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Iifungicides zihlala zisetyenziswa ngexesha lokudubula kweziqhamo zemithi kwaye zinokusongela izinambuzane ezisasaza impova. Nangona kunjalo, kuncinci okwaziwayo malunga nendlela iinyosi ezingezizo iinyosi ezisasaza impova (umz. iinyosi ezizimeleyo, ii-Osmia cornifrons) ezisabela ngayo kwiifungicides ezidityanisiweyo nezisetyenziswa rhoqo kwiiapile ngexesha lokudubula. Olu luhlu lolwazi luthintela izigqibo zomthetho ezimisela ukugxila okukhuselekileyo kunye nexesha lokutshiza ifungicides. Sivavanye iziphumo zeefungicides ezimbini ezidityanisiweyo (i-captan kunye ne-mancozeb) kunye neefungicides ezine ezidityanisiweyo/i-phytosystem (i-ciprocycline, i-myclobutanil, i-pyrostrobin kunye ne-trifloxystrobin). Iziphumo ekuzuzeni ubunzima be-larval, ukusinda, umlinganiselo wesini kunye nokwahluka kweebhaktheriya. Uvavanyo lwenziwe kusetyenziswa uvavanyo lwe-bioassay olungapheliyo apho impova yanyangwa kwiidosi ezintathu ngokusekelwe kwidosi ecetyiswayo ngoku yokusetyenziswa endle (1X), idosi yesiqingatha (0.5X) kunye nedosi ephantsi (0.1X). Zonke iidosi ze-mancozeb kunye ne-pyritisoline zinciphise kakhulu ubunzima bomzimba kunye nokusinda kwe-larval. Emva koko silandelelanise i-16S gene ukuze sichaze i-larval bacteriome ye-mancozeb, i-fungicide ebangela ukufa okuphezulu. Sifumanise ukuba ukwahlukana kweebhaktheriya kunye nobuninzi bebhaktheriya kuncitshiswe kakhulu kwi-larvae etyalwe yi-pollen ephathwe yi-mancozeb. Iziphumo zethu zelebhu zibonisa ukuba ukutshiza ezinye zezi fungicides ngexesha lokudubula kweentyantyambo kuyingozi kakhulu kwimpilo ye-O. cornifrons. Olu lwazi lubalulekile kwizigqibo zolawulo lwexesha elizayo malunga nokusetyenziswa okuzinzileyo kweemveliso zokukhusela imithi yeziqhamo kwaye lusebenza njengesiseko seenkqubo zolawulo ezijolise ekukhuseleni i-pollinators.
Inyosi ye-mason eyodwa i-Osmia cornifrons (Hymenoptera: Megachilidae) yaziswa e-United States isuka eJapan ngasekupheleni kweminyaka yoo-1970 nasekuqaleni kweminyaka yoo-1980, kwaye olu hlobo ludlale indima ebalulekileyo ekuveliseni impova kwiindawo ezilawulwayo zendalo ukusukela ngoko. Amaqela ale nyosi aqhelekileyo ayinxalenye yeentlobo ezingama-50 zeenyosi zasendle ezihambelana neenyosi ezifunxa impova kwiigadi ze-almond kunye nee-apile e-United States2,3. Iinyosi zeMason zijongene nemingeni emininzi, kubandakanya ukuqhekeka kwendawo yokuhlala, izifo ezibangelwa zizifo, kunye nezibulala-zinambuzane3,4. Phakathi kwezibulali-zinambuzane, izibulali-zinambuzane zinciphisa ukufumana amandla, ukutya ukutya5 kunye nokulungisa umzimba6,7. Nangona uphando lwakutshanje lubonisa ukuba impilo yeenyosi zeMason iphenjelelwa ngokuthe ngqo ziintsholongwane ze-commensal kunye ne-ectobactic, 8,9 kuba iintsholongwane kunye nefungi zinokuchaphazela isondlo kunye neempendulo zomzimba, iziphumo zokuvezwa kwe-fungi kwiintlobo ezahlukeneyo zeenyosi zeMason ziqala ukufundwa.
Imithi yokubulala iintsholongwane eneziphumo ezahlukeneyo (ukudibana kunye nokusebenza komzimba) itshizwa kwiigadi zezityalo ngaphambi nangexesha lokudubula ukuze kunyangwe izifo ezifana ne-apple scab, ukubola okukrakra, ukubola okumdaka kunye ne-powdery mildew10,11. Imithi yokubulala iintsholongwane ithathwa njengengenabungozi kwizityalo ezisasaza i-pollinators, ngoko ke iyacetyiswa kubalimi ngexesha lokudubula; Ukuvezwa kunye nokuginywa kwezi mithi yokubulala iintsholongwane ziinyosi kwaziwa kakhulu, njengoko kuyinxalenye yenkqubo yokubhaliswa kwezibulali zinambuzane yi-US Environmental Protection Agency kunye nezinye ii-arhente ezininzi zolawulo lwesizwe12,13,14. Nangona kunjalo, iziphumo zemithi yokubulala iintsholongwane kwiinyosi ezingezizo iinyosi aziwa kangako kuba azifuneki phantsi kwezivumelwano zokugunyaziswa kwentengiso e-United States15. Ukongeza, ngokubanzi akukho migaqo imiselweyo yokuvavanya iinyosi ezizimeleyo16,17, kwaye ukugcina iikholoni ezibonelela iinyosi ukuze zivavanye kunzima18. Uvavanyo lweenyosi ezahlukeneyo ezilawulwayo luqhutywa ngakumbi eYurophu nase-USA ukuze kufundwe iziphumo zemithi yokubulala iintsholongwane kwiinyosi zasendle, kwaye imigaqo emiselweyo isandula ukwenziwa kwi-O. cornifrons19.
