Genomic and functional insights into dietary diversification in New World leaf-nosed bats (Phyllostomidae)

Abstract

The most significant driver of adaptive radiation in the New World leaf-nosed bats (Phyllostomidae) is their remarkably diverse feeding habits, yet there remains a notable scarcity of studies addressing the genetic underpinnings of dietary diversification in this family. In this study, we have assembled a new genome for a representative species of phyllostomid bat, the fringe-lipped bat (Trachops cirrhosis), and integrated it with eight published phyllostomid genomes, along with an additional 10 genomes of other bat species. Comparative genomic analysis across 10 200 orthologus genes has unveiled that those genes subject to divergent selection within the Phyllostomidae clade are notably enriched in metabolism-related pathways. Furthermore, we identified molecular signatures of divergent selection in the bitter receptor gene Tas2r1, as well as 14 genes involved in digesting key nutrients such as carbohydrates, proteins, and fats. In addition, our cell-based functional assays conducted on Tas2r1 showed a broader spectrum of perception for bitter compounds in phyllostomids compared to nonphyllostomid bats, suggesting functional diversification of bitter taste in Phyllostomidae. Together, our genomic and functional analyses lead us to propose that divergent selection of genes associated with taste, digestion and absorption, and metabolism assumes a pivotal role in steering the extreme dietary diversification in Phyllostomidae. This study not only illuminates the genetic mechanisms underlying dietary adaptations in Phyllostomidae bats but also enhances our understanding of their extraordinary adaptive radiation.

Keywords

bats / dietary diversification / evolutionary genomics

References

[1] AdeyA,KitzmanJO, BurtonJN,Daza R,KumarA,ChristiansenL,RonaghiM, AminiS,Gunderson KL,SteemersFJ,ShendureJ. 2014. In vitro, long-range sequence information for de novo genome assembly via transposase contiguity. Genome Research 24:2041–2049.

[2] AloyP,Catasús L,VillegasV,ReverterD,Vendrell J,AvilésFX. 1998. Comparative analysis of the sequences and three-dimensional models of human procarboxypeptidases A1, A2 and B. Biological Chemistry 379:149–155.

[3] ArbourJH,CurtisAA, SantanaSE. 2019. Signatures of echolocation and dietary ecology in the adaptive evolution of skull shape in bats. Nature Communications 10:2036.

[4] AthaudaSB,TanjiM, KageyamaT,Takahashi K. 1989. A comparative study on the NH2-terminal amino acid sequences and some other properties of six isozymic forms of human pepsinogens and pepsins. Journal of Biochemistry 106:920–927.

[5] BaroneS,FussellSL, SinghAK,Lucas F,XuJ,KimC,WuX, YuY,AmlalH, SeidlerU,Zuo J,SoleimaniM. 2009. Slc2a5 (Glut5) is essential for the absorption of fructose in the intestine and generation of fructose-induced hypertension. Journal of Biological Chemistry 284:5056–5066.

[6] BensonG. 1999. Tandem repeats finder: A program to analyze DNA sequences. Nucleic Acids Research 27:573–580.

[7] BergmanCM,Quesneville H. 2007. Discovering and detecting transposable elements in genome sequences. Briefings in Bioinformatics 8:382–392.

[8] BielawskiJP,YangZ. 2004. A maximum likelihood method for detecting functional divergence at individual codon sites, with application to gene family evolution. Journal of Molecular Evolution 59:121–132.

[9] BirneyE,ClampM, DurbinR. 2004. Genewise and genomewise. Genome Research 14:988–995.

[10] BlumerM,BrownT, FreitasMB,Destro AL,OliveiraJA,MoralesAE,SchellT, GreveC,Pippel M,JebbD,HeckerN,AhmedAW, KirilenkoBM,Foote M,JankeA,LimBK,HillerM. 2022. Gene losses in the common vampire bat illuminate molecular adaptations to blood feeding. Science Advances. 8: eabm6494.

[11] BrawandD,WagnerCE, LiYI,Malinsky M,KellerI,FanS,SimakovO, NgAY,LimZW, BezaultE. 2014. The genomic substrate for adaptive radiation in African cichlid fish. Nature 513:375–381.

