Ukutholwa nokusabalalisa izakhamzimba kuhlanganisa ukudla izinambuzane kanye nezici zomlando wokuphila. Ukuze kulungiswe ukuntuleka kwezakhamzimba ezithile ezigabeni ezahlukene zokuphila, izinambuzane zingathola lezi zakhamzimba ngokudla okwengeziwe, isibonelo, ngokudla uketshezi lwezilwane ezinomgogodla enkambisweni eyaziwa ngokuthi ama-puddle. Umiyane i-Anopheles arabiani ubonakala ungondlekile kahle futhi, ngakho-ke, udinga izakhamzimba kokubili ukusetshenziswa kwe-metabolism kanye nokuzala. Inhloso yalolu cwaningo kwakuwukuhlola ukuthi ngabe ukuvuvukala kwe-An. arabiensis emchameni wenkomo ukuze kutholakale izakhamzimba kuthuthukisa izici zomlando wokuphila.
Qinisekisa ukuthi iphephile. I-arabiensis yakhangwa yiphunga lomchamo wenkomo omusha, ongamahora angu-24, amahora angu-72, kanye nowamahora angu-168 ubudala, kanye nezinsikazi ezifuna futhi ezinceliswa igazi (ezingamahora angu-48 ngemva kokudla kwegazi) zalinganiswa ku-olfactometer ye-Y-tube, kanti izinsikazi ezikhulelwe zahlolwa ukuze kuhlolwe ukuzala. Kwabe sekusetshenziswa ukuhlaziywa kwamakhemikhali kanye ne-electrophysiological ukuhlonza amakhemikhali asebenzayo emchameni wenkomo kuzo zonke izigaba ezine zeminyaka. Izingxube zokwenziwa zamakhemikhali asebenzayo zahlolwa ezivivinyweni ze-Y-tube kanye nasensimini. Ukuze kuhlolwe umchamo wenkomo kanye ne-compound urea eyinhloko equkethe i-nitrogen njengokudla okungeziwe okungenzeka kube yi-vectors ye-malaria, kwalinganiswa imingcele yokudla kanye nezici zomlando wokuphila. Kwahlolwa isilinganiso somiyane besifazane kanye nenani lomchamo wenkomo kanye ne-urea emuncwayo. Ngemva kokudla, izinsikazi zahlolwa ukuze zisinde, zahlanganiswa, futhi zazala.
Funa igazi nokudla komninimzi. Ezifundweni zelebhu nasensimini, ama-Arabhu akhangwa yiphunga lemvelo nelokwenziwa lomchamo wenkomo omusha nowasebekhulile. Abesifazane abakhulelwe babengenandaba nezimpendulo zomchamo wenkomo ezindaweni zokuzalela. Abesifazane abafuna umninimzi nabamunca igazi babamba umchamo wenkomo kanye ne-urea ngenkuthalo futhi babela lezi zinsiza ngokwemibono yomlando wokuphila njengomsebenzi wesimo somzimba sokundiza, ukusinda, noma ukuzala.
Ukutholwa kanye nokusatshalaliswa komchamo wenkomo we-Anopheles ukuze kuthuthukiswe izici zomlando wokuphila. Ukondla okwengeziwe komchamo wenkomo kuthinta amandla e-vector ngqo ngokwandisa ukusinda kwansuku zonke kanye nobuningi be-vector, futhi ngokungaqondile ngokushintsha umsebenzi wokundiza ngakho-ke kufanele kucatshangelwe kumamodeli azayo.
Ukutholwa nokusabalalisa izakhamzimba kuhlanganisa ukudla kwezinambuzane kanye nezici zomlando wokuphila [1,2,3].Izinambuzane ziyakwazi ukukhetha nokuthola ukudla futhi zenze ukudla okunxephezelayo ngokusekelwe ekutholakaleni kokudla kanye nezidingo zezakhamzimba [1, 3].Ukusatshalaliswa kwezakhamzimba kuncike enkambisweni yomlando wokuphila futhi kungaholela ezidingweni ezahlukene zekhwalithi yokudla kanye nobuningi ezigabeni ezahlukene zokuphila kwezinambuzane [1, 2].Ukuze kulungiswe ukuntuleka kwezakhamzimba ezithile, izinambuzane zingathola lezi zakhamzimba ngokudla okwengeziwe, njengodaka, indle ehlukahlukene kanye nokuphuma kwezilwane ezinomgogodla, kanye nenyama efile, inqubo eyaziwa ngokuthi ama-puddles [2].Nakuba izinhlobo ezahlukene zebhabhathane kanye ne-moth zichazwa ngokuyinhloko, imigodi yokunisela nayo iyenzeka kwezinye izinhlobo zezinambuzane, futhi ukukhangwa nokudla lezi zinhlobo zezinsiza kungaba nemiphumela ebalulekile empilweni nakwezinye izici zomlando wokuphila [2, 4, 5, 6] ,7].Umiyane we-malaria i-Anopheles gambiae sensu lato (sl) uvela njengomuntu omdala 'ongondlekile' [8], ngakho-ke ukunisela kungadlala indima ebalulekile ezicini zomlando wokuphila kwawo, kodwa lokhu kuziphatha kuze kube manje akunakwa. Ukusetshenziswa kokushukuma njengendlela yokwandisa ukudla okunomsoco kulesi sithuthi esibalulekile kudinga ukunakwa njengoba lokhu kungaba nemiphumela ebalulekile yezifo.
Ukudla i-nitrogen komiyane besifazane abadala i-Anopheles kulinganiselwe ngenxa yokugcinwa kwama-calorie aphansi athathwe kusukela esigabeni sama-larval kanye nokusetshenziswa okungalungile kokudla kwegazi [9]. I-Ann.gambiae sl yesifazane ivame ukuvala lokhu ngokunezela ngokudla kwegazi okwengeziwe [10, 11], ngaleyo ndlela ibeke abantu abaningi engozini yokuthola lesi sifo futhi ibeke omiyane engozini enkulu yokuqothulwa. Ngaphandle kwalokho, omiyane bangasebenzisa ukondla okwengeziwe kwendle yezilwane ezinomgogodla ukuze bathole amakhemikhali ane-nitrogen athuthukisa ukuzivumelanisa nezimo kanye nokuhamba kalula, njengoba kuboniswe ezinye izinambuzane [2]. Kulokhu, ukukhanga okunamandla nokuhlukile kolunye uhlobo lwezilwane ezizalwa ngaphakathi kwe-An. I-Gambian sl complex, i-Anopheles arabinis, umchamo wenkomo omusha nomdala [12,13,14], kuyathakazelisa. I-Anopheles arabinis inethuba ekuthandweni kwayo futhi yaziwa ngokuzihlanganisa nokudla izinkomo. Umchamo wenkomo uwumthombo ocebile ngamakhemikhali ane-nitrogen, i-urea ebalelwa ku-50-95% we-nitrogen iyonke emchameni omusha [15, 16].Njengoba umchamo wenkomo uguga, ama-microorganisms asebenzisa lezi zinsiza ukunciphisa ubunzima bama-compounds ane-nitrogen zingakapheli amahora angama-24 [15].Ngokwanda okusheshayo kwe-ammonia, okuhambisana nokwehla kwe-nitrogen ephilayo, ama-microorganisms e-alkalophilic (amaningi awo akhiqiza ama-compounds anobuthi komiyane) ayachuma [15], okungaba yi-Ann.arabiensis yesifazane ekhangwa kakhulu umchamo eneminyaka engamahora angama-24 noma ngaphansi [13, 14].
Kulolu cwaningo, kwafunwa i-Ans ephethwe ngumnikazi kanye negazi. Ngesikhathi somjikelezo wayo wokuqala we-gonadotropin, i-arabiensis yahlolwa ukuthi ithola kanjani amakhemikhali ane-nitrogen, okuhlanganisa ne-urea, ngokuxuba umchamo. Okulandelayo, kwenziwa uchungechunge lwezivivinyo zokuhlola ukuthi omiyane besifazane bawabela kanjani lo mthombo wezakhamzimba ongaba khona ukuze kuthuthukiswe ukusinda, ukuzala kanye nokufuna ukudla okwengeziwe. Okokugcina, iphunga lomchamo wenkomo omusha nomdala lahlolwa ukuze kutholakale ukuthi ngabe lokhu kunikeze izinkomba ezithembekile ze-An ephethwe ngumnikazi kanye negazi. Ekufuneni kwabo lo mthombo wezondlo ongaba khona, i-arabiensis yathola ubudlelwano bamakhemikhali ngemuva kokukhanga okuhlukile okubonwe. Izingxube zephunga lokwenziwa lamakhemikhali e-volatile organic (VOCs) atholakale emchameni ohlala amahora angama-24 zahlolwa kabanzi ngaphansi kwezimo zasensimini, zandisa imiphumela etholakale ngaphansi kwezimo zelebhu futhi zabonisa umphumela wephunga lomchamo wenkomo ezimweni ezahlukene zomzimba. Ukukhangwa komiyane. Imiphumela etholakele iqinisekisa ukuthi i-An. arabiensis ithola futhi isabalalise amakhemikhali ane-nitrogen atholakala emchameni wezilwane ezinomgogodla ukuze ithonye izici zomlando wokuphila. Le miphumela ixoxwa ngomongo wemiphumela engaba khona ye-epidemiological nokuthi ingasetshenziswa kanjani ekuqapheni nasekulawuleni i-vector.