Iinyosi ezineempondo zii-monocytes kwaye zisetyenziswa kwizityalo ze-carp njengesongezelelo okanye indawo yeenyosi zobusi. Ezi nyosi ziphuma phakathi kukaMatshi no-Epreli, apho iinkunzi ezisandula ukuzala zivela kwiintsuku ezintathu ukuya kwezine ngaphambi kokuba iimazi ziqhagamshele. Emva kokudibana, imazi iqokelela impova kunye ne-nectar ukuze inike uthotho lweeseli ze-brood ngaphakathi kwe-tube nest cavity (yendalo okanye yokwenziwa)1,20. Amaqanda abekwa kwi-pollen ngaphakathi kweeseli; imazi emva koko yakha udonga lodongwe ngaphambi kokulungiselela iseli elandelayo. Ii-instar larvae zokuqala zifakwe kwi-chorion kwaye zondla ulwelo lwe-embryonic. Ukusuka kwi-instar yesibini ukuya kweyesihlanu (i-prepupa), ii-larvae zondla i-pollen22. Nje ukuba umbane we-pollen uphele ngokupheleleyo, ii-larvae zenza ii-cocoons, ii-pupate kwaye ziphume njengabantu abadala kwigumbi elifanayo le-brood, ngesiqhelo ekupheleni kwehlobo20,23. Ezikhulileyo ziphuma entwasahlobo elandelayo. Ukusinda kwabantu abadala kunxulunyaniswa nokwanda kwamandla (ukwanda kobunzima) ngokusekelwe ekutyeni. Ke ngoko, umgangatho wesondlo we-pollen, kunye nezinye izinto ezifana nemozulu okanye ukuvezwa kwi-pesticides, zizinto ezibangela ukusinda kunye nempilo24.
Izibulali-zinambuzane kunye nezibulali-zinambuzane ezisetyenziswa ngaphambi kokuba zidubule ziyakwazi ukuhamba ngaphakathi kwemithambo yegazi yesityalo ukuya kumanqanaba ahlukeneyo, ukusuka kwi-translaminar (umz., ukukwazi ukuhamba ukusuka kumphezulu wamagqabi ukuya kumphezulu ongezantsi, njengezinye izibulali-zinambuzane) 25 ukuya kwimiphumo yenkqubo yokwenyani. , ezinokungena kwisithsaba ukusuka kwiingcambu, zingangena kwi-nectar yeentyatyambo zeapile26, apho zinokubulala khona ii-O. cornifrons ezindala27. Ezinye izibulali-zinambuzane nazo ziyangena kwi-pollen, zichaphazela uphuhliso lwee-corn larvae kwaye zibangele ukufa kwazo19. Ezinye izifundo zibonise ukuba ezinye izibulali-zinambuzane zinokutshintsha kakhulu indlela yokuzalela kweentlobo ezinxulumene nazo i-O. lignaria28. Ukongeza, izifundo zelebhu kunye nezifundo zasentsimini ezilinganisa iimeko zokuchayeka kwi-pesticide (kubandakanya izibulali-zinambuzane) zibonise ukuba izibulali-zinambuzane zichaphazela kakubi i-physiology22 morphology29 kunye nokusinda kweenyosi zobusi kunye nezinye iinyosi ezizimeleyo. Izitshizi ezahlukeneyo zokubulala iifungi ezisetyenziswa ngqo kwiintyatyambo ezivulekileyo ngexesha lokudubula zinokungcolisa i-pollen eqokelelwe ngabantu abadala ukuze kuphuhliswe ii-larval, iziphumo zazo ezisaza kufundwa30.
Kuya kuqatshelwa ngakumbi ukuba uphuhliso lwe-larval luphenjelelwa yi-pollen kunye ne-microbial communities kwinkqubo yokugaya ukutya. I-honeybee microbiome inefuthe kwiiparameters ezifana nobunzima bomzimba31, utshintsho lwe-metabolic22 kunye nokuchaphazeleka kwi-pathogens32. Izifundo zangaphambili zihlolisise impembelelo yesigaba sophuhliso, izondlo, kunye nokusingqongileyo kwi-microbiome yeenyosi ezizimeleyo. Ezi zifundo zityhile ukufana kwisakhiwo kunye nobuninzi be-larval kunye ne-pollen microbiomes33, kunye ne-bacterial genera eqhelekileyo i-Pseudomonas kunye ne-Delftia, phakathi kweentlobo zeenyosi ezizimeleyo. Nangona kunjalo, nangona ii-fungicides zidibene nezicwangciso zokukhusela impilo yeenyosi, iziphumo ze-fungicides kwi-larval microbiota ngokuchatshazelwa ngqo ngomlomo azikahlolwa.
