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Bakar soja tashi (Hermetia illucens, L. 1758) wani kwaro ne mai lalata da kuma babban yuwuwar yin amfani da samfuran kwayoyin halitta masu wadatar carbohydrate. Daga cikin carbohydrates, baƙar fata kwari sun dogara da sukari mai narkewa don haɓakawa da haɓakar lipid. Makasudin wannan binciken shine don kimanta tasirin sikari mai narkewa akan haɓakawa, rayuwa, da bayanin fatty acid na kudajen soja baƙar fata. Ƙara abincin kaza tare da monosaccharides da disaccharides daban. An yi amfani da Cellulose azaman sarrafawa. Larvae sun ciyar da glucose, fructose, sucrose, da maltose sun girma cikin sauri fiye da sarrafa tsutsa. Sabanin haka, lactose yana da tasirin hana abinci mai gina jiki a kan tsutsa, rage jinkirin girma da rage nauyin jikin mutum na ƙarshe. Koyaya, duk sukari mai narkewa ya sanya tsutsa mai kiba fiye da waɗanda aka ciyar da tsarin kulawa. Musamman ma, sukarin da aka gwada sun tsara bayanin martabar fatty acid. Maltose da sucrose sun ƙara yawan fatty acid abun ciki idan aka kwatanta da cellulose. Sabanin haka, lactose yana haɓaka haɓakar ƙwayoyin kitse na abinci marasa abinci. Wannan binciken shine na farko don nuna tasirin sukari mai narkewa akan abun da ke tattare da fatty acid na bakar fata tsutsa. Sakamakonmu yana nuna cewa ƙwayoyin carbohydrates da aka gwada suna da tasiri mai mahimmanci akan nau'in fatty acid na baƙar fata na sojan tsutsa don haka yana iya ƙayyade aikace-aikacen su na ƙarshe.
Bukatar makamashi da furotin dabba na duniya na ci gaba da karuwa1. Dangane da yanayin dumamar yanayi, ya zama wajibi a nemo hanyoyin da za a bi wajen samar da makamashin burbushin halittu da hanyoyin samar da abinci na gargajiya yayin da ake kara samar da kayayyaki. Kwari suna ba wa 'yan takara alkawarin magance waɗannan batutuwa saboda ƙarancin sinadarai da tasirin muhalli idan aka kwatanta da kiwon dabbobi na gargajiya2. Daga cikin kwari, kyakkyawan ɗan takara don magance waɗannan batutuwa shine jirgin soja na baƙar fata (BSF), Hermetia illucens (L. 1758), nau'in lalata da ke iya ciyar da nau'o'in nau'in kwayoyin halitta3. Don haka, haɓaka waɗannan abubuwan da ake buƙata ta hanyar kiwo na BSF na iya ƙirƙirar sabon tushen albarkatun ƙasa don biyan bukatun masana'antu daban-daban.
BSF larvae (BSFL) na iya ciyar da kayan aikin gona da masana'antu na masana'antu irin su hatsin masu shayarwa, ragowar kayan lambu, ɓangaren 'ya'yan itace da gurasar da ba a taɓa gani ba, waɗanda suka dace da ci gaban BSFL saboda babban carbohydrate (CH) 4,5, 6 abun ciki. Babban yawan samar da BSFL yana haifar da samuwar samfura guda biyu: feces, cakuɗen raƙuman ruwa da najasa waɗanda za a iya amfani da su azaman taki don shuka shuka7, da tsutsa, waɗanda galibi sun ƙunshi sunadarai, lipids da chitin. Sunadaran da lipids ana amfani da su ne a cikin noman dabbobi, biofuel da kayan shafawa8,9. Amma game da chitin, wannan biopolymer yana samun aikace-aikace a cikin sashin abinci na agri-abinci, fasahar kere-kere da kula da lafiya10.
