bioRxiv preprint doi: https://doi.org/10.1101/717322; this version posted July 28, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license.
ARTICLES PREPRINT
Evolution of a pest: towards the complete neuroethology of Drosophila suzukii and the subgenus Sophophora Ian W. Keesey1*, Jin Zhang1, Ana Depetris-Chauvin1, George F. Obiero2, Markus Knaden1‡*, and Bill S. Hansson1‡*
Comparative analysis of multiple genomes has been used extensively to examine the evolution of chemosensory receptors across the genus Drosophila. However, few studies have delved into functional characteristics, as most have relied exclusively on genomic data alone, especially for non-model species. In order to increase our understanding of olfactory evolution, we have generated a comprehensive assessment of the olfactory functions associated with the antenna and palps for Drosophila suzukii as well as sev- eral other members of the subgenus Sophophora, thus creating a functional olfactory landscape across a total of 20 species. Here we identify and describe several common elements of evolution, including consistent changes in ligand spectra as well as relative receptor abundance, which appear heavily correlated with the known phylogeny. We also combine our functional ligand data with protein orthologue alignments to provide a high-throughput evolutionary assessment and predictive model, where we begin to examine the underlying mechanisms of evolutionary changes utilizing both genetics and odorant binding affinities. In addition, we document that only a few receptors frequently vary between species, and we evaluate the justifications for evolution to reoccur repeatedly within only this small subset of available olfactory sensory neurons.
INTRODUCTION show the conserved nature of that functional neuronal type 19–23. Some more distantly related Drosophila species have begun to One of the advantages of working within the Drosophila be examined in more depth, such as D. mojavensis 24, which is a genus includes the wide-array of evolutionary specializations that model for incipient speciation; however, this species is a member these species display, especially in regard to host preference, but of an entirely different subgenus, and as such, is quite far removed also habitat choice, morphology and mate selection 1,2. As an from the more robust datasets afforded by the melanogaster additional benefit, the genus affords a vast amount of genomic clade. These factors may also mean that these distantly related data, which has been generated and accumulated since the early species are much more difficult to utilize to assess patterns 21st century 3,4. The last ten years have also brought attention and mechanisms of evolutionary selective pressure, at least in towards another novelty from this group, an agricultural pest their direct comparison to the established molecular models. species in the form of Drosophila suzukii, which has now invaded Therefore, in contrast, the detailed examination of the non- North and South America, Europe as well as Asia 5–9. Furthermore, melanogaster members of the Sophophora subgenus, which itself this pest insect has prompted both integrated pest management includes substantial variation in host and habitat choice, perhaps efforts as well as evolutionary neuroethology research, where the represents a closer group of relatives to optimize the comparisons distinct ecological niche of attacking fresh or ripening fruit has of evolutionary variation in chemosensation. Here again, D. afforded a unique opportunity for comparison among the other suzukii and its subgroup offer an ideal model for unraveling the model species within this well-studied genus of flies, which usually complexities of olfactory evolution, especially given their relative target softened, fermented host resources10–13 . phylogenetic proximity to the principal scientific models within Previous research has sought to outline either the olfactory the melanogaster clade 1,2. or gustatory systems of several species of Drosophila, either A fundamental aspect of studying evolution revolves for evolutionary or ecological comparisons of host plant, mate around understanding the processes by which genetic variation preference or reproductive isolation, including D. melanogaster, can be generated naturally, such as through mutation, genetic D. sechellia, and D. simulans 14–17. However, only a few studies drift, or recombination, and then how selective forces, have conducted electrophysiological assessments of members including environmental, ecological and developmental factors, outside of this subgroup 18, with most research examining non- subsequently act upon this variance to drive speciation and melanogaster species only in regards to a single, specific olfactory specialization. For many Drosophila species, there already exists a sensory neuron (OSN) of interest, and usually only as a means to robust library of molecular resources, as nearly complete genomic
1 Max Planck Institute for Chemical Ecology, Department of Evolutionary Neuroethology, Hans-Knöll-Straße 8, D-07745 Jena, Germany 2 Department of Biochemistry and Biotechnology, Technical University of Kenya, Haille-Sellasie Avenue, Workshop Rd, 0200, Nairobi, Kenya ‡ These authors share senior authorship * Correspondence to: [email protected] ; [email protected] ; [email protected]
1 bioRxiv preprint doi: https://doi.org/10.1101/717322; this version posted July 28, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license.