Olu phononongo luvavanye iziphumo zedosi zokwenyani zeefungicides ezintandathu ezisetyenziswa rhoqo ezibhalisiweyo ukusetyenziswa kwiziqhamo zomthi eMelika, kuquka iifungicides ezidityanisiweyo nezisetyenziswa rhoqo ezinikwa ngomlomo kwiimbungu ze-corn hornworm moth ezivela kukutya okungcolileyo. Sifumanise ukuba iifungicides ezidityanisiweyo nezisetyenziswa rhoqo zinciphisa ubunzima bomzimba weenyosi kunye nokufa okwandisiweyo, nto leyo ebangela ukuba kubekho imiphumo emibi kakhulu enxulunyaniswa ne-mancozeb kunye ne-pyrithiopide. Emva koko sithelekise ukwahluka kweentsholongwane zeembungu ezidliwa kukutya kwe-pollen ephathwe yi-mancozeb kunye nezo zidliwa kukutya okulawulayo. Sixoxa ngeendlela ezinokubakho ezibangela ukufa kunye nemiphumo kwiinkqubo zolawulo lwezinambuzane kunye ne-pollinator oludibeneyo (IPPM)36.
Ii-O. cornifrons ezindala ezihlala ebusika kwiikhokoni zifunyenwe kwiZiko loPhando lweZithelo, eBiglerville, ePA, zaza zagcinwa kwi-−3 ukuya kwi-2°C (±0.3°C). Ngaphambi kovavanyo (iikhokoni ezingama-600 zizonke). NgoMeyi 2022, iikhokoni ze-O. cornifrons ezili-100 zazidluliselwa imihla ngemihla kwiikomityi zeplastiki (iikhokoni ezingama-50 ngekomityi nganye, ubude be-DI 5 cm × 15 cm) kwaye ii-wipes zafakwa ngaphakathi kwiikomityi ukukhuthaza ukuvulwa nokubonelela ngesiseko esihlafunwayo, ukunciphisa uxinzelelo kwiinyosi ezinamatye. Beka iikomityi ezimbini zeplastiki ezineekhokoni kwikheyiji yezinambuzane (30 × 30 × 30 cm, BugDorm MegaView Science Co. Ltd., eTaiwan) kunye nee-feeders ze-10 ml eziqulethe isisombululo se-50% ye-sucrose kwaye ugcine iintsuku ezine ukuqinisekisa ukuvalwa kunye nokudibana. 23°C, ukufuma okulinganiselweyo 60%, ixesha le-photoperiod 10 l (ubunzima obuphantsi): iintsuku ezili-14. Iimazi ezili-100 kunye neenkunzi ezizalelweyo zazikhululwa rhoqo kusasa kangangeentsuku ezintandathu (100 ngosuku) zifakwe kwizidleke ezimbini zokwenziwa ngexesha lokudubula kwe-apile (isidleke setrap: ububanzi 33.66 × ukuphakama 30.48 × ubude 46.99 cm; Umfanekiso ongezelelweyo 1). Zibekwe kwiPennsylvania State Arboretum, kufutshane ne-cherry (Prunus cerasus 'Eubank' Sweet Cherry Pie™), ipesika (Prunus persica 'Contender'), iPrunus persica 'PF 27A' Flamin Fury®), ipeya (Pyrus perifolia 'Olympic', Pyrus perifolia 'Shinko', Pyrus perifolia 'Shinseiki'), coronaria apple tree (Malus coronaria) kunye neentlobo ngeentlobo zemithi yama-apile (Malus coronaria, Malus), domestic apple tree 'Co-op 30′ Enterprise™, Malus apple tree 'Co-Op 31′ Winecrisp™, begonia 'Freedom', Begonia 'Golden Delicious', Begonia 'Nova Spy'). Indlu nganye yeentaka yeplastiki eluhlaza okwesibhakabhaka ingena phezu kweebhokisi ezimbini zomthi. Ibhokisi nganye yendlwana yayineetyhubhu zephepha zekraft ezingama-800 ezingenanto (ezivulekileyo nge-spiral, 0.8 cm ID × 15 cm L) (Jonesville Paper Tube Co., Michigan) ezifakwe kwiityhubhu zecellophane ezingabonakali (0.7 OD jonga iiplagi zeplastiki (iiplagi ze-T-1X) zibonelela ngendawo yokuzalela.
Zombini iibhokisi zeentaka zazijonge empuma kwaye zazigqunywe ngocingo lwegadi oluluhlaza lweplastiki (imodeli ye-Everbilt #889250EB12, ubungakanani bokuvula buyi-5 × 5 cm, 0.95 m × 100 m) ukuthintela ukufikelela kweempuku kunye neentaka kwaye zibekwe kumphezulu womhlaba ecaleni kweebhokisi zomhlaba zebhokisi yeentaka. Ibhokisi yeentaka (Umfanekiso ongezelelweyo 1a). Amaqanda e-corn borer ayeqokelelwa yonke imihla ngokuqokelela iityhubhu ezingama-30 kwiindlwana aze azithuthe aye kwilebhu. Usebenzisa isikere, yenza ukusika ekupheleni kwetyhubhu, uze uqhawule ityhubhu ye-spiral ukuze uveze iiseli zeentaka. Amaqanda ngamanye kunye ne-pollen yawo asuswe kusetyenziswa i-spatula egobileyo (i-Microslide tool kit, i-BioQuip Products Inc., eCalifornia). Amaqanda afakwa kwiphepha lesihluzo elimanzi aze abekwe kwisitya sePetri iiyure ezi-2 ngaphambi kokuba asetyenziswe kuvavanyo lwethu (Umfanekiso ongezelelweyo 1b-d).
