Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. u atra iest fti pdri eeal itdie.I diint hs hyosre clear a of observed microbial impact they gut the Earlier this, studied 2020). to al., al., addition et et In (Kennedy Kennedy microbiome gut diet-driven. 2020). the generally species and al., is insects spider et spider prey the Sheffer the this of between 2017; of correlation microbiome al., diversity gut et bacterial the Rivera gut on ( 2019; species food al., different spider et six Hu only 2020; of diversity diversity bacterial bacterial albofasciata establish gut gut to very the explored. However, order date, was poorly in it 2011). Till is al., spiders and et group inhabiting forests Koch diverse different rain 2002; of most of al., to type this et diversity desert (Dillon This of bacterial relationships gut i.e. gut habitat 2020). their diverse, and of of host al., most role function the are et the and interpret spiders composition (Kennedy the to into) of tissue study important habitats (or liquefied to The onto model resulting of interesting fluid communities. the (EOD). an digestive behaviour microbial spiders again digestion regurgitating makes feeding back oral also and extra The sucking behaviour venom by feeding then injecting nutrients 2019). and by al., extracting prey al., prey arthropods, et important their other their et (Ndava equally most immobilize (Brinker diseases are in generally They various purposes that Spiders for 2016). medicinal from different al, agent and is et bio-control agricultural ecological spiders (Ossamy in 2017) economical, monitoring pests environment al., their insect for et on for species Yang predators indicator natural 2018; and as al., 2018), act usually et that (Michalko arthropods ecosystem are Araneae) (order Spiders fatty as acids, diversity amino bacterial lipid, gut Introduction carbohydrate, spider in driven microbiota diet gut the of gut from involvement the different metabolism. the that quite energy revealed revealed and generally are analysis study acids Wolbachia functional wild Our genus A from laglaizei. endosymbiont Eriovixia collected this, earlier. and species spiders to published mulmeinensis, spider the addition one Cyclosa of In in diversity species, observed gut. bacterial one was their alterations Acinetobacter, in in reproductive level, only genera for genera observed dominant responsible At most was the Prevotella respectively. Phylum. 22 were genus abundant bengalensis of Bacillus the highest Neoscona and presence the and Staphylococcus, as the mulmeinensis Cutibacterium, Proteobacteria revealed Cyclosa Pseudomonas, with gut Analysis in microbiome, of dominated gut studied. Chlamydiae comparison the been and in Deinococcus subfamily and have microbes Actinobacteria inter Araneinae of Araneidae Firmicutes, phyla subfamilies, genera the family the three Moreover, 364 decipher of to and based belonging (2) to families, effort species Gasteracanthinae 145 out spider largest phyla, and Twelve carried The (2) relationship. was digestion. functional Argiopinae oral sequencing their (8), extra and generation for spiders known in next remarkably diversity amplicon bacterial arthropods of 16S groups targeted diverse on most the among are Spiders Abstract 2020 12, August 2 1 Tyagi Kaomud wild Araneidae twelve family in of diversity species microbial spider gut of comparison subfamily Inter ffiito o available not Affiliation India of Survey Zoological , aioarabida Rabidosa 1 neje Tyagi Inderjeet , , aun longinqua Badumna 1 ry Prasad Priya , 1 and , hru eeas eetd h atra hl Bacteriodetes phyla bacterial The detected. also were Thermus 2 als Chandra Kailash , rip bruennichi Argiope aun longinqua Badumna ads laura Pardosa , 1 ads astrigera Pardosa r nw Kneye al., et (Kennedy known are n ia Kumar Vikas and , n bevdta the that observed and , , Nurscia 1 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. lsie lgn h eeae aooyadfauetbe ln ihmtdt l eepoesdfor Data processed Marker were using file 2017) q2-feature- al., metadata QIIME2 et with using (Dhariwal along MicrobiomeAnalyst 132) tables tool, OTUs (version feature web-based These A database and taxonomy analysis. similarity (OTUs). downstream were generated units 99% the reads SILVA The taxonomic chimeric on operational The plugin. into based classifier merging. assigned classified (ver. and were al., de-noising further QIIME2 et reads (Callahan in trimming, were non-chimeric DADA2 reads filtering, by the single quality processed and into were for filtered merged reads QIIME2 These were in species demultiplexing. pipeline using spider 2016) twelve 2019) al., of et reads (Bolyen raw 2019.10) end paired generated generated The the of analyses submission statistical Center the PRJNA638522. and National for ID Kit) Bioinformatics used The BioProject platform. V2 been the HiSeq method. under RP has Illumina sequencing to Portal Hiseq2500 the PE301+8+8+301-v2 reads GenBank (Illumina on raw 2500-RP (NCBI) PE300bp out Hiseq Information carried using Biotechnology the was sequenced for using rRNA