PB27143-PA XM 008192407.2 RPRC001623-PA PB26125-PA RPRC001154-PA Csec G17935 T1 Znev 18908 Nasvi2EG037283t1 Bger OR 67 Nasvi2EG015518t1 Mnat 17263 Nasvi2EG004443t1 Nasvi2EG036957t1 HSAL13440-PA LOCMI17618 GB50056-PA Nasvi2EG037184t1 RPRC000209-PA GB52394-PA LOCMI17404 Nasvi2EG036997t1 RPRC000331-PA RPRC001292-PA Csec G09267 T1 LOCMI17403 Znev 19013 LOCMI17595 PB23683-PA HSAL13781-PA PB23958-PA PB13489-PA RPRC003171-PA Nasvi2EG006432t1 Nasvi2EG037158t1 HSAL21281-PA FBpp0083735 Csec G11043 T1 Nasvi2EG014919t1 Csec G09183 T1 GB47984-PA Nasvi2EG037095t1 PB26468-PA PB26689-PA Nasvi2EG036975t1 GB52406-PA Nasvi2EG002476t1 XM 008198935.2 RPRC000029-PA LOCMI17622 Nasvi2EG007471t1 RPRC002113-PA PB11554-PA LOCMI17438 FBpp0087029 Nasvi2EG037085t1 RPRC002157-PA XM 015982566.1 Nasvi2EG037098t1 Mnat 14693 HSAL11730-PA XM 008194185.2 PB21572-PA Nasvi2EG037216t1 GB52413-PA LOCMI17365 XM 008201570.2 Nasvi2EG002321t1 LOCMI17399 XM 015978838.1 PB11562-PA XM 015984407.1 GB51945-PA Nasvi2EG037102t1 Mnat 01906 PB25408-PA RPRC009806-PA HSAL19347-PA Nasvi2EG036924t1 Nasvi2EG037020t1 GB52384-PA XM 015978854.1 Nasvi2EG036978t1 Bger OR 109 FBpp0079864 PB16376-PA GB41316-PA Nasvi2EG037127t1 RPRC000201-PA XM 008196899.2 Znev 19062 Nasvi2EG037040t1 LOCMI17594 Nasvi2EG037080t1 Csec G03486 T1 Nasvi2EG036958t1 LOCMI17371 GB52383-PA LOCMI17432 Nasvi2EG037023t1 Nasvi2EG000529t1 PB26471-PA Nasvi2EG037180t1 GB51075-PA Bger OR 69 Mnat 10995 Nasvi2EG037000t1 XM 015983472.1 Csec G16046 T1 Nasvi2EG037271t1 XM 015980145.1 LOCMI17366 XM 008200490.2 HSAL13728-PA HSAL17874-PA Nasvi2EG037148t1 LOCMI13114 Nasvi2EG037219t1 GB47121-PA GB52385-PA HSAL19345-PA Bger OR 53 Nasvi2EG037073t1 PB12530-PA HSAL14203-PA PB12531-PA XM 015977667.1 RPRC000579-PA Nasvi2EG037016t1 Znev 18978 GB40200-PA HSAL14209-PA Nasvi2EG011547t1 Nasvi2EG037272t1 GB52403-PA HSAL13326-PA RPRC000318-PA Nasvi2EG000533t1 FBpp0081326 HSAL13778-PA Bger OR 20 XM 015982983.1 GB52372-PA PB25409-PA LOCMI17376 Nasvi2EG014912t1 LOCMI17461 Nasvi2EG037113t1 XM 015983504.1 GB52407-PA FBpp0071932 XM 015984603.1 GB47985-PA Nasvi2EG037241t1 PB26690-PA FBpp0076149 Csec G06758 T1 Csec G10480 T1 LOCMI17383 XM 015983471.1 Bger OR 110 FBpp0301983 HSAL13741-PA Bger OR 103 FBpp0085387 FBpp0087262 Csec G08632 T1 Znev 18838 GB40197-PA PB10374-PA Znev 19068 RPRC000065-PA RPRC000053-PA PB27135-PA PB11553-PA Bger OR CO GB52408-PA PB26467-PA Csec G09266 T1 GB55277-PA FBpp0075412 Nasvi2EG037074t1 XM 015980768.1 XM 015984051.1 FBpp0070002 Nasvi2EG037321t1 Nasvi2EG037274t1 PB15431-PA Nasvi2EG015563t1 LOCMI17398 RPRC000616-PA GB52416-PA LOCMI17443 PB24602-PA GB52410-PA HSAL16501-PA PB27141-PA LOCMI17425 Bger OR 55C FBpp0076104 HSAL13636-PA Nasvi2EG037084t1 HSAL23578-PA Nasvi2EG036947t1 PB26838-PA PB21570-PA PB11546-PA PB26691-PA Nasvi2EG037030t1 Nasvi2EG037287t1 HSAL13638-PA Znev 18890 Nasvi2EG008578t1 XM 015978914.1 LOCMI17449 GB40666-PA HSAL14204-PA Bger OR 49 GB50825-PA LOCMI17614 Nasvi2EG037248t1 PB26928-PA FBpp0076026RPRC000441-PA RPRC004587-PA Csec