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A D. suzukii D. melanogaster
SensillumNeuron TypeBest Ligand SensillumNeuron TypeOlfactory ReceptorGlomerulus Best Ligand ab1 A Ethyl acetate ab1 A Or42b DM1 Ethyl acetate B Ethyl lactate B Or92a VA2 Ethyl lactate C CO2 C Gr21a/Gr63a V CO2 D Methyl salicylate D Or10a/Gr10a DL1 Methyl salicylate A Methyl acetate A Or59b DM4 Methyl acetate
) ab2 ab2 -4 B Isopentyl acetate B Or85a DM5 Ethyl-3-hydroxybutyrate ab3 A Isobutyl acetate / beta-Cyclocitral ab3 A Or22a/Or22b DM2 Ethyl hexanoate B 2-heptanol B Or85b VM5d 2-heptanol IBA (10 ab4 A E-2-hexenal ab4 A Or7a DL5 E-2-hexenal B Geosmin B Or56a/Or33a DA2 Geosmin ab5 A Geranyl acetate (weak) ab5 A Or82a VA6 Geranyl acetate B Pentyl acetate B Or47a/Or33b DM3 Pentyl acetate ab6 A 1-Octen-3-ol ab6 A Or13a DC2 1-Octen-3-ol B Guaiacol B Or49b VA5 Guaiacol ab7 A Ethyl benzoate (weak) ab7 A Or98a VM5v Ethyl benzoate B Ethyl lactate (weak) B Or67c VC4 Ethyl lactate ab8 A Ethyl E-2-butanoate ab8 A Or43b VM2 Ethyl-E-2-butanoate B 3-Hydroxy-2-butanone B Or9a VM3 3-Hydroxy-2-butanone ab9 A Linalool/Citral ab9 A Or69a/Or69b D Linalool/Citral
Antennal Sensillum Antennal B ??? B Or67b VA3 2-Phenylethanol
Or22a/Or22b ab10 A Benzyl butyrate (weak) ab10 A Or67a DM6 Benzyl butyrate B Iridomyrmecin, actinidine, nepetalactol B Or49a/Or85f DL4 Iridomyrmecin, actinidine, nepetalactol ai2 A Farnesol ai2 A Or83c DC3 Farnesol B ??? B Or23a DA3 ??? A E-2-Hexenol, Valencene A DC1 Valencene ab3A ai3 ai3 Or19a/Or19b B ??? B Or2a DA4m ??? C Methyl indole C Or43a DA4L Methyl indole at1 A cVA at1 A Or67d DA1 cVA
at4 A ML, MM at4 A Or47b VA1v ML, MM B ??? B Or65a/b/c DL3 ??? C MP C Or88a VA1d MP
pb1 A Gamma-hexalactone pb1 A Or42a VM7 Gamma-hexalactone B 4-ethylguaiacol B Or71a VC2 4-ethylguaiacol pb2 A beta-ionone pb2 A Or33c/Or85e VC1 beta-ionone B Phenol B Or46a VA7L Phenol pb3 A Benzyl cyanide pb3 A Or59c 1 Benzyl cyanide B 2-phenethyl acetate B Or85d VA4 2-phenethyl acetate Palp Sensillum Palp
Or85a VA5 VA5 B VA7m C D VA7m E
VC2 VC2 VL1 VL1
VC1 VC1 DC2 DC2 DM5 VL2p DM5 VL2p DL2v DL2v DC1 DC1 DM2 DM2
DP1l DP1l
DM1 DM1 DP1m DP1m Or22a Or22b
Or85b VA3 VA3 VA2 VA2 VA4 VA4
VM2 VC4 VC3 VM2 VC4 VC3
VM5d VM5d VM7v VM7v VM7d VM7d
DC4 DC4
DM4 DM4
Or59b D. melanogaster D. suzukii D. melanogaster D. suzukii Figure 1. Complete olfactory sensory neuron screen of Drosophila suzukii. (A) Shown are the sensillum subtypes, olfactory sensory neurons (OSNs), and best ligands for each of those olfactory channels found within either the antennae or the palps of these two adult Drosophila species. For Drosophila melanogaster, additional information is given regarding the known olfactory receptor and the corresponding glomerulus for the antennal lobe (AL) for each OSN. In total, 37 neurons were analyzed from each species using single sensillum recordings (SSR), where 86% of the receptors retain the same ligand spectra, while only five OSNs (14%) displayed variation in the odorant that produced the largest response. Responses that were different between the two species are highlighted in orange. (B-E) Here we show immunostaining with neurobiotin (green) and nc82 (red or magenta) across the antennal lobes of D. suzukii adults, which were backfilled from the ab2-like or ab3-like sensillum types. Each sensillum type was first identified with electrophysiological contact and odor response recordings from the antenna before application of the neurobiotin. (B) Spatial map of the antennal lobe (AL) atlas D.of melanogaster, with corresponding positions (marked in orange) of neurons emanating from the ab2 sensillum. (C) Neuronal wiring in D. suzukii that stem from single-sensillum backfills of the ab2-like sensillum, showing similar mapping to the model species. (D) Spatial map of the AL of D. melanogaster, with corresponding positions (marked in orange) of neurons emanating from the ab3 sensillum. (E) Neuronal wiring inD. suzukii that stem from single-sensillum backfills of the ab3-like sensillum. This spatial congruence between these two species, combined with the morphological (e.g. large basiconic identity) and odor ligand data from the other neuron housed in either ab2 or ab3 sensillum (e.g. Or59b and Or85b), continues to provide strong, corroborating evidence that the wiring forD. suzukii and D. melanogaster is unchanged for these two sensilla types. This conservation of wiring within the antennal lobe is despite changes in odorant sensitivity and ligand spectra for the ab2B and ab3A OSNs between these two species. 2 bioRxiv preprint doi: https://doi.org/10.1101/717322; this version posted July 28, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license.