was mode 16S diversity. library rapid of microbial Nextra with region of constructed sequencing gene high-throughput qualified targeted for The the gels 98 of agarose at sequencing using min visualized The were 5 products for PCR denaturation V3- The involved of cycle 98 PCR sets (5’- at The primer 806R water. the Milli-Q and ACTCCTACGGGAGGCAGCAG-3’) and using (5’- electrophoresis Tissue amplified 0.5 341F gel & primer, rRNA was each agarose Blood 16S DNA using DNeasy the 25 Qubit checked extracted Total a of GGACTACHVGGGTWTCTAAT-3’). by was The checked using regions was quality hypervariable MINI). extracted DNA the V4 was Alphalmager the and species of BioScience Thermofisher), quantification spider (Cell The (Q32866, of protocol. Fluorometer sample manufacturer’s 2.0 the pooled following each (Qiagen) of Kit DNA SMZ25 total NIKON The a sequencing species on and non-protected Prasad amplification 1982). Priya and al., Isolation, by were non-endangered et done DNA species were (Tikader was each keys study specimens starvation, available of this these using of specimens in of microscope hours ten identification used stereo 8 washing, taxonomic spiders After The After The 1). contamination. isolation. (Table vial. the DNA 4 empty trap. remove in for pan separate to stored yellow a pooled solution and and in PBS alcohol net, 100% individually with sweep in kept picking, thrice transferred hand and are sorted methods: specimen India, are each collection of following specimens State the Bengal the West by Kolkata, All field from the species spider from the collected all of species, specimens one collected except have we study, this In collection gut Sample the explore (Araneinae, to weavers). Methods subfamilies made (orb-web and spider been Araneidae three Materials has family from the (NGS) species of Sequencing 12 Gasteracanthinae) in Gen and metabolism Next Argiopinae functional through predicted effort the large and a microbiome study, present the In i.e. of spider symbiont wasp unknown the (Dominant in Tenericutes DUSA and to like microbes affiliated microbes the symbiont fluid, between bacterial antimicrobial Rivera interaction of the 2019). presence revealed sp. al., the clearly Staphylococcus et species in obtained (HU spider i.e. results style the the hosts The feeding with their of samples. peculiar associated bacteria excreta their microbiome and to gut the swaps due including studied the arthropods dominant, 2017 Thus, other most al., from the et Bacteroidetes. different was be Proterobacteria and may phylum Acidobacteria spiders the Firmicutes, that observed Actinobacteria, species Tenericutes, spider three of studies ° ,anaigfr4 t53 at s 45 for annealing C, μ fTqDAplmrs Tkr) 1 (Takara), polymerase DNA Taq of l yls bianchoria Cyclosa neceasmlswsosre.See ta. 00rpre h rsneo novel of presence the reported 2020 al., et Sheffer observed. was samples excreta in , ° ,adeogto t72 at elongation and C, hc a olce rmAdr rds tt.Teseie were specimen The state. Pradesh Andhra from collected was which , rip bruennichi Argiope μ fmxuewspeae o h C,icuig1 including PCR, the for prepared was mixture of L 2 μ fdTs 2.5 dNTPs, of l ° o 5s n nletnino i t72 at min 7 of extension final and s, 45 for C See ta. 2020) al., et (Sheffer ) ° .Sbeunl,ec pcmnwr washed were specimen each Subsequently, C. rip bruennichi Argiope tpyoocsaureus Staphylococcus aioarabida Rabidosa μ f10 of l ° olwdb 5cce o 0s 30 for cycles 35 by followed C × ue,5 go template, of ng 50 buffer, . hc hynmdas named they which , yaayigbody analysing by , nbd wp and swaps body in μ of l ° C. Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. iia rnswr bevdi ufml ripnewt osdrbecagsi h bnac forder of abundance the in changes considerable with Argiopinae subfamily in Bacilliales observed were trends Similar cus contribute (Class: majorly that orders the nobacteria them among were dataset, Firmicutes), Araneinae current subfamily the in diversity in Gammaproterobacteria bacterial detected total the were the orders this, to 81 to i.e. of addition Araneinae, total In subfamily A the Firmicutes. of and species one Proteobacteria 1b) in (Figure only i.e. observed phyla was two Chlamydiae subfamily phylum only the of of species constituted two predominantly from comes in Actinobacteria represented phylum (1 the the Bacteroidetes of all i.e. 14%), contribution of Araneinae, to guts major 1 the The from in ranges detected Araneidae. (abundance was Deinococcus Actinobacteria 35%, and second like to 10%), The taxa 14 to bacterial Gasteracanthinae. the from Other in range revealed 75% dataset species. abundances current and Araneidae the spider an Araneinae, in in with family dominated 55% Firmicutes was the Argiopinae, phylum, Proteobacteria for phylum dominant of 49% The of subfamilies families. abundance 145 relative three and with phyla in 1a). total other 22 diversity (Figure total A the for of Gasteracanthinae microbial On Gasteracanthinae. presence and 678 with of Argiopinae and Araneidae. 