Ama-Anopheles arabicans (uhlobo lwe-Dongola) agcinwe ku-25 ± 2 °C, 65 ± 5% RH kanye nomjikelezo wokukhanya nobumnyama wamahora angu-12:12. Ama-larvae akhuliswa emathileyi epulasitiki (20 cm × 18 cm × 7 cm) agcwaliswe ngamanzi acwecwe futhi ondliwe ukudla kwezinhlanzi ze-Tetramin® (Tetra Werke, Melle, DE). Ama-pupae aqoqwa ezinkomishini ezingama-30 ml (Nolato Hertila, Åstorp, SE) abese edluliselwa ezindlini ze-Bugdorm (30 cm × 30 cm × 30 cm; MegaView Science, Taichung, Taiwan) ukuze kuvunyelwe ukuvela kwabantu abadala. Abantu abadala banikezwa isisombululo se-sucrose esingu-10% ngaphambi kokuphela kwezinsuku ezi-4 ngemva kokuvela (dpe), lapho abesifazane abafuna indawo yokuhlala banikezwa ukudla ngaphambi nje kokuhlolwa, noma babulawa yindlala ubusuku bonke ngamanzi acwecwe ngaphambi kokuhlolwa, njengoba kuchaziwe ngezansi. Abesifazane abasetshenziselwa ukuhlolwa kwe-flight tube babulawa yindlala kuphela. Amahora angu-4-6 ngamanzi ngokushesha. Ukuze kulungiselelwe omiyane abamunca igazi ukuze kuhlolwe i-bioassays elandelayo, abesifazane abangu-4 be-dpe banikezwa igazi lemvu elisusa i-fibrotic (Håtunalab, Bro, SE) kusetshenziswa uhlelo lokuphakela i-membrane (Hemotek Discovery Workshops, Accrington, UK). Abesifazane ababegcwele ngokugcwele babe sebedluliselwa ezindlini zomuntu ngamunye futhi banikezwa ukudla ngqo, njengoba kuchaziwe ngezansi, noma i-10% ye-sucrose ngokushesha izinsuku ezi-3 ngaphambi kokuhlolwa okuchazwe ngezansi. Abesifazane bokugcina basetshenziselwa ukuhlolwa kwe-bioassays nge-flight tube futhi badluliselwa elabhorethri, base bethola amanzi acwengekile ngokushesha amahora angu-4-6 ngaphambi kokuhlolwa.
Ukuhlolwa kokudla kwasetshenziswa ukulinganisa ukusetshenziswa komchamo kanye ne-urea kwabesifazane abadala abangama-Arabhu. Abesifazane abafuna umbungazi futhi abanceliswa igazi banikezwa ukudla okuqukethe umchamo wenkomo omusha nomdala oxutshwe u-1%, amazinga ahlukahlukene e-urea, kanye nezilawuli ezimbili (10% sucrose namanzi) amahora angu-48. Ngaphezu kwalokho, umbala wokudla (1 mg ml-1 xylene cyanide FF; CAS 2650-17-1; Sigma-Aldrich, Stockholm, SE) wengezwe ekudleni futhi wanikezwa ku-matrix engu-4 × 4 kumashubhu e-microcentrifuge angu-250 µl (Aoxygen Scientific, Union City, CA, US; Umfanekiso 1A) Gcwalisa kuze kube semaphethelweni (~300 µl). Ukuze ugweme ukuncintisana phakathi komiyane nemiphumela engaba khona yombala wedayi, beka omiyane abayi-10 esitsheni esikhulu sePetri (12 cm ububanzi kanye no-6 cm ukuphakama; Semadeni, Ostermundigen, CH; Umfanekiso 1A) ebumnyameni obuphelele ku-25 ± 2 cm °C kanye no-65 ± 5% ngokuhlobene. umswakama. Lezi zivivinyo ziphindwe izikhathi ezi-5 kuya kweziyi-10. Ngemva kokuchayeka ekudleni, omiyane bafakwa ku--20 °C kuze kube yilapho kuhlaziywa okwengeziwe.
Bheka umchamo wezinkomo kanye ne-urea emuncwa yi-host kanye ne-Anopheles arabianus yensikazi emunca igazi. Esivivinyweni sokudla (A), omiyane besifazane banikezwe ukudla okuqukethe umchamo wezinkomo omusha nomdala, amazinga ahlukahlukene e-urea, i-sucrose (10%), kanye namanzi acwengekile (H2O). Omiyane besifazane abafuna umbungazi (B) kanye nabesifazane abanceliswa igazi (C) bancela i-sucrose eningi kunanoma yikuphi okunye ukudla okuvivinyiwe. Qaphela ukuthi abesifazane abafuna umbungazi bancela umchamo wezinkomo ongaphansi kwamahora angu-72 ngaphansi komchamo wezinkomo ongaphansi kwamahora angu-168 (B). Okuqukethwe okumaphakathi kwe-nitrogen ephelele (± ukuphambuka okujwayelekile) komchamo kumelelwe esithombeni. Omiyane abafuna umbungazi (D, F) kanye nabesifazane abamunca igazi (E, G) bancela i-urea ngendlela encike kumthamo. Amavolumu aphakathi aphefumulayo (D, E) anamagama ahlukene ezinhlamvu ayehluke kakhulu komunye nomunye (i-ANOVA yendlela eyodwa kusetshenziswa ukuhlaziywa kwe-post hoc kaTukey; p < 0.05). Amabha amaphutha amelela iphutha elijwayelekile le-mean (BE). Umugqa oqondile onemigqa umelela umugqa wokubuyela emuva we-log-linear (F, G)
Ukuze kukhishwe ukudla okumuncwe, omiyane bafakwa ngabanye emapayipini e-microcentrifuge angu-1.5 ml aqukethe amanzi acwecwe angu-230 µl kanti izicubu zaphazamiseka kusetshenziswa i-pestle elahlwayo kanye ne-cordless motor (VWR International, Lund, SE), kwalandela i-centrifugation ku-10 krpm imizuzu eyi-10. I-supernatant (200 µl) yadluliselwa kwi-microplate engu-96-well (Sigma-Aldrich) kwathi i-absorbance (λ620) yatholakala kusetshenziswa i-spectrophotometer-based microplate reader (SPECTROStar® Nano, BMG Labtech, Ortenberg, DE) nm). Ngaphandle kwalokho, omiyane bagaywa ku-1 ml wamanzi acwecwe, angu-900 µl awo adluliselwa kwi-cuvette ukuze kuhlaziywe i-spectrophotometric (λ 620 nm; UV 1800, Shimadzu, Kista, SE). Ukuze kulinganiswe ukudla okudliwayo, kwalungiswa ijika elijwayelekile ngokuncibilikisa okulandelanayo ukuze kutholakale u-0.2 µl kuya ku-2.4. µl ye-1 mg ml-1 xylene cyanide. Ngemuva kwalokho, kwasetshenziswa ubuningi bokukhanya kwamazinga aziwayo kadayi ukuze kutholakale inani lokudla omiyane ngamunye okudlayo.
Idatha yevolumu ihlaziywe kusetshenziswa ukuhlaziywa kwe-variance yendlela eyodwa (ANOVA) kulandelwe ukuqhathaniswa kukaTukey kwe-post hoc pairwise (JMP Pro, v14.0.0, SAS Institute Inc., Cary, NC, US, 1989–2007). Ukuhlaziywa kokuhlehla komugqa kuchaze ukungenisa kwe-urea okuncike ekugxilweni kanye nezimpendulo eziqhathanisiwe phakathi komiyane abafuna umbungazi kanye nabamunca igazi (GraphPad Prism v8.0.0 ye-Mac, GraphPad Software, San Diego, CA, US).
Cishe amasampula omchamo angama-20 µl avela eqenjini ngalinye lobudala aboshwe ku-Chromosorb® W/AW (10 mg 80/100 mesh, Sigma Aldrich) futhi afakwa kuma-capsule e-tin (8 mm × 5 mm). Ama-capsule afakwa ekamelweni lokushisa le-CHNS/O analyzer (Flash 2000, Thermo Fisher Scientific, Waltham, MA, US) ukuze kutholakale okuqukethwe kwe-nitrogen kumchamo omusha nowokuguga ngokwenqubo yomenzi. I-nitrogen ephelele (g N l-1) yalinganiswa ngokusekelwe kumazinga e-urea aziwayo asetshenziswa njengendinganiso.