G09263 T1 HSAL12934-PA Nasvi2EG037144t1 PB10233-PA PB26507-PA Csec G08637 T1 HSAL13787-PA XM 015983844.1 Bger OR 95 Nasvi2EG000853t1 EDAN009192-PA Nasvi2EG036932t1 RPRC002105-PA Znev 18836 Nasvi2EG037284t1 Bger OR 114 XM 015982184.1 XM 008196894.2 Bger OR 3 HSAL13441-PA LOCMI17464 Nasvi2EG012962t1 PB26458-PA GB45080-PA XM 015982193.1 PB21568-PA FBpp0288403 RPRC002136-PA LOCMI17627 HSAL10727-PA XM 015983858.1 GB40194-PA Bger OR 86 XM 015982649.1 XM 015977457.1 XM 015978574.1 Nasvi2EG036973t1 PB10096-PA PB26252-PA GB40190-PA RPRC006570-PA Nasvi2EG014910t1 XM 015983053.1 Znev 19009 Znev 18891 Nasvi2EG036983t1 GB55923-PA GB48704-PA GB40668-PA Znev 18947 GB44708-PA LOCMI17382 Bger OR 77 LOCMI17436 FBpp0077382 XM 015985329.1 LOCMI17452 HSAL23132-PA Nasvi2EG037094t1 LOCMI17596 Nasvi2EG008757t1 RPRC000490-PA HSAL12254-PA HSAL21282-PA FBpp0083734 Nasvi2EG037075t1 Znev 18872 PB23898-PA Nasvi2EG037076t1 LOCMI17427 HSAL13737-PA Csec G16797 T1 RPRC000083-PA PB11390-PA PB26466-PA PB26688-PA Bger OR 37 LOCMI17389 PB27138-PA Nasvi2EG037026t1 HSAL16296-PA PB23956-PA HSAL11023-PA Nasvi2EG037146t1 HSAL13731-PA Mnat 16988 XM 015983102.1 Bger OR 65 LOCMI17358 Nasvi2EG037018t1 HSAL13443-PA Nasvi2EG037200t1 PB14129-PA Nasvi2EG037299t1 PB26508-PA Nasvi2EG037022t1 Csec G08623 T1 Nasvi2EG036971t1 Znev 18839 XM 015984160.1 RPRC000136-PA RPRC000149-PA XM 015984938.1 Csec G04511 T1 GB40185-PA HSAL11747-PA HSAL13435-PA GB52361-PA XM 015979745.1 HSAL13733-PA HSAL16482-PA FBpp0081506 Mnat 13517 Csec G08628 T1 LOCMI17397 GB52377-PA Bger OR 74 XM 015979321.1 RPRC000041-PA RPRC001928-PA Nasvi2EG014911t1 Csec G10643 T1 PB19437-PA PB10429-PA Bger OR 87 Nasvi2EG036940t1 LOCMI17447 LOCMI17420 PB26452-PA LOCMI17630 XM 015983074.1 Bger OR 106 Bger OR 36 PB27067-PA Bger OR 91 FBpp0077542 GB52397-PA Nasvi2EG017096t1 Nasvi2EG037044t1 HSAL14205-PA Bger OR 14 Bger OR 6 GB40665-PA XM 015982986.1 GB52389-PA HSAL15954-PA Mnat 14525 HSAL11901-PA GB52396-PA Nasvi2EG017811t1 FBpp0079371 Bger OR 123 LOCMI05251 Znev 18824 RPRC000255-PA Bger OR 88 Nasvi2EG037112t1Nasvi2EG037301t1 PB16372-PA Nasvi2EG016470t1 HSAL20022-PA LOCMI17368 FBpp0078616 Nasvi2EG037286t1 Nasvi2EG037128t1 XM 015985424.1 RPRC000146-PA Nasvi2EG014927t1 Znev 19010 HSAL15958-PA GB52371-PA HSAL13788-PA PB10121-PA RPRC000202-PA FBpp0079863 Nasvi2EG037078t1 Csec G07181 T1 HSAL13740-PA HSAL17155-PA Nasvi2EG036926t1 GB52386-PA XM 008192097.2 XM 015977390.1 Nasvi2EG036925t1 FBpp0071930 Znev 18907 LOCMI17431 GB52390-PA Nasvi2EG000854t1 XM 015984780.1 GB53346-PA RPRC000165-PA LOCMI17589 LOCMI17362 PB22904-PA RPRC001767-PA LOCMI17405 FBpp0089412 FBpp0077541 GB48699-PA HSAL14208-PA GB50054-PA GB40184-PA HSAL12097-PA GB52381-PA Csec G10920 T1 HSAL14464-PA Bger OR 29 PB11559-PA GB52366-PA PB26509-PA HSAL13780-PA Mnat 09880 RPRC000376-PA RPRC001291-PA Bger OR 59 XM 015979726.1 XM 015983597.1 Nasvi2EG006530t1 PB13495-PA Nasvi2EG037027t1 GB52368-PA XM 015985440.1 LOCMI17401 Mnat 16347 Nasvi2EG037001t1 PB26460-PA Nasvi2EG037210t1 Nasvi2EG037114t1 