BSF kwaro ce mai kama da wuta, ma'ana cewa metamorphosis da haifuwar sa, musamman matakan cinye makamashi na rayuwar kwari, ana iya tallafawa gaba ɗaya ta hanyar tanadin abinci mai gina jiki da aka samar yayin girma11. Musamman ma, furotin da haɗin lipid yana haifar da haɓakar jiki mai kitse, wani muhimmin sashin ajiya wanda ke fitar da kuzari yayin lokutan ciyarwa na BSF: prepupa (watau matakin tsutsa na ƙarshe lokacin da tsutsa BSF ta zama baki yayin ciyarwa da nema). don yanayin da ya dace da metamorphosis), pupae (watau matakin da ba ya motsi lokacin da kwarin ke fuskantar metamorphosis), da manya12,13. CH shine babban tushen makamashi a cikin abincin BSF14. Daga cikin waɗannan abubuwan gina jiki, CH fibrous kamar hemicellulose, cellulose da lignin, sabanin disaccharides da polysaccharides (kamar sitaci), ba za a iya narkar da su ta BSFL15,16 ba. Narkar da CH shine muhimmin mataki na farko don shayar da carbohydrates, wanda a ƙarshe aka sanya shi cikin ruwa mai sauƙi zuwa sukari mai sauƙi a cikin hanji16. Za'a iya shayar da sikari mai sauƙi (watau ta cikin membrane peritrophic na hanji) kuma a daidaita shi don samar da kuzari17. Kamar yadda aka ambata a sama, tsutsa suna adana makamashi mai yawa kamar lipids a cikin mai mai12,18. Lipids ɗin ajiya sun ƙunshi triglycerides (tsaka-tsaki na lipids waɗanda aka samo su daga ƙwayar glycerol guda ɗaya da fatty acid guda uku) waɗanda tsutsa suka haɗe daga sikari mai sauƙi na abinci. Waɗannan CH suna ba da abubuwan acetyl-CoA da ake buƙata don fatty acid (FA) biosynthesis ta hanyar fatty acid synthase da hanyoyin thioesterase19. Bayanan martabar fatty acid na H. illukens lipids yana da rinjaye ta halitta ta hanyar cikakken fatty acid (SFA) tare da babban adadin lauric acid (C12: 0) 19,20. Saboda haka, babban abun ciki na lipid da abun da ke tattare da kitsen acid suna saurin zama abubuwan iyakancewa don amfani da duk tsutsa a cikin abincin dabbobi, musamman a cikin kiwo inda ake buƙatar polyunsaturated fatty acid (PUFA)21.
Tun lokacin da aka gano yuwuwar BSFL don rage sharar kwayoyin halitta, nazarin kan ƙimar samfuran samfuran daban-daban sun nuna cewa abun da ke cikin BSFL an tsara shi ta hanyar abincin sa. A halin yanzu, ka'idar bayanin martabar FA na H. illucens yana ci gaba da ingantawa. An nuna ikon BSFL don bioaccumulate PUFA akan abubuwan PUFA masu wadata kamar algae, sharar kifi, ko abinci irin su flaxseed, wanda ke ba da ingantaccen bayanin martaba na FA don abinci mai gina jiki na dabba19,22,23. Sabanin haka, don samfuran da ba a wadatar da su a cikin PUFA ba, ba koyaushe ake samun alaƙa tsakanin bayanan bayanan abinci na FA da tsutsa FA ba, yana nuna tasirin sauran abubuwan gina jiki24,25. A zahiri, tasirin CH mai narkewa akan bayanan bayanan FA ya kasance mara fahimta kuma ba a bincika ba24,25,26,27.
A iyakar saninmu, ko da yake jimlar monosaccharides da disaccharides suna da yawa a cikin abincin H. illusens, aikinsu na gina jiki ya kasance da rashin fahimta a cikin H. illucens abinci mai gina jiki. Manufar wannan binciken shine don bayyana tasirin su akan abinci mai gina jiki na BSFL da abun da ke ciki na lipid. Za mu ƙididdige girma, tsira, da yawan aiki na tsutsa a ƙarƙashin yanayin abinci daban-daban. Sa'an nan, za mu kwatanta abun ciki na lipid da fatty acid profile na kowane abinci don haskaka tasirin CH akan ingancin abinci mai gina jiki na BSFL.
Mun yi hasashen cewa yanayin gwajin CH zai shafi (1) haɓakar tsutsa, (2) jimlar matakan lipid, da (3) daidaita bayanin martabar FA. Ana iya ɗaukar monosaccharides kai tsaye, yayin da disaccharides dole ne a shayar da ruwa. Monosaccharides don haka sun fi samuwa a matsayin tushen makamashi kai tsaye ko abubuwan da suka faru don lipogenesis ta hanyar FA synthase da hanyoyin thioesterase, don haka inganta haɓakar H. illucens tsutsa girma da kuma inganta tarawar ajiyar lipids (musamman lauric acid).