[12] BrunA,PriceER, Gontero-FourcadeMN,Fernandez-MarinoneG,Cruz-Neto AP,KarasovWH,Caviedes-VidalE. 2014. High paracellular nutrient absorption in intact bats is associated with high paracellular permeability in perfused intestinal segments. The Journal of Experimental Biology 217:3311–3317.

[13] BurgeC,KarlinS. 1997. Prediction of complete gene structures in human genomic DNA. Journal of Molecular Biology 268:78–94.

[14] CamachoC,Coulouris G,AvagyanV,MaN,Papadopoulos J,BealerK,MaddenTL. 2009. BLAST+: Architecture and applications. BMC Bioinformatics 10:421.

[15] Caviedes-VidalE,KarasovWH, ChediackJG,Fasulo V,Cruz-NetoAP,OtaniL. 2008. Paracellular absorption: A bat breaks the mammal paradigm. PLoS One 3: e1425.

[16] ChenJ,EngleSJ, SeilhamerJJ,Tischfield JA. 1994. Cloning and recombinant expression of a novel human low molecular weight Ca(2+)-dependent phospholipase A2. The Journal of Biological Chemistry 269:2365–2368.

[17] ChengD,NelsonTC, ChenJ,Walker SG,Wardwell-SwansonJ,MeegallaR,TaubR, BillheimerJT,RamakerM,FederJN. 2003. Identification of acyl coenzyme A: monoacylglycerol acyltransferase 3, an intestinal specific enzyme implicated in dietary fatabsorption. The Journal of Biological Chemistry 278:13611–13614.

[18] CorringT. 1980. The adaptation of digestive enzymes to the diet: Its physiological significance. Reproduction, Nutrition, Development 20:1217–1235.

[19] DatzmannT,von Helversen O,MayerF. 2010. Evolution of nectarivory in phyllostomid bats (Phyllostomidae Gray, 1825, Chiroptera: Mammalia). BMC Evolutionary Biology 10:1–14.

[20] DumontER,Dávalos LM,GoldbergA,SantanaSE,RexK, VoigtCC. 2012. Morphological innovation, diversification and invasion of a new adaptive zone. Proceedings of the Royal Society B: Biological Sciences 279:1797–1805.

[21] EdgarRC,MyersEW. 2005. PILER: Identification and classification of genomic repeats. Bioinformatics 21: i152–158.

[22] EmmsDM,KellyS. 2019. OrthoFinder: Phylogenetic orthology inference for comparative genomics,Genome Biology 20:238.

[23] FengP,ZhengJ, RossiterSJ,Wang D,ZhaoH. 2014. Massive losses of taste receptor genes in toothed and baleen whales. Genome Biology and Evolution 6:1254–1265.

[24] FlemingTH,Dávalos LM,MelloMAR. 2020. Phyllostomid bats: A unique mammalian radiation. Chicago: University of Chicago Press.

[25] GautamP. 2020. Divergent evolution. In: Vonk J,Shackelford T eds. Encyclopedia of animal cognition and behavior. Cham: Springer International Publishing. 1–8.

[26] GivnishTJ. 2015. Adaptive radiation versus “radiation” and “explosive diversification”: Why conceptual distinctions are fundamental to understanding evolution. New Phytologist 207:297–303.

[27] GlendinningJI. 1994. Is the bitter rejection response always adaptive? Physiology & Behavior 56:1217–1227.

[28] GrantPR. 1999. Ecology and evolution of Darwin’s finches. Princeton: Princeton University Press.

[29] GrantPR,GrantBR. 2011. How and why species multiply: The radiation of Darwin’s finches. Princeton: Princeton University Press.

[30] GuigóR. 1998. Assembling genes from predicted exons in linear time with dynamic programming. Journal of Computational Biology 5:681–702.

[31] Gutiérrez-GuerreroYT,Ibarra-LacletteE,Martínez Del RíoC,Barrera-Redondo J,RebollarEA,OrtegaJ,León-Paniagua L,UrrutiaA,Aguirre-PlanterE,Eguiarte LE. 2020. Genomic consequences of dietary diversification and parallel evolution due to nectarivory in leaf-nosed bats. Gigascience 9: giaa059.