Ukuze kuhlolwe umthelela wokudla ekusindeni kwabesifazane abafuna umninikhaya kanye nokumunca igazi, omiyane bafakwa ngabanye ezitsheni ezinkulu zePetri (ububanzi obungu-12 cm kanye nobude obungu-6 cm; Semadeni) ngembobo embozwe nge-mesh esivalweni (ububanzi obungu-3 cm) ukuze kutholakale umoya nokudla. Ukudla kwanikezwa ngqo ngemva kwe-4 dpe futhi kwafaka umchamo wenkomo omusha nowokuguga ongu-1% oxutshwe, amazinga amane e-urea, kanye nezilawuli ezimbili, i-sucrose engu-10% namanzi. Ukudla ngakunye kwafakwa ngepayipi ku-tampon yamazinyo (DAB Dental AB, Upplands Väsby, SE) efakwe kwisirinji engu-5 ml (Thermo Fisher Scientific, Gothenburg, SE), i-plunger yasuswa, yabekwa phezu kwesitsha se-petri (isithombe 1).1A). Shintsha ukudla kwakho nsuku zonke. Gcina ilebhu njengoba kuchaziwe ngenhla. Omiyane abasindayo babalwa kabili ngosuku, kuyilapho omiyane abafile balahlwa kuze kube yilapho umiyane wokugcina ushona (n = 40 ngokwelashwa ngakunye). Ukusinda komiyane abadla ukudla okuhlukahlukene kwahlaziywa ngokwezibalo kusetshenziswa ama-curve okusinda eKaplan-Meyer kanye izivivinyo ze-log-rank ukuqhathanisa ukuqhathanisa ukusatshalaliswa kokusinda phakathi kokudla (IBM SPSS Statistics 24.0.0.0).
Imishini yokundiza yomiyane eyenziwe ngokwezifiso esekelwe ku-Attisano et al.[17], eyenziwe ngamaphaneli e-acrylic acacile angu-5 mm ubukhulu (ububanzi obungu-10 cm x ubude obungu-10 cm x ukuphakama okungu-10 cm) ngaphandle kwamaphaneli angaphambili nangemuva (Isithombe 3: phezulu). Inhlangano ejikelezayo enepayipi eliqondile elenziwe ngekholomu ye-gas chromatography (0.25 mm id; 7.5 cm L) eneziphetho ezinamathele enalitini yezinambuzane elenga phakathi kwamamagnethi amabili e-neodymium aqhelelene ngo-9 cm. Ipayipi elivundlile elenziwe ngezinto ezifanayo (6.5 cm L) lahlukanisa ipayipi eliqondile ukuze lenze ingalo eboshiwe kanye nengalo eyayiphethe ucezu oluncane lwe-aluminium foil njengesignali yokuphazamisa ukukhanya.
Izinsikazi ezilambile amahora angama-24 zanikezwa ukudla okungenhla imizuzu engama-30 ngaphambi kokuvinjelwa. Omiyane besifazane abaphakelwa ngokugcwele babe sebefakwa i-anesthesia ngayinye eqhweni imizuzu emi-2-3 bese benamathiselwa ezikhonkwaneni zezinambuzane nge-beeswax (Joel Svenssons Vaxfabrik AB, Munka Ljungby, SE) bese beboshwa ezingalweni zamapayipi avundlile. I-Flying Mill. Ukuguquguquka ngendiza ngayinye kwaqoshwa yi-data logger eyakhelwe ngokwezifiso, yabe igcinwa futhi iboniswa kusetshenziswa isoftware ye-PC-Lab 2000™ (v4.01; Velleman, Gavere, BE). I-flight mill yafakwa ekamelweni elilawulwa yisimo sezulu (12 h:12 h, ukukhanya: mnyama, 25 ± 2 °C, 65 ± 5% RH).
Ukuze kubonwe ngeso lengqondo iphethini yomsebenzi wokundiza, ibanga eliphelele eligijinywe (m) kanye nenani eliphelele lemisebenzi yokundiza elandelanayo kubalwe ngehora esikhathini samahora angama-24. Ngaphezu kwalokho, amabanga ajwayelekile agijinywe ngabesifazane ngabanye aqhathaniswa phakathi kokwelashwa futhi ahlaziywa kusetshenziswa ukuhlaziywa kwe-ANOVA kanye ne-Tukey's post hoc (JMP Pro, v14.0.0, SAS Institute Inc.), lapho ibanga elijwayelekile lalibhekwa njengokuguquguquka okuxhomeke kukho, kuyilapho ukwelashwa kuyisici esizimele. Ngaphezu kwalokho, inani elijwayelekile lemijikelezo libalwa ngokunyuka kwemizuzu eyi-10.
Ukuhlola umphumela wokudla ekusebenzeni kokuzala kwe-An.arabiensis, abesifazane abayisithupha (4 dpe) badluliselwa ngqo ezindlini ze-Bugdorm (30 cm × 30 cm × 30 cm) ngemva kokuqoqwa kwegazi base benikeza ukudla kokuhlola amahora angu-48 njengoba kuchaziwe ngenhla. Ukudla kwabe sekususwa futhi kwanikezwa izinkomishi zokuzala (30 ml; Nolato Hertila) ezigcwaliswe ngamanzi angu-20 ml acwecwe ngosuku lwesithathu amahora angu-48, zishintsha njalo emahoreni angu-24. Phinda uhlelo ngalunye lokudla izikhathi ezingu-20-50. Amaqanda abalwa futhi aqoshwa endlini ngayinye yokuhlola. Amasampula amancane amaqanda asetshenziswa ukuhlola usayizi omaphakathi kanye nokwehluka kobude bamaqanda ngamanye (n ≥ 200 ngokudla ngakunye) kusetshenziswa i-Dialux-20 microscope (DM1000; Ernst Leitz Wetzlar, Wetzlar, DE) efakwe i-Leica Camera (DFC) 320 R2; Leica Microsystems Ltd., DE). Amaqanda asele agcinwe ekamelweni elilawulwa isimo sezulu ngaphansi kwezimo ezijwayelekile zokukhulisa amahora angama-24, futhi kwalinganiswa isampula encane yezibungu zokuqala ezisanda kuvela (n ≥ 200 ngokudla ngakunye), njengoba kuchaziwe ngenhla. Inani lamaqanda kanye nosayizi wamaqanda nezibungu kwaqhathaniswa phakathi kokwelashwa kanye nokusebenzisa ukuhlaziywa kwe-ANOVA kanye ne-Tukey's post hoc (JMP Pro, v14.0.0, SAS Institute Inc.).
Ama-volatiles e-Headspace avela ku-fresh (ihora eli-1 ngemuva kokuthatha isampula), umchamo ogugile wamahora angama-24, amahora angama-72 kanye namahora angama-168 aqoqwe kumasampula aqoqwe ezinkomeni ze-Zebu, e-Arsi races. Ukuze kube lula, amasampula omchamo aqoqwe ekuseni kakhulu ngenkathi izinkomo zisesesibayeni. Amasampula omchamo aqoqwe kubantu abayi-10 kanti ama-100-200 ml esampula ngayinye adluliselwa ezikhwameni zokubhaka ze-polyamide ngazinye (i-Toppits Cofresco, i-Frischhalteprodukte GmbH kanye ne-Co., e-Minden, DE) ku-polyamide engamalitha ama-3 enesembozo Kuma-drum epulasitiki e-vinyl chloride. Ama-volatiles e-Headspace avela kusampula ngayinye yomchamo wezinkomo aqoqwe ngqo (okusha) noma ngemva kokuvuthwa ekushiseni kwegumbi amahora angama-24, amahora angama-72 kanye namahora ayi-168, okungukuthi isampula ngayinye yomchamo yayimele iqembu ngalinye leminyaka.
Ekuqoqweni kwe-headspace volatiles, kwasetshenziswa uhlelo lwe-closed-loop ukuhambisa umfudlana wegesi ohlungiwe ngekhabhoni (100 ml min-1) ngesikhwama se-polyamide uye kukholamu yokumunca amahora angu-2.5 ngokusebenzisa iphampu yokumunca i-diaphragm (KNF Neuberger, Freiburg, DE). Njengokulawula, ukuqoqwa kwe-headspace kwenziwa kusuka esikhwameni se-polyamide esingenalutho. Ikholomu yokumunca yenziwe nge-Teflon tube (5.5 cm x 3 mm id) equkethe ama-35 mg we-Porapak Q (50/80 mesh; Waters Associates, Milford, MA, US) phakathi kwama-plugs e-glass wool. Ngaphambi kokusebenzisa, ikholomu yahlanzwa nge-1 ml ye-n-hexane ephinde yahlungwa (Merck, Darmstadt, DE) kanye ne-1 ml ye-pentane (99.0% solvent GC grade, Sigma Aldrich). Ama-volatiles afakwe i-adsorbed asuswa nge-400 μl ye-pentane. Amaqoqo e-Headspace ahlanganiswa abese egcinwa ku--20°C kuze kusetshenziswe isikhathi eside. ukuhlaziywa.