Kwilabhoratri, sivavanye ubuthi bomlomo be-fungicides ezintandathu ezisetyenzisiweyo ngaphambi nangexesha le-apile blossom kumanqanaba amathathu (0.1X, 0.5X, kunye ne-1X, apho i-1X iluphawu olusetyenziswa kwiilitha ezili-100 zamanzi/ihektare. Umthamo ophezulu wentsimi = uxinano kwintsimi). , Itheyibhile 1). Uxinano ngalunye luphindwe amaxesha ali-16 (n = 16). Ii-fungicides ezimbini zoqhagamshelwano (Itheyibhile S1: mancozeb 2696.14 ppm kunye ne-captan 2875.88 ppm) kunye ne-fungicides ezine ze-systemic (Itheyibhile S1: pyrithiostrobin 250.14 ppm; trifloxystrobin 110.06 ppm; myclobutanil azole 75 .12 ppm; cyprodinil 280.845 ppm) ubuthi kwiziqhamo, imifuno kunye nezityalo zokuhombisa. Siyilungisile impova sisebenzisa isixhobo sokusila, sadlulisela i-0.20 g emthonjeni (iFalcon Plate enamaqula angama-24), saza songeza saza saxuba i-1 μL yesisombululo sokukhunta ukuze senze impova ye-pyramidal kunye nemigodi enzulu eyi-1 mm apho amaqanda abekwe khona. Beka usebenzisa i-spatula encinci (Umfanekiso ongezelelweyo 1c, d). Iipleyiti zeFalcon zigcinwe kubushushu begumbi (25°C) kunye nomswakama ohambelanayo oyi-70%. Sizithelekise neembungu ezilawulayo eziphakelwa ukutya kwempova efanayo enyangiwe ngamanzi acocekileyo. Sirekhode ukufa kwaye silinganisa ubunzima beembungu yonke imihla de iimbungu zifikelele kwiminyaka yobuntwana sisebenzisa ibhalansi yohlalutyo (Fisher Scientific, ukuchaneka = 0.0001 g). Ekugqibeleni, umlinganiselo wesini uvavanywe ngokuvula i-cocoon emva kweenyanga ezi-2.5.
I-DNA ikhutshwe kwi-O. cornifrons larvae epheleleyo (n = 3 ngemeko yonyango, i-pollen ephathwe yi-mancozeb kwaye inganyangwanga) kwaye senze uhlalutyo lokwahlukana kweentsholongwane kwezi sampuli, ingakumbi kuba kwi-mancozeb ukufa okuphezulu kubonwe kwi-larvae. ukufumana i-MnZn. I-DNA yandiswa, yahlanjululwa kusetyenziswa i-DNAZymoBIOMICS®-96 MagBead DNA kit (Zymo Research, Irvine, CA), kwaye yalandelelaniswa (ii-cycles ezingama-600) kwi-Illumina® MiSeq™ kusetyenziswa i-v3 kit. Ukulandelelana okujoliswe kuko kwe-bacterial 16S ribosomal RNA genes kwenziwe kusetyenziswa i-Quick-16S™ NGS Library Prep Kit (Zymo Research, Irvine, CA) kusetyenziswa ii-primers ezijolise kummandla we-V3-V4 we-16S rRNA gene. Ukongeza, ulandelelwano lwe-18S lwenziwe kusetyenziswa i-10% PhiX inclusion, kwaye ukwandiswa kwenziwa kusetyenziswa i-primer pair 18S001 kunye ne-NS4.
Ukungenisa kunye nokucubungula ii-reads39 ezidityanisiweyo kusetyenziswa umbhobho we-QIIME2 (v2022.11.1). Ezi reads zinqunyulwe zaza zadityaniswa, kwaye ulandelelwano lwe-chimeric lususiwe kusetyenziswa i-plugin ye-DADA2 kwi-QIIME2 (ukubhangqa ingxolo ye-qiime dada2)40. Izabelo zeklasi ze-16S kunye ne-18S zenziwe kusetyenziswa i-plugin ye-object classifier Classify-sklearn kunye ne-artifact silva-138-99-nb-classifier eqeqeshwe kwangaphambili.