275 subfamilies, classification after Argiopinae family and two Taxonomic by Argiopinae analysis of for shared with subfamilies downstream 474 were OTUs the OTUs for Araneinae, 341 features. all 35 shared for OTUs) left of by Araneinae per (131 unique were shared subfamily abundance reads were OTUs were the low 89763 OTUs hand, OTUs 1175 and 3891 of 273 OTUs) of (average that (20 while total variance revealed 106136 Gasteracanthinae, low A analysis merging, to OTUs), Venn 69573 i.e. (4641 The OTUs. steps from singletons 5967 pre-processing of ranges removal after into which the identified assigned were etc. (V3-V4) and removal, rRNA sample) 16S chimera of filtering, sequences quality 1077166 the of for total software A 2014) richness and al., composition et community (Parks Bacterial STAMP the to loaded Results was metadata the visualization. with further along file through generated characterized were pathways 2019) metabolic The predicted of The tree. ncyclopedia heat 2020) pathways. the for team metabolic of used core picturing functional was (R the beta (http://jvenn.toulouse.inra.fr) and R-based for the 2014) used the Curtis by analyzing was al., while Bray 2017) for diagram, et for al., Venn used (Bardou et the were method jvenn of measure statistical tool construction distance online based the UniFrac PERMANOVA an unweighted Further, The for Scaling diversity. method Sum Shannon measures. based Total Chao1, linkage diversity with observed, Ward’s a step using as methods resulting normalization Simpson statistical in and T-test/ANOVA and with data range analysed the quantile was Alpha-diversity rarefy inter The recovered also default abundance were have a low we OTUs on with Further, based 1326 features option. filtered of variance (TSS) OTUs. further total were low 1175 OTUs A and of These total 10 singletons. analysis. prevalence the diversity with removing bacterial after features OTUs for 5967 used of was out module (MDP) Profiling R hru)(iue2a). (Figure Thermus) cntuto of econstruction 20-22 Alphaproteobacteria (Class: aaaeadacrc vlae yteNaetSqecdTxnIdx(SI aus The values. (NSTI) Index Taxon Sequenced Nearest the by evaluated accuracy and database ), Micrococcales Flavobacteriales yls mulmeinensis Cyclosa yls spirifera Cyclosa G Bacilli nsand enes U hlm Proteobacteria), Phylum: , ) nobserved Bacteriodales , hru 2t % eedtce ntegto lvnsie pce ftefamily the of species spider eleven of gut the in detected were 6%) to (2 Thermus , hlm rtoatra,Temls(ls:Diooc,Pyu:Deinococ- Phylum: Deinococci, (Class: Thermales Proteobacteria), Phylum: , Corynebacteriales G and nms(EG Knhs ta,20;Knhs ta. 09 Kanehisa, 2019; al., et Kanehisa 2000; al, et (Kanehisa (KEGG) enomes and Bacteriodales S roii laglazei Eriovixia ae Lnil ta. 08 PCUt)wsue o rdcigthe predicting for used was (PICRUSt2) 2018) al., et (Langille tates Aaene.Temcoim ftespecies the of microbiome The (Araneinae). (Class: and (Class: Bacteroidea Propionibacteriales Bacilliales 3 hl h hlmBceidtswsms strongly most was Bacteriodetes phylum the while , Bacteroidea, Pseudomonadales and ) P and hylogenetic Rickettsiales Lactobacilliales hlm Bacteriodetes), Phylum: (Class: I Actinobacteria, vsiainof nvestigation , and eaoe otae(Foster software Metacoder (Class: Enterobacteriales (Class: rnu mitificus Araneus esoabengalensis Neoscona Alphaproteobacteria Bacilli, Phylum: C Rickettsiales ommunities K Phylum: yoto (Class: Acti- was E Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. h ld Iwt osdrbeaudneo enccu hru otisfieseis( species five contains Thermus Deinococcus of abundance considerable , a with II clade The Actinobacteria. phylum ( Araneinae while of phylogeny, ( species Argiopinae and Two of species species. abundance one three bacterial and other of the type than similar a higher of bacterial possesses was species of also the type IB in similar species subclade except a phylogeny The possesses and the IA Bacteriodetes. phylum subclade in the The of except B: abundance ( and phylogeny A Argiopinae subclades and two subfamily into abundance of divided further species was I one Clade Gasteracanthinae+ subfamily of subfamily Deinococus members of phylum the species pulchella Two of one all in abundance contain observed 4). relative was (Figure II Thermus the Araneidae Deinococus by phylum family ( of discriminated the Argiopinae abdundance of be negligible with subfamilies can I three and Clade of formed species were the clades of arrangement Gasteracanthinae phylogenetic and based and Argiopinae unifrac of unweighted richness community The the community than the higher in (Chao1), 3a-d). was observed 3–19 (Figures Araneinae changes of subfamily (p The range non-significant the were the for (Simpson). subfamilies in 0.39–0.68 three lies and of Araneidae richness (Shannon) family 0.65–1.26 the (Observed), of 3–16 Gasteracanthinae) the Argiopinae, for (Araneinae, for used ilies measures were diversity Simpson unifrac and unweighted Shannon and Observed, Chao1, of richness, ments community the genera decipher The To spiders. of dataset genus current endosymbiont the the this, in to observed addition were genera Araneinae, 