Izimpendulo zokuziphatha ze-An-host-seeking kanye ne-blood-eating-blood extracts An.Headspace volatile extracts eziqoqwe kumchamo omusha, wamahora angama-24, amahora angama-72, kanye namahora ayi-168 ubudala zihlaziywe ukuze kutholakale i-volatile extracts ezivela komiyane be-Arabidopsis kusetshenziswa i-olfactometer ye-glass tube eqondile [18]. Ukuhlolwa kwenziwe ngesikhathi se-ZT 13-15, isikhathi esiphezulu somsebenzi ka-An wokufuna ikhaya. I-Arab [19]. I-olfactometer ye-glass tube (80 cm × 9.5 cm id) ikhanyiswe ngo-3 ± 1 lx wokukhanya okubomvu okuvela phezulu. Ukugeleza komoya okuhlungiwe kwamalahle nokunomswakama (25 ± 2 °C, 65 ± 2% umswakama ohlobene) kudlule ukuhlolwa kwe-bioassay ku-30 ​​cm s-1. Umoya udlula ochungechungeni lwezikrini zensimbi engagqwali, okudala ukugeleza kwe-laminar kanye nesakhiwo se-plume esifanayo. I-dental tampon dispenser (4 cm × 1 cm; L:D; DAB Dental AB), elenga ku-coil engu-5 cm ekugcineni komoya i-olfactometer, enezinguquko zesikhuthazi njalo ngemizuzu emi-5. Ekuhlaziyeni, ama-10 μl e-headspace extract ngayinye, ancishiswe ngo-1:10, asetshenziswa njengesikhuthazi. Inani elilinganayo le-pentane lasetshenziswa njengokulawula. Omiyane abafuna umninikhaya noma abamunca igazi bafakwa emakhejini okukhululwa ngamanye amahora ama-2-3 ngaphambi kokuqala kokuhlolwa. Ikheji lokukhululwa labekwa ohlangothini lomoya we-olfactometer, futhi omiyane bavunyelwa ukujwayela umzuzu owodwa, bese i-valve yebhabhathane yekheji yavulwa ukuze ikhululeke. Ukukhangwa kokwelashwa noma ukulawula kwahlaziywa njengesilinganiso somiyane abathintane nomthombo zingakapheli imizuzu emi-5 yokukhululwa. I-headspace volatile extract ngayinye kanye nokulawula kwaphindwa okungenani izikhathi ezingama-30, futhi ukuze kugwenywe imiphumela yanoma yiluphi usuku olulodwa, inani elifanayo lokwelashwa kanye nokulawula lahlolwa ngosuku ngalunye lokuhlola. Funa izimpendulo ezivela kumninikhaya kanye nokunikezwa igazi. Amasethi e-Arabic vs headspace ahlaziywa kusetshenziswa i-nominal logistic regression elandelwa ukuqhathanisa okubili kwezilinganiso ezingavamile (JMP Pro, v14.0.0, SAS Institute Inc.).
Impendulo ka-An yokuzalela. Izingcezu zekhanda ezivela kumchamo wezinkomo omusha nomdala zahlaziywa emakhejini aseBugdorm (30 cm × 30 cm × 30 cm; MegaView Science). Izinkomishi zepulasitiki (30 mL; Nolato Hertila) ezigcwele amanzi acwecwe angu-20 mL zanikeza isisekelo sokuzalela futhi zabekwa emakhoneni aphambene ekheji, ziqhelelene ngamasentimitha angu-24. Izinkomishi zokwelapha zalungiswa nge-10 μl ye-headspace ngayinye encibilikisiwe nge-1:10 dilution. Inani elilinganayo le-pentane lasetshenziswa ukulungisa indebe yokulawula. Izinkomishi zokwelapha nezokulawula zashintshaniswa phakathi kokuhlolwa ngakunye ukuze kulawulwe imiphumela yesikhundla. Izinsikazi eziyishumi ezondliwe ngegazi zakhishwa emakhejini okuhlola ku-ZT 9-11 kanti amaqanda ezinkomishini abalwa emahoreni angu-24 kamuva. Ifomula yokubala inkomba yokuzalela yile: (inani lamaqanda abekwe endebeni yokwelapha - inani lamaqanda abekwe endebeni yokulawula)/(inani eliphelele lamaqanda abekwe). Ukwelashwa ngakunye kwaphindwa izikhathi ezingu-8.
Ukuhlaziywa kwephethini lokutholwa kwephethini ye-gas chromatographic kanye ne-electron antenna (GC-EAD) ye-An.arabiensis yesifazane kwenziwa njengoba kuchaziwe ngaphambilini [20]. Kafushane, iziqephu ezintsha ze-headspace volatile zahlukaniswa kusetshenziswa i-Agilent Technologies 6890 GC (Santa Clara, CA, US) efakwe ikholomu ye-HP-5 (30 m × 0.25 mm id, ubukhulu befilimu obungu-0.25 μm, i-Agilent Technologies). kanye nomchamo ogugayo. I-hydrogen yasetshenziswa njengesigaba sokuhamba ngesilinganiso sokugeleza okuqondile esingu-45 cm s-1. Isampula ngayinye (2 μl) yajovwa imizuzwana engama-30 kwimodi ehlukanisiwe enezinga lokushisa lokungena elingu-225 °C. Izinga lokushisa le-GC oven lahlelwa kusukela ku-35 °C (ukubamba imizuzu emi-3) kuya ku-300 °C (ukubamba imizuzu eyi-10) ku-10 °C min-1. Ku-GC effluent splitter, kwanezelwa ama-psi angu-4 e-nitrogen futhi kwahlukaniswa ngo-1:1 ku-Gerstel 3D/2 low dead volume cross (Gerstel, Mülheim, DE) phakathi kwe-flame ionization detector kanye ne-EAD. I-GC effluent capillary ye-EAD yadlula ngomugqa wokudlulisa we-Gerstel ODP-2, olandelela izinga lokushisa le-GC oven kanye no-5 °C, ungene epayipini lengilazi (10 cm × 8 mm), lapho yaxutshwa khona nomoya ohlungiwe ngekhabhoni, omanzi (1.5 l min−1). I-antenna yafakwa 0.5 cm ukusuka ekuphumeni kwepayipi. Umiyane ngamunye wawunesibalo esisodwa esiphindaphindayo, kanti komiyane abafuna umninimzi, okungenani kwaphindaphindwa kathathu kumasampula omchamo wobudala ngabunye.
Ukuhlonza amakhemikhali asebenzayo ezingxenyeni zekhanda lomchamo wezinkomo omusha nomdala kusetshenziswa i-GC ehlanganisiwe kanye ne-mass spectrometer (GC-MS; 6890 GC kanye ne-5975 MS; Agilent Technologies) ukuze kutholakale izimpendulo ze-antenna ekuhlaziyweni kwe-GC-EAD, esebenza ngemodi ye-electron impact ionization ku-70 eV. I-GC yayifakwe ikholomu ye-silica capillary ehlanganiswe ne-HP-5MS UI (ububanzi obungaphakathi obungu-60 m × 0.25 mm, ubukhulu befilimu obungu-0.25 μm) kusetshenziswa i-helium njengesigaba esihambayo esinesilinganiso sokugeleza okuqondile esingu-35 cm s-1. Isampula engu-2 μl yajovwa kusetshenziswa izilungiselelo ezifanayo ze-injector kanye nokushisa kwe-oven njengokuhlaziywa kwe-GC-EAD. Amakhemikhali atholwe ngokusekelwe esikhathini sawo sokugcina (inkomba ye-Kovát) kanye ne-mass spectra uma kuqhathaniswa nomtapo wolwazi wangokwezifiso kanye nomtapo wolwazi we-NIST14 (Agilent). Amakhemikhali atholwe aqinisekiswa ngokujova amazinga angempela (Ifayela Elingeziwe 1: Ithebula S2). Ukuze kulinganiswe, i-heptyl i-acetate (10 ng, ubumsulwa bamakhemikhali obungu-99.8%, i-Aldrich) yajovwa njengendinganiso yangaphandle.
Ukuhlola ukusebenza kahle kwengxube yephunga lokwenziwa equkethe amakhemikhali asebenzayo atholakala emchameni omusha nowokuguga ukuze kudonswe i-Ans.arabiensis efuna umninimzi futhi emunca igazi, kusetshenziswa i-olfactometer kanye nephrothokholi efanayo njengoba kushiwo ngenhla. Izingxube zokwenziwa zilingise ukwakheka kanye nezilinganiso zamakhemikhali ezikhishweni ezihlanganisiwe ze-headspace volatile zomchamo omusha, wamahora angu-24, wamahora angu-48, wamahora angu-72, kanye nowamahora angu-168 (Isithombe 5D-G; Ifayela Elingeziwe 1: Ithebula S2). Ukuze kuhlaziywe, sebenzisa i-10 μl yengxube yokwenziwa ephelele engu-1:100, enesilinganiso sokukhululwa esisukela cishe ku-140-2400 ng h-1, ukuze kuhlolwe ukukhanga komiyane abamukelayo nabamunca igazi. Ngemuva kwalokho, ukuhlolwa kwenziwa ezingxubeni eziphelele, lapho kususwa khona izingxube ezikhiphayo zamakhemikhali angawodwa engxubeni ephelele. Funa izimpendulo ezivela kumninimzi kanye ne-Ans ezidla igazi. Izingxube ze-Arab vs zokwenziwa kanye nezikhiphayo zahlaziywa kusetshenziswa ukuhlehliswa kwe-logistic okubizwa ngokuthi yi-nominal kulandelwa ukuqhathanisa okubili kokungajwayelekile. izilinganiso (JMP Pro, v14.0.0, SAS Institute Inc.).