[32] GutierrezE,SchottRK, PrestonMW,Loureiro LO,LimBK,ChangBS. 2018. The role of ecological factors in shaping bat cone opsin evolution. Proceedings of the Royal Society B: Biological Sciences 285:20172835.

[33] HaasBJ,Salzberg SL,ZhuW,PerteaM,AllenJE, OrvisJ,White O,BuellCR,WortmanJR. 2008. Automated eukaryotic gene structure annotation using EVidenceModeler and the program to assemble spliced alignments. Genome Biology 9: R7.

[34] HallgrenJ,Tsirigos KD,PedersenMD,Almagro ArmenterosJJ,Marcatili P,NielsenH,KroghA,WintherO. 2022. DeepTMHMM predicts alpha and beta transmembrane proteins using deep neural networks. BioRxiv:2022.04.08.487609.

[35] HanL,QuQ, AydinD,Panova O,RobertsonMJ,XuY,DrorRO, SkiniotisG,Feng L. 2022. Structure and mechanism of the SGLT family of glucose transporters. Nature 601:274–279.

[36] HaoX,JiaoH, ZouD,LiQ, YuanX,Liao W,JiangP,ZhaoH. 2023a. Evolution of bitter receptor genes and ontogenetic dietary shift in a frog. Proceedings of the National Academy of Sciences of the United States of America 120: e2218183120.

[37] HaoX,LuQ, ZhaoH. 2023b. A molecular phylogeny for all 21 families within Chiroptera (bats). Integrative Zoology. https://doi.org/10.1111/1749-4877.12772. Online ahead of print.

[38] HayakawaT,Suzuki-Hashido N,MatsuiA,GoY. 2014. Frequent expansions of the bitter taste receptor gene repertoire during evolution of mammals in the Euarchontoglires clade. Molecular Biology and Evolution 31:2018–2031.

[39] HoangDT,Chernomor O,von HaeselerA,MinhBQ,VinhLS. 2017. UFBoot2: Improving the Ultrafast Bootstrap approximation. Molecular Biology and Evolution 35:518–522.

[40] HongW,ZhaoH. 2014. Vampire bats exhibit evolutionary reduction of bitter taste receptor genes common to other bats. Proceedings of the Royal Society B: Biological Sciences 281:20141079.

[41] JebbD,HuangZ, PippelM,Hughes GM,LavrichenkoK,DevannaP,WinklerS, JermiinLS,Skirmuntt EC,KatzourakisA,Burkitt-GrayL,RayDA, SullivanKAM,Roscito JG,KirilenkoBM,DávalosLM,Corthals AP,PowerML,JonesG,RansomeRD, DechmannDKN,Locatelli AG,PuechmailleSJ,FedrigoO,JarvisED, HillerM,Vernes SC,MyersEW,TeelingEC. 2020. Six reference-quality genomes reveal evolution of bat adaptations. Nature 583:578–584.

[42] JiaoH,WangQ, WangB-J,Li K,LövyM,NevoE,LiQ, SuW,JiangP, ZhaoH. 2021. Local adaptation of bitter taste and ecological speciation in a wild mammal. Molecular Biology and Evolution 38:4562–4572.

[43] JiaoH,WangY, ZhangL,Jiang P,ZhaoH. 2018. Lineage-specific duplication and adaptive evolution of bitter taste receptor genes in bats. Molecular Ecology 27:4475–4488.

[44] JiaoH,ZhangL, XieH-W,Simmons NB,LiuH,ZhaoH. 2019. Trehalase gene as a molecular signature of dietary diversification in mammals. Molecular Biology and Evolution 36:2171–2183.

[45] KanehisaM,GotoS. 2000. KEGG: Kyoto encyclopedia of genes and genomes. Nucleic Acids Research 28:27–30.

[46] KarasovWH,DouglasAE. 2013. Comparative digestive physiology. Comprehensive Physiology 3:741.

[47] KimD,PerteaG, TrapnellC,Pimentel H,KelleyR,SalzbergSL. 2013. TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions. Genome Biology 14: R36.