PB11548-PA Nasvi2EG001630t1 FBpp0312615 GB47952-PA XM 015982625.1 PB26927-PA Znev 18873 HSAL10728-PA Nasvi2EG000541t1 XM 015984064.1 PB26474-PA Znev 18855 PB26475-PA LOCMI17453 PB16982-PA LOCMI17448 PB26464-PA Nasvi2EG037224t1 Nasvi2EG012961t1 Nasvi2EG036912t1 GB52378-PA PB25406-PA LOCMI17611 Csec G03321 T1 PB14770-PA GB47986-PA HSAL13729-PA PB14766-PA Bger OR 2 XM 015984385.1 HSAL16422-PA Bger OR 55B GB40730-PA Nasvi2EG015517t1 PB11551-PA PB14302-PA HSAL23797-PA XM 015978935.1 HSAL13779-PA FBpp0070028 HSAL14465-PA Nasvi2EG037233t1 RPRC000882-PA PB10377-PA Csec G16859 T1 RPRC002135-PA Nasvi2EG000834t1 XM 015984789.1 XM 015982415.1 Nasvi2EG037135t1 PB15712-PA HSAL17873-PA XM 015978289.1 RPRC006569-PA LOCMI17357 Znev 19063 Bger OR 10 GB52362-PA LOCMI17402 RPRC002161-PA LOCMI17364 Nasvi2EG014916t1 Csec G07888 T1 HSAL13439-PA Bger OR 55A RPRC000140-PA PB18751-PA Nasvi2EG037211t1 HSAL20020-PA HSAL13725-PA Znev 19065 Nasvi2EG003653t1 Nasvi2EG036960t1 Nasvi2EG036976t1 PB26321-PA GB48494-PA HSAL16423-PA FBpp0088122 GB52399-PA GB52391-PA RPRC000423-PA PB26263-PA Nasvi2EG012259t1 LOCMI17413 XM 015982808.1 HSAL13734-PA GB52411-PA Bger OR 98 FBpp0078269 Znev 19035 RPRC001726-PA Csec G08621 T1 LOCMI17422 Nasvi2EG037276t1 Nasvi2EG037305t1 Nasvi2EG037285t1 XM 015983843.1 RPRC000317-PA Nasvi2EG015001t1 Nasvi2EG037220t1 XM 015978598.1 PB26973-PA GB45224-PA LOCMI17588 Csec G13561 T1 RPRC000043-PA Csec G11491 T1 Nasvi2EG037037t1 Nasvi2EG036980t1 HSAL13735-PA PB26760-PA Nasvi2EG037004t1 XM 015983072.1 Bger OR 50D LOCMI17378 GB52400-PA GB42587-PA PB26203-PA EDAN011755-PA FBpp0082351 Csec G13064 T1 XM 015982970.1 LOCMI17625 LOCMI17408 Nasvi2EG019680t1 Nasvi2EG037247t1 Nasvi2EG037235t1 RPRC002009-PA Csec G04512 T1 LOCMI17445 HSAL13307-PA Nasvi2EG037207t1 GB48703-PA HSAL15955-PA Nasvi2EG037041t1 XM 015982611.1 LOCMI17410 PB11391-PA Nasvi2EG037289t1 Bger OR 68 FBpp0072847 LOCMI17421 RPRC001628-PA Bger OR 15 XM 015979768.1 LOCMI17373 XM 015982411.1 Nasvi2EG037228t1 GB50052-PA LOCMI17428 Nasvi2EG037193t1 RPRC002141-PA HSAL11584-PA GB52373-PA GB47953-PA PB26538-PA PB11545-PA PB26124-PA GB52374-PA LOCMI17459 Csec G08636 T1 PB26274-PA Znev 19038 Bger OR 76 HSAL13432-PA FBpp0087587 Csec G08635 T1 Nasvi2EG008584t1 Mnat 07594 XM 015982985.1 HSAL13732-PA LOCMI17367 GB52387-PA HSAL13783-PA LOCMI17451 XM 015983842.1 PB14767-PA RPRC001153-PA LOCMI17416 Bger OR 38 Csec G03320 T1 Nasvi2EG037025t1 XM 015980765.1 PB17576-PA Nasvi2EG037204t1 RPRC000200-PA LOCMI17620 XM 015978855.1 XM 015984142.1 PB26926-PA PB26470-PA GB45079-PA Znev 19044 RPRC000363-PA Znev 18892 XM 015978291.1 XM 015978955.1 Znev 18975 Csec G10896 T1 FBpp0073938 Znev 19070 FBpp0078438 RPRC000541-PA XM 015985332.1 Nasvi2EG037238t1 PB11258-PA LOCMI17437 GB43023-PA PB26765-PA PB22901-PANasvi2EG037104t1 Znev 18894 Nasvi2EG037245t1 LOCMI17460 RPRC002120-PA RPRC000585-PA Nasvi2EG007470t1 Nasvi2EG037227t1 GB40187-PA PB26459-PA PB11558-PA XM 015977943.1 Bger OR 80 GB55927-PA GB42589-PA RPRC000573-PA PB21180-PA LOCMI17435 