ARTICLES PREPRINT datasets are publically available; however, far less information is suzukii) share identical ligand spectra for the second OSN in the available describing the functional components of host ecology, same sensillum ab3B, which responds characteristically towards such as olfactory, gustatory, auditory and visual preferences, or 2-heptanol as the best ligand. We also found that although the describing behavioral or habitat variations between these species. ab2B OSN from D. suzukii deviates from D. melanogaster, that the Thus for most members across this incredibly well studied genus, ab2A OSN appears identical. As such, we believe these two sensilla more is known about their genome than their ecology. In an effort in D. suzukii (ab2-like and ab3-like) provide the strongest match to expand our knowledge about these species, here we provide a for the comparative sensilla in D. melanogaster, even given the functional olfactory landscape encompassing 20 different species deviations in ligand spectra for select neurons. This idea is further within the Sophophora subgenus, with particular focus on D. supported by the morphological structure of the sensillum (i.e. suzukii and its closest relatives, in order to examine the common large basiconic), and by the large amplitudes that are characteristic variables that are associated with the evolutionary emergence of of these sensillum types relative to the small basiconics. All this insect pest. In addition, we address how protein sequence these factors combine to strongly-support the identity of these coding of olfactory receptors correlate with functional evolution two sensillum types in D. suzukii adults, despite the variation in of odorant binding and ligand selectivity, here through high- olfactory ligand spectra, as has been previously proposed 10. throughput comparison of chemosensory variation overlaid with tertiary protein structures from the available model species. Thus, Conservation of neuronal wiring despite ligand spectrum in summary, the present study establishes the missing ingredients shifts for us to start understanding the mechanisms of olfactory We wanted to continue to explore the neural pathways for the ab2- evolution, while also providing critical chemosensory evaluations like and ab3-like OSNs in D. suzukii, we therefore next documented across 20 species within this highly influential genus of insects. the circuitry from the antenna towards the antennal lobe (AL) using the single-sensillum backfill neuronal staining technique 29. RESULTS By comparing the labeled glomeruli from D. suzukii to the known AL atlas of D. melanogaster (Figure 1 B-E) 30, we could provide Complete screening of Drosophila suzukii olfaction additional support for the ab2-like and ab3-like sensillum type In order to assess olfactory ligand spectra for all sensillum types, identification, as these two fly species shared identical wiring for we first revisited the classical model, D. melanogaster, where the these OSNs, despite the observed deviation in ligand spectra for responses of most sensilla have been previously established 14,24–28. D. suzukii adults. Here we again mapped out the response profile from each known receptor across the adult fly antenna and palps using a panel of Spatial mapping of sensillum subtypes 80 odorants (Figure 1). We found that out of 37 unique olfactory We next mapped the position of all sensillum subtypes on the sensory neurons (OSNs), only 3 are still currently without a D. suzukii and D. melanogaster antenna and palps using several strong ligand candidate (i.e. Or23a, Or2a, Or65a/b/c), although in individuals to create an aggregate diagram. The heads of both addition, several co-expressed receptors also have poorly defined D. suzukii and D. melanogaster were positioned in four ways to response profiles (e.g. Or33a which is found in ab4B). Next, we completely map the spatial pattern for sensillum