Yonke idatha yovavanyo ihlolwe ukuba iqhelekile na (uShapiro-Wilks) kunye nokuhambelana kweenguqu (uvavanyo lukaLevene). Ngenxa yokuba iseti yedatha ayihlangabezananga neengqikelelo zohlalutyo lwe-parametric kwaye utshintsho aluzange lukwazi ukulinganisa iintsalela, senze i-ANOVA yeendlela ezimbini engeyiyo i-parametric (iKruskal-Wallis) eneempawu ezimbini [ixesha (amanqaku exesha lesithathu le-2, 5, kunye neentsuku ezi-8) kunye ne-fungicide] ukuvavanya isiphumo sonyango kubunzima obutsha be-larval, emva koko uthelekiso lwe-post hoc nonparametric pairing lwenziwa kusetyenziswa uvavanyo lweWilcoxon. Sisebenzise imodeli yomgca oqhelekileyo (i-GLM) enokusasazwa kwePoisson ukuthelekisa imiphumo ye-fungicide ekusindeni kwiingxinano ezintathu ze-fungicide41,42. Uhlalutyo lobuninzi obuhlukeneyo, inani lee-variants ze-amplicon sequence (ii-ASV) lawa kwinqanaba le-genus. Uthelekiso lobuninzi bokwahluka phakathi kwamaqela kusetyenziswa i-16S (inqanaba lohlobo) kunye nobuninzi bokuthelekisa be-18S lwenziwe kusetyenziswa imodeli yokongeza ngokubanzi yesikhundla, isikali, kunye nemilo (GAMLSS) kunye nokusasazwa kosapho lwe-beta zero-inflated (BEZI), oluye lwalinganiswa kwi-macro. kwi-Microbiome R43 (v1.1). 1). Susa iintlobo ze-mitochondrial kunye ne-chloroplast ngaphambi kohlalutyo lokwahluka. Ngenxa yamanqanaba ahlukeneyo e-taxonomic e-18S, kuphela inqanaba eliphantsi le-taxon nganye elisetyenziselwe uhlalutyo lokwahluka. Lonke uhlalutyo lwezibalo lwenziwe kusetyenziswa i-R (v. 3.4.3., iprojekthi ye-CRAN) (Iqela lika-2013).
Ukuvezwa kwi-mancozeb, i-pyrithiostrobin, kunye ne-trifloxystrobin kunciphisa kakhulu ukuqokelelwa kobunzima bomzimba kwi-O. cornifrons (Umzobo 1). Ezi ziphumo zibonwe rhoqo kuzo zonke iidosi ezintathu ezivavanyiweyo (Umzobo 1a–c). I-Cyclostrobin kunye ne-myclobutanil azizange zinciphise kakhulu ubunzima bee-larvae.
Ubunzima obuqhelekileyo be-stem borer larvae bulinganiswa ngamaxesha amathathu phantsi konyango olune lokutya (ukutya kwe-pollen efanayo + i-fungicide: ulawulo, iidosi eziyi-0.1X, 0.5X kunye ne-1X). (a) Idosi ephantsi (0.1X): indawo yokuqala (usuku 1): χ2: 30.99, DF = 6; P < 0.0001, indawo yesibini (usuku 5): 22.83, DF = 0.0009; ixesha lesithathu; indawo (usuku 8): χ2: 28.39, DF = 6; (b) isiqingatha sedosi (0.5X): indawo yokuqala (usuku 1): χ2: 35.67, DF = 6; P < 0.0001, indawo yesibini (usuku lokuqala). ): χ2: 15.98, DF = 6; P = 0.0090; inqaku lesithathu (usuku 8) χ2: 16.47, DF = 6; (c) Indawo okanye idosi epheleleyo (1X): inqaku lesihlandlo sokuqala (usuku 1) χ2: 20.64, P = 6; P = 0.0326, inqaku lesibini (usuku 5): χ2: 22.83, DF = 6; P = 0.0009; inqaku lesithathu (usuku 8): χ2: 28.39, DF = 6; uhlalutyo olungelulo lweparametric lomahluko. Iibha zimele i-mean ± SE yothelekiso lwesibini (α = 0.05) (n = 16) *P ≤ 0.05, **P ≤ 0.001, ***P ≤ 0.0001.
Kwidosi ephantsi (0.1X), ubunzima bomzimba we-larval buncitshiswe ngama-60% nge-trifloxystrobin, ama-49% nge-mancozeb, ama-48% nge-myclobutanil, kunye nama-46% nge-pyrithistrobin (Umfanekiso 1a). Xa bevezwa kwisiqingatha sedosi yasendle (0.5X), ubunzima bomzimba we-mancozeb larvae buncitshiswe ngama-86%, i-pyrithiostrobin ngama-52% kunye ne-trifloxystrobin ngama-50% (Umfanekiso 1b). Idosi epheleleyo yasendle (1X) ye-mancozeb incitshiswe ngama-82%, i-pyrithiostrobin ngama-70%, kunye ne-trifloxystrobin, i-myclobutanil kunye ne-sangard malunga nama-30% (Umfanekiso 1c).
Ukufa bekuphezulu phakathi kweempungutye ezityiwa yi-pollen ephathwe yi-mancozeb, kulandele i-pyrithiostrobin kunye ne-trifloxystrobin. Ukufa kunyuke ngokunyusa iidosi ze-mancozeb kunye ne-pyritisoline (Umzobo 2; Itheyibhile 2). Nangona kunjalo, ukufa kwe-corn borer kunyuke kancinci njengoko ubuninzi be-trifloxystrobin bunyuka; i-cyprodinil kunye ne-captan azizange zinyuse kakhulu ukufa xa kuthelekiswa nonyango lolawulo.