HSAL13777-PA PB26837-PA HSAL21283-PA Nasvi2EG037170t1 FBpp0087602 RPRC000895-PA Nasvi2EG036931t1 XM 015983603.1 PB26763-PA Nasvi2EG036972t1 Nasvi2EG037093t1 FBpp0081382 Nasvi2EG036970t1 LOCMI17434 HSAL12712-PA LOCMI17608 RPRC000420-PA Bger OR 30 LOCMI17415 HSAL23580-PA Nasvi2EG037226t1Nasvi2EG037221t1 XM 015984433.1 Bger OR 32 RPRC000108-PA GB43025-PA Nasvi2EG037079t1 Nasvi2EG037288t1 Znev 11756 Mnat 15635 RPRC000208-PA HSAL13775-PA LOCMI17454 Nasvi2EG014221t1 LOCMI17417 FBpp0312370 GB51179-PA Nasvi2EG036942t1 XM 008201099.2 FBpp0084595 Nasvi2EG036982t1 Znev 18846 PB26473-PA PB26322-PA HSAL10726-PA Znev 18988 Bger OR 96 GB40183-PA XM 015983692.1 Nasvi2EG037086t1 Nasvi2EG036979t1 GB43354-PA XM 015983954.1 GB52375-PA XM 015985427.1 Nasvi2EG037046t1 Nasvi2EG015565t1 XM 015984532.1 Nasvi2EG037010t1 Nasvi2EG036933t1 HSAL12627-PA XM 964219.2 RPRC000449-PA LOCMI17605 Nasvi2EG036981t1 Bger OR 62 GB52367-PA PB10268-PA Nasvi2EG037145t1 LOCMI17388 XM 015983191.1 GB45078-PA Nasvi2EG036987t1 GB52382-PA Nasvi2EG015560t1 LOCMI17377 HSAL23740-PA PB26572-PA XM 015983588.1 Bger OR 7B PB25414-PA Nasvi2EG037217t1 Nasvi2EG036977t1 HSAL12188-PA LOCMI16592 GB52415-PA Bger OR 45 Bger OR 34 FBpp0079862 Nasvi2EG017812t1 PB18678-PA RPRC000733-PA Nasvi2EG037234t1 XM 015984558.1 Znev 19061 GB41940-PA Nasvi2EG037002t1 PB10467-PA XM 015984382.1 PB18167-PA Nasvi2EG037171t1 PB26762-PA PB27136-PA GB40201-PA Nasvi2EG036921t1 XM 015982864.1 Nasvi2EG007012t1 Znev 18874 HSAL12625-PA GB55924-PA XM 015980762.1 Csec G08622 T1 HSAL18849-PA LOCMI17604 PB16373-PA GB52405-PA Nasvi2EG037125t1 Csec G06759 T1 Nasvi2EG004895t1 RPRC000413-PA PB20272-PA RPRC000459-PA LOCMI17418 Mnat 12200 Nasvi2EG037096t1 PB11550-PA Nasvi2EG036415t1 Bger OR 73 XM 015985327.1 Csec G08624 T1 Nasvi2EG000548t1 FBpp0076029 LOCMI17455 Nasvi2EG003774t1 LOCMI17396 PB26716-PA PB25407-PA RPRC000607-PA Nasvi2EG037290t1 PB27045-PA PB20273-PA Bger OR 90 HSAL14206-PA Znev 18893 Bger OR 7A PB27137-PA PB26592-PA Znev 18906 GB40193-PA Znev 19069 LOCMI17360 Bger OR 11 PB10171-PA FBpp0071134 HSAL13789-PA HSAL14210-PA Nasvi2EG037119t1 HSAL21036-PA Nasvi2EG008426t1 FBpp0076712 Nasvi2EG037354t1 Bger OR 22 FBpp0087212 Bger OR 43 LOCMI17386 Znev 19048 LOCMI17609 Nasvi2EG037212t1 RPRC000079-PA Nasvi2EG037089t1 RPRC000362-PA PB26463-PA Bger OR 107 LOCMI17598 LOCMI17400 LOCMI17450 Znev 18875 RPRC002158-PA HSAL16502-PA Nasvi2EG037205t1 LOCMI17424 Nasvi2EG033867t1 XM 015983499.1 Csec G16798 T1 HSAL15956-PA Nasvi2EG036943t1 XM 015981313.1 LOCMI17409 LOCMI17623 XM 015984396.1 LOCMI17419 LOCMI17426 Nasvi2EG037179t1 RPRC000095-PA GB49018-PA RPRC000337-PA XM 015985429.1 HSAL20721-PA RPRC000093-PA Nasvi2EG037029t1 Nasvi2EG037134t1 PB27175-PA RPRC001293-PA Nasvi2EG010113t1 FBpp0070381 PB14127-PA Csec G08626 T1 LOCMI17374 GB46324-PA RPRC000120-PA Nasvi2EG037143t1 GB43024-PA Nasvi2EG037138t1 Nasvi2EG037130t1 Csec G09666 T1 XM 015978627.1 LOCMI17616 RPRC006981-PA HSAL16480-PA PB16374-PA Bger OR 48 PB26461-PA LOCMI17393 HSAL15953-PA FBpp0076713 