Ukuhlola ukuthi umchamo wezinkomo ungasebenza yini njengendawo yokuhlala yomiyane we-malaria, umchamo wezinkomo omusha nogugile, oqoqwe njengoba kuchaziwe ngenhla, bese amanzi afakwa emabhakedeni ama-3 l ahlanganisiwe (100 ml) bese efakwa ezicuphweni zokudoba. (Inguqulo ye-BG-HDT; i-BioGents, i-Regensburg, i-DE). Izicupho eziyishumi zibekwe eceleni kwamamitha angu-50 emadlelweni, amamitha angu-400 ukusuka emphakathini wasemakhaya (iSilay, i-Ethiopia, 5°53´24´´N, 37°29´24´´E) futhi kungekho zinkomo, ezindaweni zokuzalela ezihlala njalo nasemadolobheni. Izicupho ezinhlanu zashiswa ukuze kulingiswe ukuba khona komninimzi, kanti izicupho ezinhlanu zashiywa zingashiswanga. Indawo ngayinye yokwelapha ijikeleziswa ubusuku ngabunye ubusuku obuyisihlanu. Izinombolo zomiyane ezithathwe ezicuphweni ezifakwe umchamo wobudala obuhlukene zaqhathaniswa kusetshenziswa i-logistic regression ngokusatshalaliswa kwe-beta binomial (i-JMP Pro, v14.0.0, i-SAS Institute Inc.).
Edolobhaneni elivame ukuba khona umalaleveva eduze kwedolobha laseMaki, esifundeni sase-Oromia, e-Ethiopia (8° 11′ 08″ N, 38° 81′ 70″ E; Isithombe 6A). Ucwaningo lwenziwe phakathi kuka-Agasti naphakathi kukaSepthemba ngaphambi kokufuthwa kwezindlu okusele minyaka yonke, kanye nesizini ende yemvula. Izindlu ezinhlanu (eziqhelelene ngamamitha angu-20-50) ezisemaphethelweni edolobhana zakhethwa kulolu cwaningo (Isithombe 6A). Izindlela ezisetshenzisiwe ukukhetha izindlu yilezi: azivunyelwe izilwane endlini, azivunyelwe ukupheka endlini (ukukhipha izinkuni noma amalahle) (okungenani ngesikhathi sokuhlolwa), kanye nezindlu ezinabantu ababili kuphela, abalala ezibulala-zinambuzane. ngaphansi kwenethi yomiyane elashwe. Imvume yokuziphatha inikezwe yiBhodi Yokubuyekezwa Kwezimiso Zokuziphatha Zocwaningo Lwezikhungo (IRB/022/2016) ye-Faculty of Natural Sciences (CNS-IRB), e-Addis Ababa University, ngokuhambisana neziqondiso ezisungulwe yi-World Medical Association Declaration of Helsinki. Imvume evela enhloko yomndeni ngamunye itholakale ngosizo lwabasebenzi bezempilo. Yonke inqubo ivunyelwe abaphathi bendawo ezingeni lesifunda nelewadi ('kebele'). Umklamo wokuhlola ulandele umklamo wesikwele sesiLatini esingu-2 × 2, lapho izingxube zokwenziwa kanye nezilawuli zanikezwa izindlu ezibhangqiwe ngobusuku bokuqala futhi zashintshaniswa phakathi kwezindlu ngobusuku bokuhlola obulandelayo. Le nqubo yaphindwa izikhathi eziyishumi. Ngaphezu kwalokho, ukuze kulinganiswe umsebenzi womiyane ezindlini ezikhethiwe, izicupho ze-CDC zabekwa ukuthi zisebenze ubusuku obuhlanu obulandelanayo ekuqaleni, maphakathi nasekupheleni kokuhlolwa kwensimu ngesikhathi esifanayo sosuku.
Ingxube yokwenziwa equkethe amakhemikhali ayisithupha asebenzayo yancibilikiswa ku-heptane (97.0% solvent GC grade, Sigma Aldrich) yabe isikhishwa ku-140 ng h-1 kusetshenziswa i-cotton wick dispenser [20]. I-wick dispenser yavumela wonke amakhemikhali ukuthi akhishwe ngokulingana phakathi nokuhlolwa kwamahora ayi-12. I-Heptane yasetshenziswa njengendlela yokulawula. I-vial yalengiswa eduze kwendawo yokungena ye-Centers for Disease Control and Prevention (CDC) light trap (John W. Hock Company, Gainesville, FL, US; Isithombe 6A). Izingibe zalengiswa 0.8 - 1 m ngaphezu komhlaba, eduze konyawo lombhede, kanti isisebenzi sokuzithandela salala ngaphansi kwenethi yomiyane engalashwanga futhi sasebenza phakathi kuka-18:00 no-06:30. Omiyane ababanjwe ubulili nesimo somzimba (abangondli, abadliwe, abakhulelwe kancane, nabakhulelwe [21] bahlolwa kamuva kusetshenziswa ukuhlaziywa kwe-polymerase chain reaction (PCR) ukuze kutholakale uhlobo oluhlonzwe ngokwesimo se-morphological njenge-A. gambiae sl. Amalungu e-complex [23]. Esifundweni sasensimini, ukubanjwa kwezicupho zezindlu ezibhangqiwe kwahlaziywa kusetshenziswa imodeli yokulingana kwempahla ejwayelekile, lapho ukukhanga kwakuyi-variable exhomeke khona kanye nokwelashwa (ingxube yokwenziwa vs ukulawula) kwaba umphumela ohleliwe (JMP® 14.0. 0. SAS Institute Inc.). Lapha, sibika amanani e-χ2 kanye ne-p kusukela ekuhlolweni kwesilinganiso sokwenzeka.
Hlola ukuthi iphephile yini. I-arabiensis ikwazile ukuthola umchamo, umthombo wayo oyinhloko we-nitrogen, i-urea, ngokuwudla ngokuqondile, zingakapheli amahora angu-48 ngemuva kokuphathwa izinsuku ezi-4 ngemuva kokuhlolwa kokudla kwabesifazane (dpe) kokufuna umninimzi kanye nokuncela igazi (Isithombe 1A). Bobabili abesifazane abafuna umninimzi kanye nabamunca igazi bamunca i-sucrose eningi kakhulu kunanoma yikuphi okunye ukudla noma amanzi (F(5,426) = 20.15, p < 0.0001 kanye no-F(5,299) = 56.00, p < 0.0001, ngokulandelana; Isithombe 1B,C). Ngaphezu kwalokho, abesifazane abafuna umninimzi badle kancane emchameni emahoreni angu-72 uma kuqhathaniswa nomchamo emahoreni angu-168 (Isithombe 1B). Lapho benikezwa ukudla okuqukethe i-urea, abesifazane abafuna umninimzi bamunca inani elikhulu kakhulu le-urea ku-2.69 mM uma kuqhathaniswa nazo zonke ezinye izingxube namanzi, kuyilapho kungenakuhlukaniswa ne-sucrose engu-10% (F(10,813) = 15.72, p < 0.0001; Umfanekiso 1D). Lokhu bekungafani nokusabela kwabesifazane abanceliswa ngegazi, ababevame ukumunca ukudla okuqukethe i-urea kakhulu kunamanzi, yize kungaphansi kakhulu kwe-10% ye-sucrose (F(10,557) = 78.35, p < 0.0001; Umfanekiso 1).1E).Ngaphezu kwalokho, lapho kuqhathaniswa phakathi kwezimo ezimbili zomzimba, abesifazane abane-phlebotomized bamunca i-urea eningi kunabesifazane abafuna umninikhaya ngamanani aphansi kakhulu, futhi laba besifazane bamunca amanani afanayo e-urea ngamanani aphezulu (F(1,953)= 78.82, p < 0.0001; Umfanekiso 1F, G).Ngenkathi ukudla okuvela ekudleni okuqukethe i-urea kubonakala kunenani elihle kakhulu (Umfanekiso 1D,E), abesifazane kuzo zombili izimo zomzimba bakwazile ukulinganisa inani le-urea elimuncwa kulo lonke uhla lwamazinga e-urea ngendlela ye-log-linear (Umfanekiso 1F,G). ).Ngokunjalo, omiyane babonakala belawula ukumuncwa kwe-nitrogen yabo ngokulawula inani lomchamo omuncwayo, njengoba inani le-nitrogen emchameni libonakala enanini elimuncwayo (Isithombe 1B, C kanye ne-B insets).