CH da aka gwada ya shafi matsakaicin nauyin jiki na tsutsa yayin girma (Fig. 1). FRU, GLU, SUC da MAL sun ƙaru tsutsa nauyin jiki daidai da abincin kulawa (CEL). Sabanin haka, LAC da GAL sun bayyana don jinkirta ci gaban tsutsa. Musamman ma, LAC yana da mummunar tasiri akan ci gaban tsutsa idan aka kwatanta da SUC a duk tsawon lokacin girma: 9.16 ± 1.10 MG tare da 15.00 ± 1.01 MG a ranar 3 (F6,21 = 12.77, p <0.001; Fig. 1), 125.11 ± 4.4. MG da 211.79 ± 14.93 MG, bi da bi, a ranar 17 (F6,21 = 38.57, p <0.001; Fig. 1).
Amfani da monosaccharides daban-daban (fructose (FRU), galactose (GAL), glucose (GLU), disaccharides (lactose (LAC), maltose (MAL), sucrose (SUC)) da cellulose (CEL) azaman sarrafawa. Girman larvae da ake ciyar da baƙar fata soja tsutsa. Kowace aya akan lanƙwasa tana wakiltar ma'anar nauyin mutum ɗaya (mg) da aka lissafta ta hanyar auna 20 zaɓaɓɓun tsutsa daga yawan 100 tsutsa (n = 4). Kuskuren sanduna suna wakiltar SD.
Abincin CEL ya ba da kyakkyawar rayuwa na tsutsa na 95.5 ± 3.8%. Bugu da ƙari, an rage rayuwar H. illucens ciyar da abinci mai ɗauke da CH mai narkewa (GLM: χ = 107.13, df = 21, p <0.001), wanda MAL da SUC (disaccharides) suka haifar a cikin binciken CH. Yawan mace-mace ya yi ƙasa da na GLU, FRU, GAL (monosaccharide), da LAC (EMM: p <0.001, Hoto 2).
Boxplot na rayuwa na baƙar fata soja larvae gardama bi da daban-daban monosaccharides (fructose, galactose, glucose), disaccharides (lactose, maltose, sucrose) da cellulose a matsayin controls. Jiyya tare da wasiƙar guda ba ta bambanta da juna ba (EMM, p> 0.05).
Duk abincin da aka gwada sun ba da damar tsutsa su isa matakin farko. Koyaya, CHs ɗin da aka gwada sun kasance suna haɓaka haɓakar tsutsa (F6,21=9.60, p<0.001; Table 1). Musamman ma, tsutsa da ke ciyar da GAL da LAC sun ɗauki tsawon lokaci don isa matakin farko idan aka kwatanta da tsutsa da aka haifa akan CEL (CEL-GAL: p<0.001; CEL-LAC: p<0.001; Table 1).
CH da aka gwada kuma yana da tasiri daban-daban akan nauyin jiki na tsutsa, tare da nauyin jiki na tsutsa da aka ciyar da abincin CEL ya kai 180.19 ± 11.35 mg (F6,21 = 16.86, p <0.001; Fig. 3). FRU, GLU, MAL da SUC sun haifar da matsakaicin matsakaicin nauyin tsutsa na ƙarshe fiye da 200 MG, wanda ya fi girma fiye da na CEL (p <0.05). Ya bambanta, tsutsa da ke ciyar da GAL da LAC suna da ƙananan nauyin jiki, matsakaicin 177.64 ± 4.23 MG da 156.30 ± 2.59 MG, bi da bi (p <0.05). Wannan tasirin ya fi bayyana tare da LAC, inda nauyin jiki na ƙarshe ya kasance ƙasa da tare da abinci mai sarrafawa (CEL-LAC: bambanci = 23.89 MG; p = 0.03; Hoto 3).