FBpp0081383 LOCMI17406 RPRC006542-PA XM 015985428.1 XM 015979747.1 RPRC000166-PA RPRC001689-PA Znev 19021 Nasvi2EG014125t1 Bger OR 118 RPRC002162-PA LOCMI17370 Nasvi2EG001283t1 Nasvi2EG036974t1 XM 015982836.1 RPRC000229-PA Nasvi2EG037072t1 Nasvi2EG037218t1 PB10095-PA GB52392-PA Nasvi2EG036953t1 XM 015982615.1 Nasvi2EG010712t1 Bger OR 89 Nasvi2EG037088t1 RPRC000476-PA Nasvi2EG036986t1 PB23847-PA RPRC000235-PA PB26615-PA Nasvi2EG036930t1 XM 015978571.1 LOCMI17466 XM 962666.2 XM 008201090.2 PB27144-PA XM 015984128.1 GB52412-PA Csec G04894 T1 Bger OR 84 XM 015983496.1 GB52380-PA Nasvi2EG002317t1 GB52364-PA Nasvi2EG037277t1 Csec G09269 T1 RPRC000245-PA LOCMI17442 Nasvi2EG036922t1 Mnat (5) Csec (26) Znev (25)Bger (16) Bger (20) Znev (9)Csec (5)Mnat (2) Bger (37) Znev (18)Csec (14)Mnat (3) Csec (5), Znev (3), Bger (6), Lmig (2) Mnat (1), Csec (3), Znev (3), Bger (5) OR co-receptors d N CCsec N C TM1 TM2 TM3 TM4 TM5 TM6 TM7 Mnat C N TM1 TM2 TM3 TM4 TM5 TM6 TM7 * c M1 P M2 M3 S1S2 TM1 P TM2 TM3 C N N C * * * S1 TM1 TM2 TM3P S2 Mnat a not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint PC2 (13.4%) PC3 (11.7%) PC1 (44.1%) Znev Mnat Csec Lmig Edan Rpro Bger low CpGo/e T ermites Blattella germanica high CpGo/e a b Sensory perception Gene regulation Transposition Development Immune response Other biological proc esses Signalling DNA repair transcription initiation from RNA polymerase II promoter regulation of transcription from RNA polymerase II promoter nucleobase−containing compound metabolic process proteolysis involved in cellular protein catabolic process oxidation−reduction process signal transduction cytoskeleton organization protein dephosphorylation protein import into mitochondrial inner membrane protein ubiquitination post−translational protein modification protein deacetylation retrograde vesicle−mediated transport, Golgi to ER negative regulation of microtubule depolymerization ATP synthesis coupled proton transport pentose−phosphate shunt, non−oxidative branch regulation of protein metabolic process proton−transporting ATP synthase complex assembly chromatin silencing metabolic process tryptophan catabolic process to kynurenine apoptotic process biosynthetic process 7−methylguanosine RNA capping purine nucleotide biosynthetic process cholesterol biosynthetic process positiv e regulation of apoptotic process pseudouridine synthesis calcium−mediated signaling cell proliferation cell redox homeostasis L−cysteine metabolic process DNA replication initiation prenylcysteine catabolic process metal ion transport cellular iron ion homeostasis RNA meth ylation tRNA methylation iron ion transport cell cycle regulation of transcription, DNA−templated G−protein coupled receptor signaling sensory perception of taste DNA integr ation homophilic cell adhesion via plasma membrane adhesion molecules Wnt signaling pathway, calcium modulating path way multicellular organism de velopment chromatin assembly or disassembly sensory perception of smell cyclic