Ukuze kuhlolwe imiphumela yomchamo kanye ne-urea ekusindeni komiyane abafuna umninikhaya kanye nabamunca igazi, abesifazane banikwa umchamo wabo bonke ubudala (okusha, amahora angama-24, amahora angama-72, kanye namahora ayi-168 ngemuva kokudonswa) kanye nohlu lwamazinga e-urea, kanye namanzi acwengekile kanye ne-sucrose engu-10% esetshenziswa njengokulawula (Isithombe 2A). Lokhu kuhlaziywa kokusinda kubonise ukuthi ukudla kube nomthelela omkhulu ekusindeni okuphelele kwabesifazane abafuna umninikhaya (umchamo: χ2 = 108.5, df = 5, p < 0.0001; urea: χ2 = 122.8, df = 5, p < 0.0001; Isithombe 2B, C) kanye nabesifazane abanceliswa igazi (umchamo: χ2 = 93.0, df = 5, p < 0.0001; urea: χ2 = 137.9, df = 5, p < 0.0001; Isithombe 2D,E).Kuzo zonke izivivinyo, abesifazane abaphakelwa umchamo, i-urea, kanye namanzi babenezinga lokusinda eliphansi kakhulu uma kuqhathaniswa nabesifazane abaphakelwa ukudla kwe-sucrose (Isithombe 2B-E). Abesifazane abafuna umbungazi abaphakelwa umchamo omusha nowomile babonise amazinga okusinda ahlukene, lapho labo abaphakelwa umchamo omude amahora angu-72 (p = 0.016) benethuba eliphansi kakhulu lokusinda (Isithombe 2B). Ngaphezu kwalokho, abesifazane abafuna umbungazi abaphakelwa i-urea engu-135 mM baphile isikhathi eside kunokulawula amanzi (p < 0.04) (Isithombe 2C). Uma kuqhathaniswa namanzi, abesifazane abaphakelwa umchamo omusha kanye nomchamo wamahora angu-24 baphile isikhathi eside (p = 0.001 kanye no-p = 0.012, ngokulandelana; Isithombe 2D), kuyilapho abesifazane abaphakelwa umchamo wamahora angu-72 baphile isikhathi eside kunalabo abaphakelwa umchamo omusha omfushane wesifazane kanye nomchamo onamahora angu-24 ubudala (p < 0.0001 kanye no-p = 0.013, ngokulandelana; Isithombe 2D).Lapho bephakelwa i-urea engu-135 mM, abesifazane abaphakelwa igazi baphile isikhathi eside kunabo bonke amanye amazinga e-urea namanzi (p < 0.013; Umfanekiso 2E).
Ukusinda komiyane abamukelayo kanye nabamunca igazi i-Anopheles arabinis yensikazi edla umchamo wenkomo kanye ne-urea. Ekuhlolweni kwe-bioassay (A), omiyane besifazane banikezwa ukudla okuqukethe umchamo wenkomo omusha nogugile, amazinga ahlukahlukene e-urea, i-sucrose (10%) kanye namanzi acwengekile (H2O). Ukusinda komiyane abafuna ukwamukelayo (B, C) kanye nabamunca igazi (D, E) kwaqoshwa njalo emahoreni ayi-12 kuze kube yilapho bonke abesifazane bedla umchamo (B, D) kanye ne-urea (C, E), kanye nokulawula, i-Sucrose namanzi, sekufile.
Ibanga eliphelele kanye nenani lemijikelezo enqunywe ekuhlolweni kwe-flight mill esikhathini samahora angama-24 kwahluka phakathi komiyane abafuna umninikhaya kanye nabamunca igazi, okwabonisa umsebenzi omncane wokundiza jikelele (Isithombe 3). Omiyane abafuna umninikhaya abanikeza umchamo omusha nogugile noma i-sucrose namanzi babonise amaphethini okundiza ahlukile (Isithombe 3), kanti abesifazane abadla umchamo omusha basebenza kakhulu ekuseni, kuyilapho labo abadla amahora angama-24 kanye nama-168 ubudala Omiyane abadla umchamo babonise amaphethini okundiza ahlukene futhi ikakhulukazi babesebenza usuku ngalunye. Omiyane besifazane abanikeza i-sucrose noma umchamo wamahora angama-72 babonise umsebenzi phakathi nesikhathi samahora angama-24, kuyilapho abesifazane abanikeza amanzi babesebenza kakhulu phakathi nesikhathi. Omiyane abadla i-sucrose babonise amazinga aphezulu omsebenzi ebusuku nasekuseni kakhulu, kuyilapho labo abadla umchamo wamahora angama-72 ubudala bebhekene nokwehla okuqhubekayo komsebenzi emahoreni angama-24 (Isithombe 3).
Ukusebenza kokundiza kwensikazi efuna igazi emunca igazi i-Anopheles arabinis edla umchamo wenkomo kanye ne-urea. Ekuhlolweni komshini wokundiza, omiyane besifazane abadla umchamo wenkomo omusha nomdala, amazinga ahlukahlukene e-urea, i-sucrose (10%), kanye namanzi acwengekile (H2O) aboshwe ezingalweni ezivundlile, ezijikelezayo ngokukhululekile (ngenhla). Kwabesifazane abafuna igazi (kwesobunxele) kanye nabamunca igazi (kwesokudla), ibanga eliphelele kanye nenani lezindiza ngehora lokudla ngakunye esikhathini samahora angama-24 kwaqoshwa (omnyama: grey; ukukhanya: mhlophe). Ibanga elimaphakathi kanye nenani elimaphakathi lezikhathi zokuhlasela kuboniswa ngakwesokudla kwegrafu yomsebenzi we-circadian. Amabha amaphutha amelela iphutha elijwayelekile lesilinganiso. Ukuhlaziywa kwezibalo bheka umbhalo
Ngokuvamile, umsebenzi wokundiza jikelele wezinsikazi ezifuna umninikhaya ulandela iphethini efana neyebanga lokundiza esikhathini samahora angama-24. Ibanga eliphakathi lokundiza lithinteke kakhulu ngokudla okudliwe (F(5, 138) = 28.27, p < 0.0001), kanti izinsikazi ezifuna umninikhaya ezidle umchamo amahora angama-72 zihambe amabanga amade kakhulu uma kuqhathaniswa nazo zonke ezinye izinhlobo zokudla (p < 0.0001), kanti omiyane abanceliswa yi-sucrose bandiza isikhathi eside kunomiyane abasha (p = 0.022) kanye nomchamo oneminyaka engama-24 (p = 0.022). Ngokungafani nephethini yomsebenzi wokundiza echazwe ekudleni komchamo, izinsikazi ezifuna umninikhaya ezinceliswa yi-urea zibonise umsebenzi wokundiza oqhubekayo esikhathini samahora angama-24, zifinyelela phezulu phakathi nengxenye yesibili yesigaba sobumnyama (Isithombe 3). Nakuba amaphethini omsebenzi ayefana, izinsikazi ezifuna umninikhaya ezinceliswa yi-urea zandise kakhulu ibanga eliphakathi lokundiza kuye ngokuthi zingakanani izinhlayiya ezimuncwayo (F(5, 138) = 1310.91, p < 0.0001). Ukufuna umninikhaya abesifazane abanceliswa noma yiliphi izinga le-urea bandiza isikhathi eside kunabesifazane abanceliswa amanzi noma i-sucrose (p < 0.03).
Umsebenzi wokundiza womiyane abamunca igazi wawuzinzile futhi waqhubeka amahora angama-24 kuzo zonke izinhlobo zokudla, kanye nomsebenzi womchamo owandisiwe phakathi nengxenye yesibili yesikhathi sobumnyama kwabesifazane abadla amanzi kanye nakwabesifazane abadla ama-fresh namahora angama-24 ubudala (isithombe 3). Ngenkathi ukudla komchamo kuthinte kakhulu ibanga eliphakathi lokundiza kwabesifazane abadla igazi (F(5, 138) = 4.83, p = 0.0004), ukudla kwe-urea akuzange (F(5, 138) = 1.36, p = 0.24).kanye neminye imichamo kanye nokudla okulawulayo (okusha, p = 0.0091; amahora angama-72, p = 0.0022; amahora ayi-168, p = 0.001; i-sucrose, p = 0.0017; dH2O, p = 0.036).
Imiphumela yokudla komchamo kanye ne-urea ezilinganisweni zokuzala ihlolwe ekuhlolweni kwe-bioassays yamaqanda (Isithombe 4A) futhi yahlolwa ngokwenani lamaqanda abekwe yinsikazi ngayinye, usayizi weqanda, kanye nezibungu zokuqala ezisanda kuchanyuselwa. Inani lamaqanda abekwe. Izinsikazi zase-Arabhu ezifuywa ngomchamo zahlukahluka ngokudla (F(5,222) = 4.38, p = 0.0008; Isithombe 4B). Izinsikazi zondliwa ngomchamo wamahora angu-24, ukudla kwegazi kwabekwa amaqanda amaningi kakhulu kunabesifazane abafuywa okunye ukudla komchamo futhi zazifana nalezo ezifuywa nge-sucrose (Isithombe 4B). Ngokufanayo, usayizi wamaqanda abekwe yizinsikazi ezifuywa ngomchamo wahlukahluka ngokudla (F(5, 209) = 12.85, p < 0.0001), ngomchamo wamahora angu-24 kanye nezinsikazi ezifuywa nge-sucrose zibeka amaqanda amakhulu kakhulu kunabesifazane abafuywa ngamanzi, kuyilapho amaqanda ezinsikazi ezifuywa ngomchamo wamahora angu-168 ayemancane kakhulu (Isithombe 4C). Ngaphezu kwalokho, ukudla komchamo kuthinte kakhulu usayizi wezibungu (F(5, 187) = 7.86, p < 0.0001), lapho izibungu ezinkulu kakhulu ziphuma emaqandeni ezizalelwe yizinsikazi ezineminyaka engu-24 no-72 ubudala ezinceliswa umchamo kunamaqanda ezizalelwe izibungu ezinceliswa umchamo. Izinsikazi ezinceliswa amanzi kanye nezinceliswa umchamo amahora angu-168 (Isithombe 4D).