abundance, shifted our focus towards the antenna and palps ofD. suzukii using including antennal arista down, arista side, arista up as well as across the same diverse panel of 80 odorants in order to examine any the maxillary palp (Figure 2A-D). Here, as we had established ligand changes in functional ligand spectra between these two species. spectra for all OSNs of both species, we used SSR data to document Here we determined that of the 37 OSNs found in D. suzukii, the position of each sensillum type on the antenna, which provided approximately 86% are functionally conserved between the two spatial information as well as the relative abundance, and this data species (Figure 1A). Moreover, we identified only five OSNs that matches previous studies of D. melanogaster antenna 19,31. The showed robust functional deviation from the D. melanogaster positioning of each specific sensillum type was nearly identical in model, including the ab2B-like, ab3A-like, ab9B-like, ab10A-like, both species and arranged in concentric circles or zones; however, and ai3A-like sensory neurons (Figure 1A). We also document we did observe large variations in the abundance of sensillum minor shifts across ab5 and ab7, though not as strong, and we note types between species (Figure 2 E-H). For example, within the that we found very few ab8 sensilla across D.suzukii trials. Two of large basiconics, while the ab1 sensillum counts were nearly the larger deviations in olfactory response forD. suzukii have been identical, we found that D. suzukii had almost twice as many ab2 previously described, including ab2B and ab3A 10. Interestingly, in as D. melanogaster (22 and 12, respectively). In contrast, we found the present study we identify two separately responding ab3A OSN that D. melanogaster had more than twice as many ab3 sensilla types, with an approximately 60:40 split ratio for OSN abundance, compared to D. suzukii (13 and 6, respectively). It was possible where the first (type i) is tuned towards isobutyl acetate (IBA) and to identify large basiconics due to physical metrics, like width, tip the second (type ii) responds most strongly to beta-cyclocitral shape and length (Supplementary Figure 1). In addition, we could (βCC). Both of these ab3A type OSNs in D. suzukii are tuned quite also positively identify large basiconics based on the amplitude of differently than those found within the corresponding sensillum of SSR spike response (Figure 2 I-K). Here the ab2 and ab3 sensilla D. melanogaster, which is tuned instead towards ethyl hexanoate are also uniquely identifiable based on the ratio of the A neuron (EH). However, both species (and both type i and type ii, in D. to B neuron spike sizes, where ab2 has an exaggerated disparity
3 bioRxiv preprint doi: https://doi.org/10.1101/717322; this version posted July 28, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license.
ARTICLES PREPRINT between the two neurons, while the ab3 neurons are much interest we counted from were the same during the comparison of closer in spike size (Figure 2 I-K). Moreover, the ab1 sensillum is the two species 25. As such, these counts represent strong relative straightforward to identify due to its unique housing of four OSNs, values for species-specific comparisons of sensillar abundance. In 32 as well as the characteristic response to CO2 odor cues . While addition, it is important to note that the flies do differ in absolute our sensillum counts are not the absolute total number of large size, as does their antennal surface area 1. However, we still show basiconics, this was provided previously for these two species a consistent difference that is not explained by the larger size of 1, and we are confident that the preparations and the zones of D. suzukii relative to D. melanogaster adults, therefore,Figure additional 2 A B C D
arista down arista side arista up maxillary palp
E D. melanogaster F G H D. suzukii D. suzukii D. melanogaster D. suzukii D. melanogaster D. melanogaster ab1 ab2 ab3 ab4 ab5 ab6 ab7 ai2 ab8 ai3 pb1 ab9 at1 pb2 ab10 at4 pb3 D. suzukii
4 4 I JK EH ab2 ab3 D. melanogaster (ab2) D. melanogaster (ab3) A 2 A 2
B B 0 0
- 2 melanogaster D. - 2 Amplitude (mV)Amplitude (mV)Amplitude
- 4 - 4
4 4 D. suzukii (ab2) IBA D. suzukii (ab3) A 2 A 2
B B
0 0 D. suzukii D.