nucleotide biosynthetic process lipopolysaccharide biosynthetic process pheromone biosynthetic process ecdysis, chitin −based cuticle purine nucleobase metabolic process glutamine metabolic process carbo xylic acid metabolic process neuropeptide signaling path way chitin metabolic process transmembrane tr ansport peptidoglycan catabolic process Notch signaling path way innate immune response transposition, DNA −mediated sex differentiation nucleosome assembly peptide cross −linking sodium ion transport DNA replication ion tr ansport citrate transport lipid transport cell adhesion proteolysistransport growth pathway regulation of transcription from RNA polymerase II promoter tRNA aminoacylation for protein translation phospholipid biosynthetic process protein phosphorylation mitochondrial electron transport, ubiquinol to cytochrome c translational initiation microtubule cytoskeleton organization regulation of mitotic metaphase/anaphase transition arginyl−tRNA aminoacylationtranslation thiamine diphosphate biosynthetic process DNA topological change 'de novo' IMP biosynthetic process cobalamin biosynthetic process arginine biosynthetic process inositol biosynthetic process lipopolysaccharide biosynthetic process regulation of tr anslational fidelity mature ribosome assembly DNA replication Golgi organization sulfate assimilationmetabolic process chromatin remodeling phagocytosis carbon utilization heme oxidation citrate transport DNA repair endocytosis oxidation−reduction process metabolic process transmembrane transport tRNA aminoacylation for protein translation Mo−molybdopterin cofactor biosynthetic pathway carbohydrate metabolic process double−strand break repair via nonhomologous end−joining retrograde vesicle−mediated tr ansport, Golgi to ER ER to Golgi vesicle−mediated transport folic acid−containing compound biosynthesis nucleobase−containing compound metabolic process microtubule−based movement GPI anchor biosynthetic process intracellular protein transport protein phosphorylation UMP biosynthetic process fatty acid beta−oxidation cellular protein metabolic process regulation of protein metabolic process phenylalanyl−tRNA aminoacylation post −translational protein modification integrin−mediated signaling path way alanyl−tRNA aminoacylation vesicle−mediated transport fatty acid metabolic process superoxide metabolic process cellular metabolic process glycerol metabolic process iron−sulfur cluster assembly base−excision repair biosynthetic process oligopeptide tr ansport protein deacetylation cell redox homeostasis protein meth ylation cation transport protein localization proteolysis chromatin silencing glycolytic process metal ion tr ansport catabolic process rRNA processing RNA processing protein processing protein tr ansport DNA repair iron ion transport protein folding meth ylation not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which wasthis version posted November 15, 2017. ; https://doi.org/10.1101/181909doi: bioRxiv preprint