364 subfamily , level, the genus of the species At one only Araneinae, in Pseudomon- subfamily reflected was the and Prevotellaceae of mulmeinensis Burkholderiaceae family species the diver- one families, of 47% in Two tribution Propionibacteriaceae, remaining observed The Staphylococcaceae, not etc. were Prevotellaceae, spiders. adaceae, including Thermaceae of families, subfamilies Micrococcaceae, other three Pseudomonadaceae Corynebacteriaceae, by all (9%), contributed in Bacillaceae was observed (13%), sity were Enterobacteriaceae (5%), (20%), Burkholderiaceae Moraxellaceae (6%), level, family the sub- At to similar 2c). quite in Deinococcus (Figure also observed Phylum: were was Deinococci, abdundance Gasteracanthinae order subfamily in of micutes), differences case considerable in Few (Class: abundance Araneinae. order family of pattern The 2b). (Figure ) h ryCri ae MSodnto ltas ugse iia yeo eut sidctdb un- by indicated as results of type similar a suggested Cyanobacteria. also and plot bacterial Chalamydiae of ordination Phylum type NMDS similar of have based diversity and the Bray-Curtis B in and The A changes subclades few two except into phylogeny divided sub and further diversity clade This Firmicutes. lum species N Pseudomonas . bengalensis Gammaproterobacteria rip pulchella Argiope Siphingomonadales +toseiso ufml rnia ( Araneinae subfamily of species two )+ ytohr cicatrosa Cyrtrophora Fgr 1b) (Figure Wolbachia , Cutibacterium , atrcnh kuhli Gasteracantha yls spirifera Cyclosa Agoia)i ld Iwstefis rnhotdet h ihaudneo phy- of abundance high the to due out branch first the was II Clade in (Argiopinae) 2 )wsfudol in only found was %) (28 Prevotella (Class: hlm Proteobacteria), Phylum: , ytpoacicatrosa Cyrtophora β- , iest nlsswscridott netgt h omnt richness community the investigate to out carried was analysis diversity Staphylococcus Thermus), +altemmeso ufml rnia nti ld hl clade while clade this in Araneinae subfamily of members the all )+ 6% a bevdol noespecies, one in only observed was (60%) yls bianchoria Cyclosa α- Alphaproteobacteria a rnhdotfo hssbld u otemr bnac of abundance more the to due subclade this from out branched was , and atrcnh hasselti Gasteracantha β- β- iest nlsswr are u.Tedvriymeasure- diversity The out. carried were analyses diversity Bacteriodales iest nlss The analysis. diversity esoanautica Neoscona and , yls mulmeinensis Cyclosa > roii laglaizei Eriovixia nsbld Bhv iia yeo atra diversity bacterial of type similar have IB subclade in ) 4 .5.Mroe,temcoim omnt richness community microbiome the Moreover, 0.05). Bacillus nwihteaudneo hlmProteobacteria phylum of abundance the which in , hlm rtoatra,Temls(Class: Thermales Proteobacteria), Phylum: , Lactobacilliales (Class: osiuetemjrgtmcoim.In microbiome. gut major the constitute and roii excelsa Eriovixia and α- Bacteroidea, (Araneinae). rip pulchella Argiope iest nlss hl Bray-Curtis while analysis, diversity rnu mitificus Araneus .bengalensis N. α- hc iegddet increased to due diverged which iest o he pdrsubfam- spider three for diversity (Class: yls mulmeinensis Cyclosa hlm Bacteriodetes) Phylum: Bacilli, and Fgr 4). (Figure ) yls spirifera Cyclosa .Tebac of branch The ). h ao con- major The . esoanautica Neoscona Enterobacteriales hlm Fir- Phylum: Acinetobacter Thermus. Cyclosa Argiope while ) Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 06.Oeseiso h genus 2011). the Foster, of species One 2016). Bacillus 2018). al., except et non-pathogenic, Voirol usually (Paniagua their earlier of reported ratio as sex the altering genera by The genus 2020) 2019). Wang, endosymbiont al., & The et (Bi (Hu 2017). activities reproduction al., transmitting host and et proliferative strains (Xia their few activity al., for enzyme a known et Moreover, catalase genus (Briones-Roblero or 2010). the weevils dismutase al., of superoxide like et members insects Furthermore, (Anand other silkmoth Ham- 2007). in and & 2016; earlier genus 2005), Dexter al., reported the Van with al., et of as 2013; et line Ruokolainen Morgan, 2018) (Delalibera in & Cerambycidae 2016; al., (Engel is 2017), et al., insecticides gut Itoh et based 2019; spiders’ Chen organophosphate Boopathy, of the 2019; genera, degradation The al., in and et Proteobacteria metabolism 2017). (Hu phylum al., arthropods 2020). et the other mer al. of of et representatives (Young studies larvae of previous grub abundance grass of high digestion The to due modified was which microbiome mulmeinensis genus the this, to observed was it ( 0.055), species = two Stress the (NMDS from obtained matrix samples distance gut on i.e. Based species two that measures. diversity unifrac weighted a bevdol noesie pce i.e. species spider one and in were only Proteobacteria observed phylum was Bacteroidetes in Actinobacteria, 22 genera Firmicutes, of dominant Proteobacteria, obacter presence func- were the the predicted phyla reveals Moreover, dominant their analysis The Argiopinae and amplicon etc. Araneinae, microbiome rRNA families. 