[48] KoblmüllerS,Schliewen UK,DuftnerN,SefcKM,KatongoC, SturmbauerC. 2008. Age and spread of the haplochromine cichlid fishes in Africa. Molecular Phylogenetics and Evolution 49:153–169.

[49] KorfI. 2004. Gene finding in novel genomes. BMC Bioinformatics 5:59.

[50] KriesK,BarrosMA, DuytschaeverG,OrkinJD,JaniakMC, PessoaDM,Melin AD. 2018. Colour vision variation in leaf-nosed bats (Phyllostomidae): Links to cave roosting and dietary specialization. Molecular Ecology 27:3627–3640.

[51] KuokkanenM,Kokkonen J,EnattahNS,Ylisaukko-OjaT,KomuH, VariloT,Peltonen L,SavilahtiE,JarvelaI. 2006. Mutations in the translated region of the lactase gene (LCT) underlie congenital lactase deficiency. American Journal of Human Genetics 78:339–344.

[52] LamichhaneyS,Berglund J,AlménMS,MaqboolK,Grabherr M,Martinez-BarrioA,PromerováM,Rubin C-J,WangC,ZamaniN. 2015. Evolution of Darwin’s finches and their beaks revealed by genome sequencing. Nature 518:371–375.

[53] LiL,ChiH, LiuH,XiaY, IrwinDM,Zhang S,LiuY. 2018. Retention and losses of ultraviolet-sensitive visual pigments in bats. Scientific Reports 8:11933.

[54] LinharesBS,RibeiroSP, de FreitasRMP,PugaL,SartoriSSR, FreitasMB. 2021. Aspects regarding renal morphophysiology of fruit-eating and vampire bats. Zoology 144:125861.

[55] LöytynojaA. 2014. Phylogeny-aware alignment with PRANK. Methods in Molecular Biology 1079:155–170.

[56] LuQ,JiaoH, WangY,Norbu N,ZhaoH. 2021. Molecular evolution and deorphanization of bitter taste receptors in a vampire bat. Integrative Zoology 16:659–669.

[57] LuoR,LiuB, XieY,LiZ, HuangW,Yuan J,HeG,ChenY,PanQ, LiuY,TangJ, WuG,ZhangH, ShiY,LiuY, YuC,WangB, LuY,HanC, CheungDW,Yiu SM,PengS,XiaoqianZ,LiuG, LiaoX,Li Y,YangH,WangJ,LamTW, WangJ. 2012. SOAPdenovo2: An empirically improved memory-efficient short-read de novo assembler. Gigascience 1:18.

[58] MajorosWH,PerteaM, SalzbergSL. 2004. TigrScan and GlimmerHMM: Two open source ab initio eukaryotic gene-finders. Bioinformatics 20:2878–2879.

[59] MarçaisG,Kingsford C. 2011. A fast, lock-free approach for efficient parallel counting of occurrences of k-mers. Bioinformatics 27:764–770.

[60] MarínI. 2015. Origin and diversification of Meprin proteases. PLoS One 10: e0135924.

[61] MeyerhofW,BatramC, KuhnC,Brockhoff A,ChudobaE,BufeB,Appendino G,BehrensM. 2009. The molecular receptive ranges of human TAS2R bitter taste receptors. Chemical Senses 35:157–170.

[62] MinhBQ,SchmidtHA, ChernomorO,Schrempf D,WoodhamsMD,von HaeselerA,LanfearR. 2020. IQ-TREE 2: New models and efficient methods for phylogenetic inference in the genomic era. Molecular Biology and Evolution 37:1530–1534.

[63] PotterJHT,DaviesKTJ, YoheLR,Sanchez MKR,RengifoEM,StruebigM,WarrenK, TsagkogeorgaG,LimBK,Dos Reis M,DávalosLM,RossiterSJ. 2021. Dietary diversification and specialization in Neotropical bats facilitated by early molecular evolution. Molecular Biology and Evolution 38:3864–3883.

[64] PriceAL,JonesNC, PevznerPA. 2005. De novo identification of repeat families in large genomes. Bioinformatics 21: i351–i358.