- 2 - 2 Amplitude (mV)Amplitude Amplitude (mV)Amplitude
- 4 - 4
Figure 2. Olfactory sensory neuron mapping across the antennae of Drosophila suzukii. (A-D) The antennae of each Drosophila species were prepared in several different positions to optimize access to the various sensillar subtypes, including the four positions forD. suzukii that are shown. (E) Schematic drawing of the third antennal segment from both species, where spatial distribution and sensillum abundance were identified during single sensillum recordings (SSR). Each color denotes a unique subtype. Shown are the sensillar mappings from the arista down configuration from both target species. (F) Arista side configuration. (G) Arista up preparation. (H) Maxillary palp preparation. (I) Shown are example traces of the ab2 sensillum type, here we note maximum amplitudes well over 2 mV for each species. (J) Consistent ratio of amplitude differences between the A and B neurons for both ab2 and ab3 sensillum. Here we observed that the ratio for A:B in ab2 was larger than for ab3 recordings. We also note that ab3 has smaller amplitudes than ab2 types. (K) Shown are example traces of the ab3 sensillum type, where we note maximum amplitudes at or near 2 mV for each species. Here the relative size of the ab3B neuron amplitude is much larger when compared to ab2B, in relation to their respective A neurons for ab2 and ab3 sensillum types. In general, these stereotyped response dynamics appear conserved between species, and add an additional layer of confirmation to the chemical and morphological identification of sensillum type in these novel Drosophila species.
4 bioRxiv preprint doi: https://doi.org/10.1101/717322; this version posted July 28, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license.
ARTICLES PREPRINT evolutionary factors are in play concerning sensillum abundance. tuned (Figure 4 B). Other neurons within this sensillum type, such as ab1A (which expresses Or42b in D. melanogaster), were Proportional analyses of sensilla acrossSophophora functionally identical in each new species that we tested, with all As we were interested in the evolution of olfaction in D. suzukii, species responding most strongly to ethyl acetate. However, two we next sought to examine 18 additional species within the OSNs were consistently variable over the Sophophora subgenus, Sophophora subgenus in order to generate an evolutionary including the same neurons that we originally show were different framework of sensillar variation Figure ( 3). Here we screened in D. suzukii, namely ab2B and ab3A (Figure 4 C,D). each new species for ab1, ab2 and ab3-like sensilla using a wide In general, ab3A (which co-expresses both Or22a and panel of odorants. The total sample sizes across individuals (n) and Or22b in D. melanogaster) was the most commonly changed across large basiconic sensillum recordings (R) is listed above each OSN within our 20 fly species Figure( 4), and the response usually species (Figure 3), and we present the data via a proportion of fell into one of four main categories of best odorant profile (i.e. these total SSR basiconic contacts. As has been shown previously methyl hexanoate (MH), ethyl hexanoate (EH), isobutyl acetate 19, we confirm the overrepresentation of the ab3 sensillum for the (IBA) or beta-cyclocitral (βCC)). The different ligands for ab3A often entirety of the melanogaster clade (Figure 3; ab3 bias; shown in seemed to be conserved between closely related species. Again, red), including the confirmation of the most heavily ab3-biased we noted two different functional ab3A types in D. suzukii, where species, D. sechellia and D. erecta. Intriguingly, we also document one was tuned to IBA (type i) and the other type was tuned best that each of the spotted wing species within the suzukii clade all towards βCC (type ii) (Supplementary Figure 2 A-D). We observed display, in contrast to the melanogaster clade, a reduction in ab3 these two types of ab3-like sensilla within the same individual and a corresponding overrepresentation of the ab2-like sensillum animal, with a roughly 60:40 split between those detecting IBA (Figure 3; ab2 bias; shown in green). We believe the ab3 bias of and βCC, respectively. Further work is needed to confirm and the melanogaster clade might be linked to ecologically associated reconstruct the neural circuit for these two sensillum types in D. detection of strong fermentation components, while the ab2 suzukii, as while they map to the same glomerulus (DM2-like), it