16S i.e. gut 145 Targeted subfamilies and the spider phyla Araneidae. map three family to under of species made Gasteracanthinae 12 been in and has activities metabolic effort in largest pathways subfamily the tional between study, present included the bacteria In biosynthesis. gut acid of amino pathways degradation, and metabolic Discussion metabolism the carbohydrate were Whereas, involved Argiopinae amino bacteria and in the metabolism. Araneinae involved than other bacteria abundances relative gut the higher The showed in metabolism of metabolism. carbohydrate nucleotide metabolism and were and degradation, metabolism Gasteracanthinae metabolism, and acid and biosynthesis energy acids Araneinae vitamins, amino and subfamily metabolism, cofactors between acids included fatty bacteria acids metabolism, gut carbohydrate sub- amino of three Isolucine), pathways all metabolic Lysine, 6). The among (Arginine, (Figure biosynthesis shared metabolism acid acids were Argiopinae fatty pathways amino while and metabolic metabolism, Gasteracanthinae, degradation Seven carbohydrate with Araneinae pathways subfamily i.e. Gasteracanthinae. the 77 families, with that and Comparative pathways predict Argiopinae families 16 Gastercanthinae. with three shared pathways Argiopinae, these metabolic Araneinae, of abundance 11 metabolic i.e. gene shared in subfamilies, functional differences three metabolic and of of resemblances studies the microbiome decipher the to of used profiles was approach ( computational Argiopinae PICRUSt2 family The of species ( one Argiopinae with of resemblance species close one with ( resemblance Araneinae close in were canthinae Bacillus ihcnieal bnac hogottesie pce,wiea noybotgenus endosymbiont an while species, spider the throughout abundance considerable with eersosbefrteezm n hshlpd eaoim(irse l,21;Fvr tal., et Favaro 2018; al., et (Nigris metabolism phospholipids and enzyme the for responsible were esoanautica Neoscona Staphylococcus Pseudomonas pyu imcts and Firmicutes) (phylum hc a edet ietdgasgu ave hsrpr sas bevdi ua gut human in observed also is report This larvae. grub grass digested to due be may which Prevotella rnu mitificus Araneus hwdatv naoimtwrsetmptoei ug Idrgnh tal., et (Indiragandhi fungi entomopathogenic towards antagonism active showed and pyu atriee)wsms bnati n pce fsie i.e. spider of species one in abundant most was Bacteroidetes) (phylum and tpyoocsaureus Staphylococcus Acinetobacter Bacillus Bacillus .bengalensis N. Cutibacterium and , a lob novdi h rtaeatvt ntegto spiders of gut the in activity protease the in involved be also may .thuringiensis, B. roii laglzei Eriovixia Enterobacter atrcnh kuhli Gasteracantha roii laglaizei Eriovixia and Staphylococcus .Rs ftemmeso ufml rnia eein were Araneinae subfamily of members the of Rest ). Pseudomonas H ta. 03.Frhroe h pce ftegenus the of species the Furthermore, 2013). al., et (He 5 Atnbcei)wr h otabundant most the were (Actinobacteria) yrpoacicatrosa Cytrophora eersosbefrezm eaoimthrough metabolism enzyme for responsible were scmol sda nisc ahgn(on& (Joan pathogen insect an as used commonly is eei ls eebac ihec te.The other. each with resemblance close in were usqety eealike genera Subsequently, . otisGa-oiiebcei hc are which bacteria Gram-positive contains eekonfrterrl neeg,lipid energy, in role their for known were and rip pulchella Argiope Acinetobacter G . hassleti Fgr 5). (Figure ) fsbaiyGastera- subfamily of ) , Pseudomonas n w pce of species two and ) Staphylococcus Wolbachia . naddition In Wolbachia Cyclosa , Enter- is Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. adu . aite . suie . jme,C,Kop .(04.jen nitrcieVn diagram Venn interactive an jvenn: (2014). C. Klopp. C., Djemiel, Escudi´e, F., J., viewer. digestion. of on Mariette, gut impact P., their the Bardou, C.J., and from Geoffrey, starch bacteria T., and of Raghuraman pectin characterization P.T., xylan, and Vasan, cellulose, Isolation D.I.G., Prabhu, (2010). S.G., S.E. Sankar, Vendan, S.J., Vennison, A.A.P., under Anand, GenBank (NCBI) Information Biotechnology for References Center National PRJNA638522. ID: to BioProject submitted the were reads raw acquisition. The Funding and project the Accessibility supervised Manuscript Data VK analysis, and Bioinformatics Conceptualiza- in KC involved in VK involved Editing. and VK and IT and KT, writing KT Software; isolation; Methodology, DNA Curation, and Data identification tion, collection, specimen in involved Ministry PP Kolkata, language India, for of Survey Manchester Contributions Zoological of of Author University Change. funding Breitling, Climate core Rainer by and Forest Prof. supported Environment financially thank and of We support is constant study providing studies. The for the Kolkata, editing. India, out of carry Survey get to Zoological to Director, facilities the microbiome to spiders. gut thankful in their are microbiome Authors for gut mapped the be of to role needs detected the Acknowledgement(s) as species spider and spiders spider driven diversity of of diet of gut number the the picture also in larger from clear diversity study a different