[65] PriceER,BrunA, FasuloV,Karasov WH,Caviedes-VidalE. 2013. Intestinal perfusion indicates high reliance on paracellular nutrient absorption in an insectivorous bat Tadarida brasiliensis. Comparative Biochemistry and Physiology. Part A, Molecular & Integrative Physiology 164:351–355.

[66] PriceER,BrunA, Gontero-FourcadeM,Fernández-MarinoneG,Cruz-NetoAP,KarasovWH, Caviedes-VidalE. 2015. Intestinal water absorption varies with expected dietary water load among bats but does not drive paracellular nutrient absorption. Physiological and Biochemical Zoology 88:680–684.

[67] RhieA,WalenzBP, KorenS,Phillippy AM. 2020. Merqury: Reference-free quality, completeness, and phasing assessment for genome assemblies. Genome Biology 21:245.

[68] Rodriguez-PeñaN,Price ER,Caviedes-VidalE,Flores-OrtizCM,KarasovWH. 2016. Intestinal paracellular absorption is necessary to support the sugar oxidation cascade in nectarivorous bats. The Journal of Experimental Biology 219:779–782.

[69] RundleHD,NosilP. 2005. Ecological speciation. Ecology Letters 8:336–352.

[70] SchluterD. 2000. The ecology of adaptive radiation. Oxford: Oxford University Press.

[71] SchluterD. 2001. Ecology and the origin of species. Trends in Ecology & Evolution 16:372–380.

[72] SchondubeJE,Herrera-M LG,Martínez del RioC. 2001. Diet and the evolution of digestion and renal function in phyllostomid bats. Zoology 104:59–73.

[73] SchwenkRW,Holloway GP,LuikenJJ,BonenA,GlatzJF. 2010. Fatty acid transport across the cell membrane: regulation by fatty acid transporters. Prostaglandins Leukot Essent Fatty Acids 82:149–154.

[74] SeehausenO. 2006. African cichlid fish: A model system in adaptive radiation research. Proceedings of the Royal Society B: Biological Sciences 273:1987–1998.

[75] SimãoFA,Waterhouse RM,IoannidisP,KriventsevaEV,ZdobnovEM. 2015. BUSCO: Assessing genome assembly and annotation completeness with single-copy orthologs. Bioinformatics 31:3210–3212.

[76] SimoesBF,FoleyNM, HughesGM,Zhao H,ZhangS,RossiterSJ,TeelingEC. 2019. As blind as a bat? Opsin phylogenetics illuminates the evolution of color vision in bats. Molecular Biology and Evolution 36:54–68.

[77] SlaterGSC,BirneyE. 2005. Automated generation of heuristics for biological sequence comparison. BMC Bioinformatics 6:31.

[78] SmitAF,HubleyR, GreenP. 2008. RepeatModeler Open-1.0[online]. Available from www.repeatmasker.org. [accessed 13 October 2020].

[79] SpringerMS,MurphyWJ, EizirikE,O’Brien SJ. 2003. Placental mammal diversification and the Cretaceous-Tertiary boundary. Proceedings of the National Academy of Sciences of the United States of America 100:1056–1061.

[80] StankeM,WaackS. 2003. Gene prediction with a hidden Markov model and a new intron submodel. Bioinformatics 19(Suppl 2): ii215–i225.

[81] SunYB. 2017. FasParser: A package for manipulating sequence data. Zoological Research 38:110–112.

[82] TeelingEC,Springer MS,MadsenO,BatesP,O’Brien SJ,MurphyWJ. 2005. A molecular phylogeny for bats illuminates biogeography and the fossil record. Science 307:580–584.

[83] ThorensB. 2015. GLUT2, glucose sensing and glucose homeostasis. Diabetologia 58:221–232.

[84] TianS,ZengJ, JiaoH,Zhang D,ZhangL,LeiCQ,Rossiter SJ,ZhaoH. 2023. Comparative analyses of bat genomes identify distinct evolution of immunity in Old World fruit bats. Science Advances 9: eadd0141.

[85] TrapnellC,Williams BA,PerteaG,MortazaviA,KwanG, van BarenMJ,Salzberg SL,WoldBJ,PachterL. 2010. Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nature Biotechnology 28:511–515.