Ukufa kweempukane ze-borer fly kuthelekiswe emva kokufunxa i-pollen nganye nganye enyangwe ngamayeza amathandathu ahlukeneyo okubola. I-Mancozeb kunye ne-pentopyramide zazinovelwano ngakumbi xa zichatshazelwa ngomlomo ziimpethu zombona (GLM: χ = 29.45, DF = 20, P = 0.0059) (umgca, i-slope = 0.29, P < 0.001; i-slope = 0.24, P <0.00)).
Ngokomyinge, kuzo zonke iindlela zonyango, ama-39.05% ezigulana yayingamabhinqa kwaye ama-60.95% yayingamadoda. Phakathi konyango lolawulo, inani labasetyhini yayiyi-40% kwizifundo zedosi ephantsi (0.1X) kunye nedosi yesiqingatha (0.5X), kunye nama-30% kwizifundo zedosi yasentsimini (1X). Kwidosi ye-0.1X, phakathi kwezibungu ezityiswe ngepollen eziphathwe nge-mancozeb kunye ne-myclobutanil, ama-33.33% abantu abadala yayingamabhinqa, ama-22% abantu abadala yayingamabhinqa, ama-44% ezibungu ezindala yayingamabhinqa, ama-44% ezibungu ezindala yayingamabhinqa. Amabhinqa, ama-41% ezibungu ezindala yayingamabhinqa, kwaye abalawuli babeyi-31% (Umzobo 3a). Xa idosi iphindwe kayi-0.5, ama-33% eempethu ezindala kwiqela le-mancozeb kunye ne-pyrithiostrobin yayingamabhinqa, ama-36% kwiqela le-trifloxystrobin, ama-41% kwiqela le-myclobutanil, kunye nama-46% kwiqela le-cyprostrobin. Eli nani lalingama-53% kwiqela. kwiqela le-captan kunye nama-38% kwiqela lolawulo (Umzobo 3b). Kwidosi ye-1X, ama-30% eqela le-mancozeb yayingamabhinqa, ama-36% eqela le-pyrithiostrobin, ama-44% eqela le-trifloxystrobin, ama-38% eqela le-myclobutanil, ama-50% eqela lolawulo yayingabafazi - ama-38.5% (Umzobo 3c).
Ipesenti yezilwanyana ezifunxayo eziziimazi neziziinkunzi emva kokuba zichaphazeleke kwi-fungicide kwinqanaba le-larval stage. (a) Idosi ephantsi (0.1X). (b) Idosi esiqingatha (0.5X). (c) Idosi yasentsimini okanye idosi epheleleyo (1X).
Uhlalutyo lwe-16S sequence lubonise ukuba iqela leebhaktheriya lahlukile phakathi kwee-larvae ezityiswa nge-pollen ephathwe yi-mancozeb kunye nee-larvae ezityiswa nge-pollen enganyangwanga (Umzobo 4a). I-microbial index yee-larvae ezinganyangwanga ezityiswa nge-pollen yayiphezulu kuneye-larvae ezityiswa nge-pollen ephathwe yi-mancozeb (Umzobo 4b). Nangona umahluko obonweyo kubutyebi phakathi kwamaqela wawungabalulekanga ngokwezibalo, wawuphantsi kakhulu kunowo ubonwe kwii-larvae ezityiswa nge-pollen enganyangwanga (Umzobo 4c). Ubuninzi obuthelekisekayo bubonise ukuba ii-microbiota zee-larvae ezityiswa nge-pollen elawulwayo zahlukile kakhulu kuneze-larvae ezityiswa kwi-larvae ephathwe yi-mancozeb (Umzobo 5a). Uhlalutyo oluchazayo lubonise ukuba kukho ii-genera ezingama-28 kwiisampulu zolawulo kunye ne-mancozeb ephathwe ngayo (Umzobo 5b). c Uhlalutyo olusebenzisa ulandelelwano lwe-18S alubonisanga mahluko abalulekileyo (Umfanekiso ongezelelweyo 2).
Iiprofayili ze-SAV ezisekelwe kulandelelwano lwe-16S zithelekiswe nobutyebi beShannon kwaye zabona ubutyebi kwinqanaba le-phylum. (a) Uhlalutyo lwe-Principal coordinate (PCoA) olusekelwe kwisakhiwo soluntu olupheleleyo lwe-microbial kwi-pollen-fed okanye ulawulo (oluhlaza okwesibhakabhaka) kunye ne-mancozeb-fed larvae (orenji). Inqaku ngalinye ledatha limele isampuli eyahlukileyo. I-PCoA ibalwe kusetyenziswa umgama weBray-Curtis wokusasazwa kwe-t ye-multivariate. Ii-ovals zimele inqanaba lokuzithemba le-80%. (b) I-Boxplot, idatha ye-Shannon wealth eluhlaza (amanqaku) kunye ne-c. Ubutyebi obubonakalayo. Ii-Boxplot zibonisa iibhokisi zomgca ophakathi, uluhlu lwe-interquartile (IQR), kunye ne-1.5 × IQR (n = 3).