bacterial a Our study, high quite the present habitat. are microbiota Given the and diversity spider earlier. in host bacterial published the like gut as that factors diversity spider environmental bacterial suggesting collected gut and field species, behavior the these feeding that of the revealed relationship by phylogenetic influenced the be may with correlated (V3- not rRNA is genus for 16S the spiders unifrac NGS unweighted of of the the screening species using the another spiders for in prompts of reported species been 12 previously of genus diversity The bacterial Wolbachia the data. mapped sequence we V4) environment study, natural our the in In from data spiders baseline actual of as of gut used terms the be Conclusion in specimens. in can significance reared present it great laboratory diversity Moreover, of of bacterial al., metabolism. that be the et and in will distinguish (Hu role field to earlier potential the future indicated from their the collected and as diversity specimens detoxification activities bacterial this, and metabolic gut to different digestion for addition enemies, responsible influenced In natural turn greatly in 2019). be against which can self-defence host microbiota and to gut behaviour the related feeding that habitat, indicated, like spiders factors in by assessment diversity bacterial case in gut ( the histolysis The beetle In the bark 2017). for in also. al., stages responsible developmental analysis et being adult PICRUSt2 as (Briones-Roblero the by reported in ) indicated histogenesis been and has as stages Prevotella 2019) larval genus the al., the et Bacteroidetes, phylum Martin the against genus (Rocha of The metabolites Actinobac- metabolism bioactive of 2009). acid and Members (Kaltenpoth, fatty synthesis bacteria spider. nutrient the pathogenic of enzymes, the gut processing of the invasion food in the producing taxon for abundant known most are third teria the was Actinobacteria phylum The M Bioinf. BMC eetdfrtefis iein time first the for detected 5 293–300. 15, , β- iest nlssceryidctsta h u irboacmoiini spiders in composition microbiota gut the that indicates clearly analysis diversity Prevotella Wolbachia roii laglaizei Eriovixia a bevdol in only observed was n t oei eaoim h edormbsdon based dendrogram The metabolism. in role its and 6 roii,E cavaleriei E. Eriovixia, hsapartoatraedsmin had endosymbiont alphaproteobacteria This . Cyclosamulmeinensis netSci., Insect J Cutibacterium 0 107. 10, obxmori Bombyx edotnsrhizophagus Dendroctonus Wn ta,21) It 2010). al, et (Wang a ersosbefor responsible be may n endosymbiont and , htdegrade that Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. niaadi . nnhm . ahia,M,Pogzai . i,GH,Srvnn ..&Sa, & V.S. Saravanan, and nutrition. insect prothiofos-susceptible G.H., host prothiofos-resistant, and Kim, of fungi gut S., entomopathogenic larval moth, diamondback with Poonguzhali, of associated populations M., bacteria field-caught Cultivable Madhaiyan, (2007). R., species: spiders. T. spider Anandham, of three style of P., feeding microbiota Indiragandhi, gut special the the into to insight corresponding Taking found (2019). 8146–8156. was Y. symbiont Peng, characteristic Y., Yun, No L., Zhang, community G., bacterial Hu, gut the of analysis diversity and Composition techniques. gut armyworm, (2013). resident Oriental M. a Li, the Z., lack of Zhang, Caterpillars X., Nan, (2017). C., N. He, Fierer, S.P., Jaffe, W., Hallwachs, manip- microbiome. D.H., and B. Janzen, visualization Baldan, for T.J., package Hammer, P., R data. An diversity Pastore, taxonomic Metacoder: A., community (2017). of Squartini, N.J. ulation Grunwald E., T.J., Sharpton Baldan, Z.S.L., Foster S., spectrometry. by Nigris, mass produced tandem I.M., accurate lipopeptides Gangi, with of Di Characterization function. S., and (2016). structure Bogialli, in insects–diversity G., of Favaro, locust microbiota a gut of The components (2013). Rev., produce N.A. Microbiol. bacteria Moran, gut & P., note: Engel, A web- (2002). a A.K. - pheromone. Charnley, ”MicrobiomeAnalyst cohesion C.T., Vennard, (2017). data”, R.J., J. microbiome Dillon, of Xia. meta-analysis L.B., and Agellon, visual 45,180–188. I., statistical, , King, comprehensive for S., tool Habib, microorgan- based J., gut Chong, of A., activities Dhariwal, cellulolytic in Contrasts (2005). frontalis of borer, Dendroctonus K.F. activity wood and Raffa, the Biodiversity J., between herbivore (2016). isms Handelsman insect Y. the Jr, Shao, of I. B., history Wilhelm, Delalibera life X., the Lu, across S., beet- microbiota Hu, C., gut bark Sun, the High- T., DADA2: the Beng-Soon, (2016) S.P. B., of data, Holmes, Chen, amplicon A.J., microbiota Illumina Johnson, from A.W., bacterial inference Han, sample M.J., gut resolution Rosen, P.J., the McMurdie B.J., of Callahan, dynamics and Structure le, Rivera-Ordu˜na, (2017). Z´u˜niga, L.V., G. Soto-Robles, F.N., R., Hern´andez-Garc´ıa, Gonzalez-Escobedo, C.I., De- J.A., Briones-Roblero, (2017). of F. gut the Orduna, from Rivera- isolated G., yeasts Scolytinae). and Zuniga, bacteria microbiome: J.A., of Santiago-Cruz capacities the gradation R., and RodrIguez-Diaz, symbionts, C.I., Host, Briones-Roblero (2019). extensible J.F. and interaction. Salles, tripartite scalable L.W., missing interactive, Beukeboom, the Reproducible, M.C., Fontaine, (2019). P., al., Brinker, 2. et QIIME M.R., using science Dillon, data J.R., microbiome hosts. insect Rideout, of E., behavior the Bolyen, on Wolbachia endosymbiont the of effect The (2020). science Y.F. Wang, & J., Bi, edotnsrhizophagus Dendroctonus 1-13. , .Isc Sci. Insect J. oi Microbiol. Folia rc al cd Sci., Acad. Natl. Proc. 7 699-735. 