[86] WangK,TianS, Galindo-GonzálezJ,DávalosLM,Zhang Y,ZhaoH. 2020. Molecular adaptation and convergent evolution of frugivory in Old World and neotropical fruit bats. Molecular Ecology 29:4366–4381.

[87] WeadickCJ,ChangBS. 2012. An improved likelihood ratio test for detecting site-specific functional divergence among clades of protein-coding genes. Molecular Biology and Evolution 29:1297–1300.

[88] WeisenfeldNI,KumarV, ShahP,Church DM,JaffeDB. 2017. Direct determination of diploid genome sequences. Genome Research 27:757–767.

[89] WickhamH. 2016. ggplot2: Elegant eraphics for data analysis. Cham: Springer International Publishing.

[90] WilmanH,Belmaker J,SimpsonJ,de la RosaC,Rivadeneira MM,JetzW. 2014. EltonTraits 1.0: Species-level foraging attributes of the world’s birds and mammals: Ecological Archives E095-178. Ecology 95:2027.

[91] WuJ,JiaoH, SimmonsNB,Lu Q,ZhaoH. 2018. Testing the sensory trade-off hypothesis in New World bats. Proceedings of the Royal Society B: Biological Sciences 285:20181523.

[92] XuZ,WangH. 2007. LTR_FINDER: An efficient tool for the prediction of full-length LTR retrotransposons. Nucleic Acids Research 35: W265–W268.

[93] YangZ. 1998. Likelihood ratio tests for detecting positive selection and application to primate lysozyme evolution. Molecular Biology and Evolution 15:568–573.

[94] YangZ. 2007. PAML 4: Phylogenetic analysis by maximum likelihood. Molecular Biology and Evolution 24:1586–1591.

[95] YangZ,NielsenR. 1998. Synonymous and nonsynonymous rate variation in nuclear genes of mammals. Journal of Molecular Evolution 46:409–418.

[96] YangZ,NielsenR. 2002. Codon-substitution models for detecting molecular adaptation at individual sites along specific lineages. Molecular Biology and Evolution 19:908–917.

[97] YuG,SmithDK, ZhuH,GuanY, LamTT-Y. 2017. ggtree: An r package for visualization and annotation of phylogenetic trees with their covariates and other associated data. Methods in Ecology and Evolution 8:28–36.

[98] YuXH,QianK, JiangN,Zheng XL,CayabyabFS,TangCK. 2014. ABCG5/ABCG8 in cholesterol excretion and atherosclerosis. Clinica Chimica Acta 428:82–88.

[99] YuXJ,ZhengHK, WangJ,Wang W,SuB. 2006. Detecting lineage-specific adaptive evolution of brain-expressed genes in human using rhesus macaque as outgroup. Genomics 88:745–751.

[100] ZhangC,RabieeM, SayyariE,Mirarab S. 2018. ASTRAL-III: Polynomial time species tree reconstruction from partially resolved gene trees. BMC Bioinformatics 19:153.

[101] ZhangJ,NielsenR, YangZ. 2005. Evaluation of an improved branch-site likelihood method for detecting positive selection at the molecular level. Molecular Biology and Evolution 22:2472–2479.

[102] ZhangJ,XuD, NieJ,CaoJ, ZhaiY,Tong D,ShiY. 2014. Monoacylglycerol acyltransferase-2 is a tetrameric enzyme that selectively heterodimerizes with diacylglycerol acyltransferase-1. The Journal of Biological Chemistry 289:10909–10918.

[103] ZhangZ,XiaoJ, WuJ,ZhangH, LiuG,WangX, DaiL. 2012. ParaAT: A parallel tool for constructing multiple protein-coding DNA alignments. Biochemical and Biophysical Research Communications 419:779–781.

[104] ZhouY,ZhouB, PacheL,Chang M,KhodabakhshiAH,TanaseichukO,BennerC, ChandaSK. 2019. Metascape provides a biologist-oriented resource for the analysis of systems-level datasets. Nature Communications 10:1523.

RIGHTS & PERMISSIONS

2024 The Author(s). Journal of Systematics and Evolution published by John Wiley & Sons Australia, Ltd on behalf of Institute of Botany, Chinese Academy of Sciences.