Ma'anar nauyin ƙarshe na kowane tsutsa wanda aka bayyana azaman tsutsa (mg) da ƙudaje na soja baƙar fata wanda aka bayyana azaman histogram (g) suna ciyar da monosaccharides daban-daban (fructose, galactose, glucose), disaccharides (lactose, maltose, sucrose) da cellulose (a matsayin iko). Haruffa ginshiƙi suna wakiltar ƙungiyoyi daban-daban a cikin jimlar nauyin tsutsa (p <0.001). Haruffa masu alaƙa da tsutsa suna wakiltar ƙungiyoyi masu ma'aunin ma'aunin tsutsa daban-daban (p <0.001). Kuskuren sanduna suna wakiltar SD.
Matsakaicin nauyin mutum ɗaya ya kasance mai zaman kansa daga matsakaicin matsakaicin jimlar ƙwayar tsutsa. A gaskiya ma, abincin da ke dauke da FRU, GLU, MAL, da SUC ba su kara yawan nauyin tsutsa da aka samar a cikin tanki ba idan aka kwatanta da CEL (Figure 3). Duk da haka, LAC ya rage mahimmancin jimlar nauyin (CEL-LAC: bambanci = 9.14 g; p <0.001; Hoto 3).
Shafin 1 yana nuna yawan amfanin ƙasa (lavae / day). Abin sha'awa, mafi kyawun amfanin CEL, MAL da SUC sun kasance iri ɗaya (Table 1). Sabanin haka, FRU, GAL, GLU da LAC sun rage yawan amfanin ƙasa idan aka kwatanta da CEL (Table 1). GAL da LAC sun yi mafi muni: an rage yawan amfanin ƙasa zuwa kawai 0.51 ± 0.09 g tsutsa / rana da 0.48 ± 0.06 g tsutsa / rana, bi da bi (Table 1).
Monosaccharides da disaccharides sun haɓaka abun ciki na lipid na tsutsa CF (Table 1). A kan abincin CLE, an samo tsutsa tare da abun ciki na lipid na 23.19 ± 0.70% na abun ciki na DM. Don kwatanta, matsakaicin abun ciki na lipid a cikin larvae da aka ciyar da sukari mai narkewa ya wuce 30% (Table 1). Koyaya, CH ɗin da aka gwada sun ƙara yawan kitsen su daidai gwargwado.
Kamar yadda aka zata, batutuwa na CG sun shafi bayanin martaba na FA na tsutsa zuwa digiri daban-daban (Fig. 4). Abubuwan da ke cikin SFA ya kasance mai girma a duk abincin abinci kuma ya kai fiye da 60%. MAL da SUC basu daidaita bayanin martabar FA ba, wanda ya haifar da karuwa a cikin SFA. Game da MAL, a gefe guda, wannan rashin daidaituwa ya haifar da raguwa a cikin abun ciki na fatty acids (MUFA) (F6,21 = 7.47; p <0.001; Fig. 4). A gefe guda kuma, ga SUC, raguwa ya kasance mafi daidaituwa tsakanin MUFA da PUFA. LAC da MAL suna da sabanin tasiri akan bakan FA (SFA: F6,21 = 8.74; p <0.001; MUFA: F6,21 = 7.47; p <0.001; PUFA: χ2 = 19.60; Df = 6; p <0.001; Figure) 4). Ƙananan rabo na SFA a cikin larvae masu ciyar da LAC ya bayyana yana ƙara abun ciki na MUFA. Musamman, matakan MUFA sun kasance mafi girma a cikin larvae masu ciyar da LAC idan aka kwatanta da sauran masu ciwon sukari banda GAL (F6,21 = 7.47; p <0.001; Hoto 4).
Amfani da monosaccharides daban-daban (fructose (FRU), galactose (GAL), glucose (GLU), disaccharides (lactose (LAC), maltose (MAL), sucrose (SUC)) da cellulose (CEL) azaman sarrafawa, makircin akwatin kitse. abun da ke ciki ciyar da baƙar fata soja tashi tsutsa. An bayyana sakamakon a matsayin kashi na jimlar FAME. Jiyya da aka yiwa alama da haruffa daban-daban sun bambanta sosai (p <0.001). (a) Matsakaicin cikakken fatty acid; (b) Mononsaturated fatty acids; (c) Polyunsaturated fatty acids.