Ukusebenza kokuzala kwe-Anopheles arabinis yesifazane edla umchamo wenkomo kanye ne-urea. Omiyane besifazane abanceliswa ngegazi banikwa ukudla okuqukethe umchamo wenkomo omusha nomdala, amazinga ahlukahlukene e-urea, i-sucrose (10%), kanye namanzi acwengekile (H2O) amahora angu-48 ngaphambi kokufaka ukuhlolwa kwe-bioassay nokuthola izisekelo zokubeka amaqanda. Amahora angu-48 (A). Inombolo yeqanda (B, E), usayizi weqanda (C, F) kanye nosayizi wezibungu (D, G) kwathinteka kakhulu ekudleni okunikeziwe (umchamo wenkomo: BD; urea: EG). Izindlela zepharamitha ngayinye ezilinganiswe kusetshenziswa amagama ezinhlamvu ahlukene zazihluke kakhulu komunye nomunye (i-ANOVA yendlela eyodwa kusetshenziswa ukuhlaziywa kwe-post hoc kaTukey; p < 0.05). Amabha amaphutha amelela iphutha elijwayelekile lesilinganiso
Njengengxenye enkulu ye-nitrogen yomchamo, i-urea, uma inikezwa njengokudla kwabesifazane abanceliswa ngegazi, yathinta kakhulu imingcele yokuzala kuzo zonke izifundo. Inani lamaqanda abekwe ngabesifazane abanceliswa i-urea, ngemva kokudla kwegazi, kuye ngokuthi bangakanani abanceliswa i-urea (F(11, 360) = 4.69; p < 0.0001), abesifazane abanceliswa amazinga e-urea aphakathi kuka-134 µM no-1.34 mM babeke amaqanda amaningi (Isithombe 4E). Abesifazane abanceliswa amazinga e-urea angu-134 µM noma ngaphezulu babeke amaqanda amakhulu kunabesifazane abanceliswa ngamanzi (F(10, 4245) = 36.7; p < 0.0001; Isithombe 4F), kanye nosayizi wama-larval, yize bethintwe amazinga afanayo e-urea komama (F(10, 3305) = 37.9; p < 0.0001) kwakuhlukahluka kakhulu (Isithombe 4G).
Ukukhanga okuphelele kwe-headspace volatile extracts yomchamo wezinkomo ezifuna umninikhaya. I-arabiensis ehlolwe ku-olfactometer yengilazi (Isithombe 5A) yathinteka kakhulu ubudala bomchamo (χ2 = 15.9, df = 4, p = 0.0032; Isithombe 5B). Ukuhlaziywa kwangemva kwesikhathi kubonise ukuthi iphunga lomchamo elidala emahoreni angama-24 libangele amazinga aphezulu kakhulu okukhanga uma kuqhathaniswa nazo zonke ezinye izindlela zokwelapha (amahora angama-72: p = 0.0060, amahora ayi-168: p = 0.012, pentane: p = 0.00070), Ngaphandle kwephunga lomchamo omusha (p = 0.13; Isithombe 5B). Nakuba ukukhanga okuphelele komiyane abamunca igazi ephunga lomchamo kwakungafani kakhulu (χ2 = 8.78, df = 4, p = 0.067; Isithombe 5C), laba besifazane batholakale bekhanga kakhulu kwi-headspace volatile extracts uma kuqhathaniswa nomchamo onamahora angama-72 ubudala uma kuqhathaniswa kuya ezilawulini (p = 0.0066; Umfanekiso 5C).
Izimpendulo zokuziphatha ephunga lomchamo wenkomo wemvelo nowokwenziwa ekufuneni i-Anopheles arabianus ephethwe ngumnikazi kanye negazi. I-Schematic ye-olfactometer ye-glass tube (A). Ukukhangwa kwe-headspace volatile extracts kusuka kumchamo wenkomo omusha nomdala kuya komiyane abamukela igazi (B) kanye nabamunca igazi (C). Thola ukusabela kwe-tentacle ye-Lord An. I-Headspace extracts ehlukaniswe kumchamo wenkomo omusha (D), amahora angu-24 (E), amahora angu-72 (F), kanye namahora angu-168 (G) asebekhulile kuyaboniswa. Izimpawu zokutholwa kwe-electron antenna (EAD) zibonisa izinguquko ze-voltage ekuphenduleni kwamakhemikhali asebenzayo endaweni yekhanda akhishwe ku-chromatograph yegesi futhi atholwe yisitholi se-ionization selangabi (FID). Ibha yesikali imelela i-amplitude yokuphendula (mV) uma kuqhathaniswa nesikhathi sokugcina (s). Izakhiwo kanye namazinga okukhululwa (µg h-1) kwamakhemikhali asebenzayo ngokwezinto eziphilayo kuyaboniswa. I-asterisk eyodwa (*) ikhombisa impendulo ehambisanayo ye-amplitude ephansi. I-asterisks ephindwe kabili (**) ikhombisa izimpendulo ezingaphindeki. Thola i-host (H) kanye ne-blood-muncting (I) I-An.arabiensis inezikhalo ezahlukene ezingxubeni zokwenziwa zephunga lomchamo wenkomo omusha nomdala. Izilinganiso ezimaphakathi zomiyane ezidonswa ngamagama ezinhlamvu ezahlukene zazihluke kakhulu komunye nomunye (i-ANOVA yendlela eyodwa kusetshenziswa ukuhlaziywa kwe-post hoc kukaTukey; p < 0.05). Amaphutha amelela iphutha elijwayelekile lesikali
I-Ann.arabiensis yesifazane, emahoreni angu-72 kanye namahora angu-120 ngemva kokudla kwegazi, ngesikhathi sokuzala, akukho ukukhetha okubonisiwe kokukhishwa kwe-headspace volatile okuvela kumchamo wenkomo omusha nomdala uma kuqhathaniswa nokulawula kwe-pentane (χ2 = 3.07, p > 0.05; Ifayela elengeziwe 1: Umfanekiso S1).
Ku-Ann.arabiensis yesifazane, ukuhlaziywa kwe-GC-EAD kanye ne-GC-MS kuthole amakhemikhali ayisishiyagalombili, ayisithupha, amathathu kanye namathathu asebenzayo (Isithombe 5D-G). Nakuba umehluko enanini lamakhemikhali abangela izimpendulo ze-electrophysiological ubonwe, iningi lala makhemikhali lalikhona ku-headspace volatile extract ngayinye eqoqwe kumchamo omusha nomdala. Ngakho-ke, ku-extract ngayinye, amakhemikhali kuphela akhiqiza impendulo yomzimba evela kuma-antennae esifazane angaphezu komkhawulo afakiwe ekuhlaziyweni okwengeziwe.
Izinga lokukhishwa okuguquguqukayo kwama-compounds asebenzayo eqoqweni le-headspace likhuphuke lisuka ku-29 µg h-1 kumchamo omusha laya ku-242 µg h-1 kumchamo onamahora angu-168, ikakhulukazi ngenxa ye-p-cresol kanye ne-m-formaldehyde Phenol iyanda kanye ne-phenol. Ngokuphambene nalokho, amazinga okukhishwa kwamanye ama-compounds, njenge-2-cyclohexen-1-one kanye ne-decanal, ehla ngokukhula komchamo, okuhambisana nokwehla okubonakalayo kokuqina kwesignali (ubuningi) ku-chromatogram (Isithombe 5D)-G kwesobunxele) kanye nezimpendulo zomzimba kula ma-compounds (Isithombe 5D-G kwesobunxele).
Sekukonke, ingxube yokwenziwa yayinesilinganiso esifanayo semvelo samakhemikhali asebenzayo atholakala ezikhishweni eziguquguqukayo zezindawo zomchamo ezintsha nezindala (Isithombe 5D–G) futhi akubonakali sengathi idale ukukhanga okukhulu ekufuneni umninimzi (χ2 = 8.15, df = 4, p = 0.083; Isithombe 5H) noma omiyane abamunca igazi (χ2 = 4.91, df = 4, p = 0.30; Isithombe 5I). Kodwa-ke, ukuqhathaniswa kwangemva kwe-hoc phakathi kokwelashwa kubonise ukuthi omiyane abafuna umninimzi babekhanga kakhulu engxubeni yokwenziwa yomchamo ondala wamahora angama-24 uma kuqhathaniswa nezilawuli ze-pentane (p = 0.0086; Isithombe 5H).