37, .Ap.Microbiol., Appl. J. Clotr:Curculionidae). (Coleoptera: 3 165. 13, , yhmaseparata Mythimna 2 1–9. 62, , Cruinde cltne costerlf stages. life their across Scolytinae) (Curculionidae: aed vestita Saperda rnsMicrobiol. Trends 1,9641–9646. 114, a.Biotechnol., Nat. ai omnMs Sp Mass Commun Rapid 2 759-763. 92, Clotr:eabcde,adtebr beetles, bark the and (Coleoptera:Cerambycidae), ltlaxylostella Plutella eemndb utr-needn n culture-dependent and culture-independent by determined , LSCmu.Biol. Comput. PLoS .Ap.Microbiol., Appl. J. 7 480–488. 27, , 7 nio Entomol., Environ ailslicheniformis Bacillus a.Methods Nat. 7 852–857. 37, pdpealittoralis Spodoptera n hi oeta o,atgns towards antagonism for, potential their and edotnsrhizophagus Dendroctonus 0 2237–2252. 30, , 3 e1005404. 13, , 3 581–583. 13, , 0,2664–2675. 103, 4 541–547. 34, sn iudchromatography liquid using lSone PloS . c.Rep. Sci. uli cd Res. Acids Nucleic 2 e0175470. 12, , (Curculionidae: cl Evol., Ecol. p pini Ips ,29505. 6, , FEMS Insect and 9: Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. uklie,L,Ioe,S,Mkoe,H,Hnk,I 21) avlgot aei soitdwt the with associated is rate growth butterfly. Larval fritillary glanville (2016). the I. in Hanski, microbiota (2019). gut H., the C. Makkonen, of Lacroix, S., composition C., Ikonen, Braegger, L., A., Ruokolainen, Geirnaert, E., 1-14. Voney, 95, L., , Krych, C., of Schwab, Colonization of V.N., Community Microbial Martin, the at Rocha Look First Arkansas, A Searcy, (2017). in A. Spider Hug, Statistical R., for Foundation Stork, R P., Rivera, computing. statistical http://www.R-project.org/. for URL environment and Austria. language Vienna, A and R: Computing, taxonomic (2020). of Team. analysis Core statistical R STAMP: (2014). R.G. Beiko P., profiles, symbionts Hugenholtz functional host, Bacterial G.W., the Tyson (2018). on D.H., N.E. impact Parks Fatouros, and & transmission, as M., diversity, spider Hilker, Their M., of Lepidoptera: Kaltenpoth, role in E., Frago, potential L.R., the Voirol, Egypt. Paniagua Assessing Said, Port of (2016). Area, traits F.M. Protected Biocontrol Natural Semida, 112. (2018), Gamil G.M., B. Ashtoumel Telatin, Orabi, in Baldan, N., S.M., bioindicators G., Bordin, Elbanna, Valle, V., S., A., Guidolin, Ossamy, Squartini, G., Favaro, M., N., Zottini, Vitulo, S., licheniformis F., Marcato, Bacillus Zanella, E., A., biological Barizza, Tondello, as A., E., spiders of Baldan, role S., The Nigris, control: mosquito of review. future A The in predators agents: (2018). P. generalist control Manyanga, as S.D., spiders Llera, on J., Ndava, perspective Human updated the An of (2018). control. M.H. Profiles biological Entling, Functional S., Pekar, and R., Taxonomic Michalko, between Linkages and Exploring diversity low Microbiome, (2018). bumblebees: M.G.I. European central Langille, transient in highly communities A Bacterial eat? specificity. (2011). you high P.S. what Hempel, you H., Are spider Koch, house (2020). grey the H. in Krehenwinkel, microbiome R., gut prey-influenced Gillespie, and S., Tsau, S.R. organisms. Kennedy, cellular of evolution and origin understanding the 1951. for understanding Toward approach New M; Kanehisa, (2019). M. Tanabe, KEGG. K., in Morishima, variations M. Furumichi, genome Genomes, Y., and Sato, Genes M., of Kanehisa, Encyclopedia Kyoto KEGG: (2000). 27–30. S. 28, Goto, insects?. & for M. healthcare Kanehisa, general mutualists: Norton as 529–535. Edition), Actinobacteria 17, (2nd Science (2009). Evolving M. An Kaltenpoth of Microbiology: dynamics (2011), Infection J.W., spraying. (2018). Foster, insecticide Y. L.S., to Kikuchi, Joan response M., Hayatsu, in insects K., Tago, to A., soil Nagayama, from symbionts Y., insecticide-degrading Sato, T., Hori, H., Itoh, mSystems, uiatru avidum Cutibacterium irb Ecol., Microb. Oecologia, Bioinformatics L7,aclual noht of endophyte culturable a GL174, ,e00163-17. 3, n.J oqioRes. Mosquito J. Int. ora fteAkna cdm fScience of Academy Arkansas the of Journal 8,21–36. 189, uli cd Res Acids Nucleic 2 121-133. 62, 0 3123–3124. 30, , uigifn u irboaestablishment. microbiota gut infant during ,6–11. 5, , ,4,D590-D595. 47, ., 8 ii vinifera Vitis aun ognu,Ml Ecol. Mol. longinqua, Badumna Oecologia v Glera”. cv. r n.Microbiol., Front. 1 13. 71, , ninJ Arachnol., J. Indian 8,895–903. 181, , M Microbiol., BMC aioarabida Rabidosa ESMcoil Ecol. Microbiol. FEMS rti Sci Protein SEJ. ISME uli cd Res. Acids Nucleic rnsMicrobiol. Trends ,556. 