Ukwakheka kweendawo zeentsholongwane zeembungu ezitya impova enyangiweyo nenganyangwanga yi-mancozeb. (a) Ubuninzi beentlobo zeentsholongwane bufundeka kwiimpungutye. (b) Imephu yobushushu yeendawo zeentsholongwane ezichongiweyo. I-Delftia (umlinganiselo weengxaki (OR) = 0.67, P = 0.0030) kunye ne-Pseudomonas (OR = 0.3, P = 0.0074), i-Microbacterium (OR = 0.75, P = 0.0617) (OR = 1.5, P = 0.0060); Imigca yemephu yobushushu idityaniswe ngokusebenzisa umgama wokunxibelelana kunye noqhagamshelwano oluphakathi.
Iziphumo zethu zibonisa ukuba ukuchatshazelwa ngomlomo kwi-contact (mancozeb) kunye ne-systemic (pyrostrobin kunye ne-trifloxystrobin), ezisetyenziswa kakhulu ngexesha lokudubula, kunciphise kakhulu ukutyeba kunye nokwanda kokufa kwe-corn larvae. Ukongeza, i-mancozeb inciphise kakhulu ukwahluka kunye nobutyebi be-microbiome ngexesha lesigaba sokulungiselela. I-Myclobutanil, enye i-systemic fungicide, inciphise kakhulu ukutyeba komzimba we-larvae kuzo zonke iidosi ezintathu. Esi siphumo sabonakala kwixesha lesibini (ngosuku lwesi-5) kunye nelesithathu (ngosuku lwesi-8). Ngokwahlukileyo koko, i-cyprodinil kunye ne-captan azizange zinciphise kakhulu ukutyeba okanye ukusinda xa kuthelekiswa neqela lolawulo. Ngokolwazi lwethu, lo msebenzi ngowokuqala ukumisela imiphumo yamanqanaba entsimi ye-fungicides ezahlukeneyo ezisetyenziselwa ukukhusela izityalo zombona ngokuchatshazelwa ngqo yi-pollen.
Zonke iindlela zonyango lwe-fungicide zinciphise kakhulu ukwanda kobunzima bomzimba xa kuthelekiswa nonyango lolawulo. I-Mancozeb ibe nefuthe elikhulu ekunyukeni kobunzima bomzimba we-larval ngokunciphisa okuphakathi kwe-51%, kulandele i-pyrithiostrobin. Nangona kunjalo, ezinye izifundo azizange zibike iziphumo ezimbi zeedosi zentsimi ze-fungicides kwizigaba ze-larval44. Nangona i-dithiocarbamate biocides ibonakalisiwe ukuba inetyhefu ephantsi kakhulu45, i-ethylene bisdithiocarbamates (EBDCS) efana ne-mancozeb inokonakala ibe yi-urea ethylene sulfide. Ngenxa yeziphumo zayo zokuguqulwa kwezinye izilwanyana, le mveliso yokonakala inokuba noxanduva lweziphumo ezibonwe46,47. Izifundo zangaphambili zibonise ukuba ukwakheka kwe-ethylene thiourea kuphenjelelwa zizinto ezifana nobushushu obuphezulu48, amanqanaba omswakama49 kunye nobude bokugcinwa kwemveliso50. Iimeko ezifanelekileyo zokugcina i-biocides zinokunciphisa ezi ziphumo bezingalindelekanga. Ukongeza, i-European Food Safety Authority ibonakalise inkxalabo malunga nobuthi be-pyrithiopide, ebonakalisiwe ukuba ibangela umhlaza kwiinkqubo zokugaya zezinye izilwanyana51.
Ukufakwa ngomlomo kwe-mancozeb, i-pyrithiostrobin, kunye ne-trifloxystrobin kwandisa ukufa kwee-larvae ze-corn borer. Ngokwahlukileyo koko, i-myclobutanil, i-ciprocycline kunye ne-captan azizange zibe nefuthe ekufeni. Ezi ziphumo zahlukile kwezo zikaLadurner et al.52, ababonise ukuba i-captan inciphise kakhulu ukusinda kwe-O. lignaria yabantu abadala kunye ne-Apis mellifera L. (Hymenoptera, Apisidae). Ukongeza, ii-fungicides ezifana ne-captan kunye ne-boscalid zifunyenwe zibangela ukufa kwee-larval52,53,54 okanye zitshintshe indlela yokutya55. Olu tshintsho, ngokunye, lunokuchaphazela umgangatho wesondlo se-pollen kwaye ekugqibeleni ufumane amandla kwinqanaba le-larval. Ukufa okubonwe kwiqela lolawulo bekungqinelana nezinye izifundo 56,57.
Umlinganiselo wesini othandwa ngamadoda obonwe kumsebenzi wethu unokucaciswa zizinto ezinje ngokungalingani kokuzala kunye neemeko zemozulu ezimbi ngexesha lokudubula kweentyatyambo, njengoko bekucetyisiwe ngaphambili kwi-O. cornuta nguVicens noBosch. Nangona iimazi kunye neenkunzi kwisifundo sethu bezineentsuku ezine zokuzala (ixesha eliqhelekileyo elithathwa njengelaneleyo lokuzala ngempumelelo), sinciphise ngabom ukukhanya ukuze sinciphise uxinzelelo. Nangona kunjalo, olu tshintsho lunokuphazamisana ngempazamo nenkqubo yokuzala61. Ukongeza, iinyosi zifumana iintsuku ezininzi zemozulu embi, kubandakanya imvula kunye namaqondo obushushu aphantsi (<5°C), anokuchaphazela kakubi impumelelo yokuzala4,23.