Ukuze kuhlolwe indima yezingxenye ngazinye ezingxubeni zokwenziwa zomchamo onamahora angama-24, kuhlolwe izingxube eziyisithupha zokususa ezixutshwe ngokuqhathanisa nezingxube eziphelele ekuhlolweni kwe-Y-tube, lapho kususwe khona izingxube ngazinye. Komiyane abafuna umninimzi, ukukhipha izingxube ngazinye engxubeni ephelele kube nomthelela omkhulu ezimpendulweni zokuziphatha (χ2 = 19.63, df = 6, p = 0.0032; Ifayela elengeziwe 1: Umfanekiso S2A), zonke izingxube zokususa zazikhanga kakhulu kuneZincane kunezixutshwe ngokuphelele. Ngokuphambene nalokho, ukususwa kwezingxube ngazinye engxubeni yokwenziwa ngokugcwele akuzange kuthinte izimpendulo zokuziphatha zomiyane abamunca igazi (χ2 = 11.38, df = 6, p = 0.077), ngaphandle kwe-decanal, okwaholela emazingeni aphansi uma kuqhathaniswa nengxube ephelele. Ukukhanga (p = 0.022; Ifayela Elingeziwe 1: Umfanekiso S2B).
Edolobhaneni elihlala umalaleveva e-Ethiopia, ukusebenza kahle kwengxube yokwenziwa yomchamo wezinkomo wamahora angama-24 ekudonseni omiyane ngaphansi kwezimo zasensimini kwahlolwa ubusuku obuyishumi (Isithombe 6A). Kwabanjwa futhi kwatholakala omiyane abangu-4,861, abangu-45.7% kubo kwakuyi-Anthropus.gambiae sl, abangu-18.9% kwakuyi-Anopheles pharoensis kanti abangu-35.4% kwakuyi-Culex spp. (Ifayela elengeziwe 1: Ithebula S1). I-Anopheles arabinis iyilungu elilodwa kuphela le-An.Gambian species complex ehlonzwe ngokuhlaziywa kwe-PCR. Ngokwesilinganiso, omiyane abangu-320 babanjwa ngobusuku ngabunye, lapho izicupho ezinezicupho zokwenziwa zibamba omiyane abaningi kunezicupho ezibhangqiwe ngaphandle kwengxube (χ2(0, 3196) = 170.0, p < 0.0001). Izicupho ezingabhangqiwe zabekwa ubusuku ngabunye bokulawula obuhlanu ekuqaleni, maphakathi, nasekupheleni kokuhlolwa. Inani elifanayo lomiyane labanjwa kubhangqa ngalinye. kwezicupho, okubonisa ukungabi nabandlululo phakathi kwezindlu (χ2(0, 1665) = 9 × 10-13, p > 0.05) futhi akukho ukwehla kwenani labantu phakathi nesikhathi socwaningo. Uma kuqhathaniswa nezicupho zokulawula, inani lomiyane ababanjwe ezicuphweni eziqukethe ingxube yokwenziwa landa kakhulu: ukufuna umbungazi (χ2(0, 2107) = 138.7, p < 0.0001), ukuncela igazi kwakamuva (χ2(0, 650) = 32.2, p < 0.0001) kanye nokukhulelwa (χ2(0, 228) = 6.27, p = 0.0123; Ifayela elengeziwe 1: Ithebula S1). Lokhu kubonakala futhi enanini eliphelele lomiyane ababanjwe: ukufuna umbungazi > ukumunca igazi > ukhulelwe > ukhulelwe kancane > owesilisa.
Ukuhlolwa kwensimu kokusebenza kahle kwengxube yephunga lomchamo wenkomo eyenziwe ngamahora angama-24. Ukuhlolwa kwensimu kwenziwe eningizimu-maphakathi ne-Ethiopia (imephu), eduze kwedolobha laseMaki (faka), kusetshenziswa isicupho esikhanyayo se-Centers for Disease Control (CDC) (kwesokudla) ezindlini ezibhangqiwe, ngomklamo wesikwele waseLatin (isithombe se-aerial) (A). Ama-phototrap e-CDC afakwe iphunga adonsela futhi abambe ama-Anopheles arabesques (B) abesifazane, kodwa hhayi ama-Anopheles farroes (C), ngendlela ehlukile, umphumela oxhomeke esimweni somzimba. Ngaphezu kwalokho, lezi zicupho zibambe inani elikhulayo kakhulu lomiyane baseCulex. (D) Uma kuqhathaniswa nokulawula. Amabha angakwesobunxele amelela isilinganiso sokukhetha somiyane ababanjwe ngamabhangqa esicupho esinuka kamnandi (oluhlaza okotshani) kanye nezicupho zokulawula (ezivulekile) (N = 10), kuyilapho amabha angakwesokudla emelela isilinganiso sokukhetha ngamabhangqa ezicupho zokulawula (ezivulekile; N = 5). Ama-Asterisk abonisa amazinga okubaluleka kwezibalo (*p = 0.01 kanye ***p < 0.0001)
Lezi zinhlobo ezintathu zibanjwe ngendlela ehlukile ezicuphweni eziqukethe izingxube zokwenziwa. Ukufuna umsingathi (χ2(1, 1345) = 71.7, p < 0.0001), ukuphakelwa kwegazi (χ2(1, 517) = 16.7, p < 0.0001) kanye nokukhulelwa (χ2(1, 180) = 6.11, p = 0.0134) i-.arabiensis yayibanjwe ogibeni ikhipha ingxube yokwenziwa (Isithombe 6B), kuyilapho inani le-An lingahlukanga. I-Pharoensis ezimweni ezahlukene zomzimba yatholakala (Isithombe 6C). Ku-Culex, ukwanda okukhulu kuphela kwenani lomiyane abafuna abasingathi kwatholakala ezicuphweni ezifakwe ingxube yokwenziwa (χ2(1,1319) = 12.6, p = 0.0004; Isithombe 6D), uma kuqhathaniswa nezingibe zokulawula.
Izicupho zokudoba ezitholakala ngaphandle kwezindawo ezingaba khona phakathi kwezindawo zokuzalela kanye nemiphakathi yasemakhaya e-Ethiopia zasetshenziswa ukuhlola ukuthi omiyane be-malaria basebenzisa iphunga lomchamo wenkomo njengesibonakaliso sendawo yokuhlala yomninimzi. Uma kungekho zimpawu zomninimzi, ukushisa, kanye nokuba khona noma ngaphandle kwephunga lomchamo wenkomo, awekho omiyane ababanjwe (Ifayela elengeziwe 1: Umfanekiso S3). Kodwa-ke, lapho kukhona izinga lokushisa eliphezulu kanye nephunga lomchamo wenkomo, omiyane be-malaria besifazane badonswa futhi babanjwa, yize babembalwa, kungakhathalekile ubudala bomchamo (χ2(5, 25) = 2.29, p = 0.13; Ifayela elengeziwe 1: Umfanekiso S3). Ngokuphambene nalokho, izilawuli zamanzi azibambanga omiyane be-malaria emazingeni okushisa aphezulu (Ifayela elengeziwe 1: Umfanekiso S3).
Omiyane be-malaria bathola futhi basabalalise amakhemikhali aqukethe i-nitrogen ngokudla okuncishisiwe emchameni wenkomo (okungukuthi, ama-chiddle) ukuze bathuthukise izici zomlando wokuphila, ezifana nezinye izinambuzane [2, 4, 24, 25, 26]. Umchamo wenkomo uwumthombo ovuselelekayo otholakala kalula ohlotshaniswa eduze nezindawo zokuphumula zezilwane ezithwala i-malaria, njengezibaya zenkomo nezitshalo ezinde eduze kwamakhaya asemakhaya nezindawo zokuzalela. Omiyane besifazane bathola lo mthombo ngephunga futhi bayakwazi ukulawula ukumuncwa kwamakhemikhali e-nitrogen emchameni, okuhlanganisa i-urea, ingxenye enkulu ye-nitrogen emchameni [15, 16]. Kuye ngesimo somzimba somiyane wesifazane, izakhamzimba emchameni zabelwe ukuthuthukisa umsebenzi wokundiza nokusinda komiyane besifazane abafuna umninimzi, kanye nezici zokusinda nokuzala zabantu abanceliswa igazi ngesikhathi somjikelezo wokuqala we-gonadotropic. Ngakho-ke, ukuxuba umchamo kudlala indima ebalulekile yokudla okunempilo kwezilwane ezithwala i-malaria ezivalekile njengabantu abadala abangondlekile [8], njengoba kunikeza omiyane besifazane ikhono lokuthola amakhemikhali abalulekile e-nitrogen ngokuhlanganyela ekudleni okunengozi encane. Lokhu okutholakele kunemiphumela ebalulekile ye-epidemiological, njengoba abesifazane bandisa ubude besikhathi sokuphila, umsebenzi kanye nomphumela wokuzala, konke okuthinta amandla e-vector. Ngaphezu kwalokho, lokhu kuziphatha kungaba yisisulu sezinhlelo zokuphatha i-vector zesikhathi esizayo.