9, 9 1001–1015. 29, , 2 909–920. 12, , 8 1947- 28, . Wolf a , 8 133. 18, ,100– 5, , , Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. a enue odanteefiue n ute dtdi db Photoshop. Adobe in Metacoder edited (http://www.R-project.org/) further software 2c. and based figures Figure R these The drawn to Araneidae. used family been has the Photoshop. of Adobe subfamilies in Metacoder three edited (http://www.R-project.org/) further software and 2b. based figures Figure R these The drawn to Araneidae. used family been has the of subfamilies three 2a. Figure Species species. spider (a) 1. Figure Captions Figure code Specimen Subfamily code. their No. with study Sl. this in studied species the of Feeding Front.List Model. (2020). Fermentation P. vitro in Payne, an 1 N. in Table Bell, Microbiome Gut J., Human Gathercole, the p.1763. E., Modify 11, Insects Rettedal, Microbiol., Edible identification M.R., Molecular Bugs: McNeill, to (2017). Bugs S.K., J. Arojju, Chen. W., I., Agnarsson, Young, D., Li, on Y., preying Zhang, of spiders L.-yu, sequencing of Yuan, Metagenomic J., Liu, herbivory. (2017). T.B., for al., Yang, adaptations et holobiont B., key Qin, of unveils H., phylogenetics microbiome Zhong, 663. the D., gut and Zheng, moth detection L., back Vasseur, Molecular diamond G.M., Gurr, (2010). X., Y. Xia, Peng, C., Jian, Y, li-Yun, Wolbachia C., Deng, by Z.Y., phenol Wang, of Biodegradation termite. India. the (2019). of of 1-293. R. fauna gut Calcutta, Boopathy, The India, S., In: of Dexter, orb-weavers. Survey Van Typical Zoological II. (Argiopidae). Vol. Araneidae Family Araneae. 1. Spiders: Part Population- (1982) Tissue-and B.K. (2020). Tikader, M.M. Bengtsson, Spider T., Wasp Urich, the T., Symbiont, of Lueders, Analysis S., Microbiome Prost, Level G., Uhl, M.M., Sheffer, Microorganisms nCieesies(Araneae), spiders Chinese in ette t re ee)o ufml rnia odmntaetegtbceildvriyin diversity bacterial gut the demonstrate to Araneinae subfamily of level) order (to tree Heat ette t re ee)o ufml ripnet eosrt h u atra iest in diversity bacterial gut the demonstrate to Argiopinae subfamily of level) order (to tree Heat ette t re ee)o ufml atrcnhnet eosrt h u bacterial gut the demonstrate to Gasteracanthinae subfamily of level) order (to tree Heat endarm()Audneo u atra iest ttepyaadfml ee ftwelve of level family and phyla the at diversity bacterial gut of Abundance (b) diagram Venn nio c oltRes. Pollut Sci Environ mosavitis Empoasca ,8. 8, , Gasteracanthinae Argiopinae Araneinae nataplantation.” tea a in Arachnol J 6 34067–72. 26, rip bruennichi Argiope 9 8 237-241. 38, , A1164 AA 154 AA 29 AA 136 AA 397 AA 1873 AA 1438 AA 2116 AA 598 AA 795 AA 787 AA 2217 AA c.Rep., Sci. cntbce tandoii Acinetobacter dnie oe oiatBacterial Dominant Novel a Identified ,1-10. 7, atrcnh hasselti Gasteracantha kuhli Gasteracantha cicatrosa Cyrtophora pulchella Argiope nautica Neoscona bengalensis Neoscona laglaizei Eriovixia excelsa Eriovixia bianchoria Cyclosa mulmeiensis Cyclosa spirifera Cyclosa mitificus Araneus rn.Microbiol., Front. sltdfo the from isolated 8, Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. aio ewe he ufmle ffml rnia.Alsqec ed eeue opeitfunc- predict to PICRUSt2 used in were implemented reads is sequence which All (http://www.genome.jp/kegg/), package. database software Araneidae. KEGG bioinformatics family (https://github.com/picrust/picrust2) the of against subfamilies tions three between parison 6 Figure species. spider twelve 5. Figure diversity. the represent bacterial whiskers gut median. phylum the the with of represents ends box The the 4. inside Araneidae. Figure line family the of while subfamilies maximum three and Photoshop. in minimum diversity Adobe in bacterial edited gut (http://www.R-project.org/) further the software and based figures R 3. these The Figure drawn to Araneidae. used family been the has of Metacoder subfamilies three in diversity edormof Dendrogram ryCri ismlrt-ae o-ercMliiesoa cln ND)odnto ltof plot ordination (NMDS) Scaling Multidimensional Non-metric dissimilarity-based Bray-Curtis o ltfrteapadvriyidx()osre,()Ca1 c hno n d ipo of Simpson (d) and Shannon (c) Chao1, (b) observed, (a) index alpha-diversity the for plot Box tce ounbrgahrpeetn h rdce ucinlmtblcptwycom- pathway metabolic functional predicted the representing graph bar column Stacked . β- iest ftev pce rmtresbaiiso h aiyAaede along Araneidae, family the of subfamilies three from species twelve of diversity 10 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 11 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 12 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 13 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 14 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 15 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 16 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 17 Posted on Authorea 12 Aug 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.159724530.01998846 — This a preprint and has not been peer reviewed. Data may be preliminary. 18