sensilla is perhaps more tightly associated with fresh fruit esters was unclear from our neuronal backfills whether type i and type or the fruit ripening process, but more work is needed to examine ii map to different regions within this same glomerulus. We note this in behavioral trials. Our screen of the Sophophora subgenus that several functional types of this ab3A OSN have been recently also provided evidence for an ab1 bias in several species (Figure 3; reported within D. melanogaster populations 40,41. One variant shown in yellow). We do not have enough ecological information expression is a chimeric gene, Or22ab, which arises from the fusion about many of these species, but we can speculate that perhaps of both the Or22a and the Or22b genes into a single receptor there exists a common association with higher altitudes and protein 40,41. Moreover, we note that the ligand spectrum reported altered CO2 concentrations in these alpine habitats (e.g. D. birchii) for this Or22ab variant is functionally similar to our current data 33, or that floral-, foliage-, and yeast-derived host signals34–36 might in cases where we observe that IBA is the best ligand for many play a role in this association with 2 CO sensitivity. Lastly, there of these Sophophora species, including D. suzukii adults. Thus, it were four species that had a roughly even proportion of basiconic is possible that this variant receptor type also occurs within non- sensillum types, which are shown in grey. Again, we are greatly melanogaster members of this genus. Another consideration limited due to the unknown ecology of most species, thus it is is that Or22c, which is usually expressed only in larvae, detects currently unclear what the evolutionary or ecological rationale is 2-acetylpyridine which bears aromatic, chemical similarities to for these differences in sensillum abundance. βCC 26; therefore, another chimeric form, such as Or22ac, may also be possible in nature (Supplementary Figure 2 B-E). Overall, the Functional ligand spectra for sensillum types across olfactory changes for ab3A were more of a gradient or moderate Sophophora shift between species, such as the transition from EH to MH as As we had established that some of the main olfactory differences the best ligand (e.g. D. melanogaster and D. sechellia). We also between D. suzukii and D. melanogaster were related to large note two species for which we could not identify any strong basiconics, we sought to test this hypothesis by looking at olfactory ligand for ab3A, D. elegans and D. subobscura. Here, not much is ligand variation across our 20 species within the Sophophora known about the ecology, and despite efforts to screen a variety of subgenus (Figure 4). Here we screened each species with a diverse headspace collections from flowers, fruits and tree sap (including panel of odorants, including high-throughput testing that utilized in total several thousand compounds), we could not identify any fruit, foliage and floral headspace extracts via gas-chromatography suitable ligand candidates for this OSN in these two species. mass-spectrometry combined with single-sensillum recordings Intriguingly, we also observed several changes in the (GC-SSR). Of the ten OSN types we examined from each of the 20 ab2B neuron across our 20 examined species. In this case, species, only three showed any dynamic variation in ligand spectra most species displayed exactly the same ligand (i.e. ethyl-3- or ligand sensitivities (i.e. ab1C, ab2B, ab3A; Figure 4). We note hydroxybutyrate (E3HB)). However, seven Sophophora members that the ab1C neuron (which co-expresses Gr21a and Gr63a in D. had an acute change in ligand spectra, where none of these seven species shared any overlap for their novel ligand, making melanogaster) displayed a large variation in sensitivity towards 2CO between our species, which has been suggested previously 10,37–39; each of these adaptations entirely species-specific. For example, 10 however, the best ligand for this receptor was found to be identical as was published previously , we again demonstrated that D. between all those tested, and ab1C appears to remain narrowly suzukii ab2B has changed ligand spectrum towards the detection 5 bioRxiv preprint doi: https://doi.org/10.1101/717322; this version posted July 28, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license.
ARTICLES PREPRINT
n=44345443433435443434 R=(177) (117) (83) (169) (121) (168) (187) (57) (167) (69) (59) (88) (89) (80) (54) (86) (34)(82) (60) (53) 1 a 3