Nangona uphando lwethu lugxile kwi-microbiome yonke ye-larval, iziphumo zethu zibonelela ngengqiqo kubudlelwane obunokubakho phakathi kweendawo zebhaktiriya obunokuba luncedo kakhulu kwisondlo seenyosi kunye nokuvezwa kwe-fungicide. Umzekelo, i-pollen ephathwe yi-mancozeb enikwe iimbungu iye yanciphisa kakhulu isakhiwo soluntu lwe-microbial kunye nobuninzi xa kuthelekiswa ne-pollen ephathwe yimbungu enganyangwanga. Kwi-pollen etya i-pollen enganyangwanga, amaqela eebhaktiriya i-Proteobacteria kunye ne-Actinobacteria ayenamandla kwaye ayehlala e-aerobic okanye e-facultatively aerobic. Iibhaktiriya ze-Delft, ezihlala zinxulunyaniswa neentlobo zeenyosi ezizimeleyo, zaziwa ngokuba nomsebenzi we-antibiotic, nto leyo ebonisa indima enokubakho yokukhusela kwi-pathogens. Olunye uhlobo lwebhaktiriya, i-Pseudomonas, yayininzi kwi-pollen ephathwe yi-mungu enganyangwanga, kodwa yancitshiswa kakhulu kwi-pollen ephathwe yi-mancozeb. Iziphumo zethu zixhasa izifundo zangaphambili ezichonga i-Pseudomonas njengenye yezona genera zininzi kwi-O. bicornis35 kunye nezinye ii-wolps34 ezizimeleyo. Nangona ubungqina bovavanyo ngendima yePseudomonas kwimpilo ye-O. cornifrons bungakhange bufundwe, le bhaktiriya ibonakalisiwe ukuba ikhuthaza ukwenziwa kweetyhefu ezikhuselayo kwi-beetle Paederus fuscipes kwaye ikhuthaza i-arginine metabolism kwi-vitro 35, 65. Ezi ngqwalasela zibonisa indima enokubakho ekukhuseleni iintsholongwane kunye neebhaktiriya ngexesha lokuphuhliswa kwe-O. cornifrons larvae. I-Microbacterium lolunye uhlobo oluchongiweyo kuphononongo lwethu oluxelwe ukuba lukhona ngamanani aphezulu kwi-black soldier fly larvae phantsi kweemeko zendlala66. Kwi-O. cornifrons larvae, i-microbacteria inokuba negalelo ekulinganisweni nasekuqineni kwe-microbiome yamathumbu phantsi kweemeko zoxinzelelo. Ukongeza, i-Rhodococcus ifumaneka kwi-O. cornifrons larvae kwaye yaziwa ngamakhono ayo okukhupha ubuthi67. Olu hlobo lukwafumaneka emathunjini e-A. florea, kodwa luncinci kakhulu68. Iziphumo zethu zibonisa ukuba kukho umahluko omkhulu we-genetic kwiintlobo ezininzi ze-microbial taxa ezinokutshintsha iinkqubo ze-metabolic kwi-larvae. Nangona kunjalo, kufuneka ukuqonda ngcono ukwahluka kokusebenza kwe-O. cornifrons.
Ngamafutshane, iziphumo zibonisa ukuba i-mancozeb, i-pyrithiostrobin, kunye ne-trifloxystrobin zinciphise ukwanda kobunzima bomzimba kunye nokwanda kokufa kwee-corn borer larvae. Nangona kukho inkxalabo ekhulayo malunga nemiphumo ye-fungicides kwi-pollinators, kukho imfuneko yokuqonda ngcono imiphumo ye-metabolites eseleyo yale mixube. Ezi ziphumo zinokufakwa kwiingcebiso zeenkqubo zolawulo lwe-pollinator ezidibeneyo ezinceda abalimi ukuba baphephe ukusebenzisa ii-fungicides ezithile ngaphambi nangexesha lokudubula kwemithi yeziqhamo ngokukhetha ii-fungicides kunye nokutshintsha ixesha lokusetyenziswa, okanye ngokukhuthaza ukusetyenziswa kwezinye iindlela ezingenabungozi 36. Olu lwazi lubalulekile ekuphuhliseni iingcebiso malunga nokusetyenziswa kwezibulali-zinambuzane, njengokulungisa iinkqubo zokutshiza ezikhoyo kunye nokutshintsha ixesha lokutshiza xa ukhetha ii-fungicides okanye ukukhuthaza ukusetyenziswa kwezinye iindlela ezingenabungozi bungako. Kufuneka uphando olongezelelweyo kwimiphumo emibi ye-fungicides kumlinganiselo wesini, indlela yokutya, i-gut microbiome, kunye neendlela ze-molecular ezibangela ukulahleka kobunzima kunye nokufa kwe-corn borer.
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