Daga cikin fatty acids da aka gano, lauric acid (C12: 0) ya kasance mafi rinjaye a duk abubuwan da aka gani (fiye da 40%). Sauran SFAs na yanzu sune palmitic acid (C16: 0) (kasa da 10%), stearic acid (C18: 0) (kasa da 2.5%) da capric acid (C10: 0) (kasa da 1.5%). MUFAs yawanci suna wakiltar oleic acid (C18: 1n9) (kasa da 9.5%), yayin da PUFAs galibi sun ƙunshi linoleic acid (C18:2n6) (kasa da 13.0%) (duba Ƙarin Tebura S1). Bugu da ƙari, ƙananan ƙananan mahadi ba za a iya gano su ba, musamman ma a cikin nau'i na CEL larvae, inda lambar fili na 9 (UND9) ba a san shi ba ya kai matsakaicin 2.46 ± 0.52% (duba Ƙarin Ƙarin S1). Binciken GC×GC-FID ya nuna cewa yana iya zama fatty acid 20-carbon tare da ɗakuna biyar ko shida (duba Ƙarin Hoto S5).
Binciken PERMANOVA ya bayyana ƙungiyoyi daban-daban guda uku dangane da bayanan fatty acid (F6,21 = 7.79, p <0.001; Hoto 5). Babban nazarin abubuwan da ke ciki (PCA) na bakan TBC ya kwatanta wannan kuma an yi bayanin su ta hanyar sassa biyu (Hoto na 5). Babban abubuwan da aka gyara sun bayyana 57.9% na bambance-bambancen kuma sun haɗa, a cikin tsari mai mahimmanci, lauric acid (C12: 0), oleic acid (C18: 1n9), palmitic acid (C16: 0), stearic acid (C18: 0), da kuma linolenic acid (C18: 3n3) (duba hoto S4). Bangare na biyu ya bayyana 26.3% na bambance-bambancen kuma ya haɗa da, a cikin tsari mai mahimmanci, decanoic acid (C10: 0) da acid linoleic (C18: 2n6 cis) (duba Ƙarin Hoto S4). Bayanan bayanan abincin da ke ɗauke da sikari mai sauƙi (FRU, GAL da GLU) sun nuna halaye iri ɗaya. Sabanin haka, disaccharides sun samar da bayanan martaba daban-daban: MAL da SUC a daya hannun da LAC a daya bangaren. Musamman, MAL shine kawai sukari wanda ya canza bayanin martabar FA idan aka kwatanta da CEL. Bugu da kari, bayanin martabar MAL ya bambanta sosai da bayanan FRU da GLU. Musamman ma, bayanin martaba na MAL ya nuna mafi girman rabo na C12: 0 (54.59 ± 2.17%), yana yin daidai da CEL (43.10 ± 5.01%), LAC (43.35 ± 1.31%), FRU (48.90 ± 1.97%) da kuma GLU (48.38 ± 2.17%) bayanan martaba (duba Ƙarin Teburi S1). Hakanan MAL bakan ya nuna mafi ƙarancin C18: 1n9 abun ciki (9.52 ± 0.50%), wanda ya ƙara bambanta shi da LAC (12.86 ± 0.52%) da CEL (12.40 ± 1.31%). An lura da irin wannan yanayin don C16: 0. A cikin kashi na biyu, ƙirar LAC ta nuna mafi girman C18: 2n6 abun ciki (17.22 ± 0.46%), yayin da MAL ya nuna mafi ƙasƙanci (12.58 ± 0.67%). C18: 2n6 kuma ya bambanta LAC daga sarrafawa (CEL), wanda ya nuna ƙananan matakan (13.41 ± 2.48%) (duba Ƙarin Table S1).
PCA makirci na fatty acid profile na baƙar fata soja tashi larvae tare da daban-daban monosaccharides (fructose, galactose, glucose), disaccharides (lactose, maltose, sucrose) da cellulose a matsayin iko.
Don nazarin tasirin sinadirai masu narkewa akan H. illucens larvae, an maye gurbin cellulose (CEL) a cikin abincin kaza tare da glucose (GLU), fructose (FRU), galactose (GAL), maltose (MAL), sucrose (SUC), da kuma lactose (LAC). Duk da haka, monosaccharides da disaccharides suna da tasiri daban-daban akan ci gaba, rayuwa, da abun ciki na tsutsa HF. Misali, GLU, FRU, da nau'ikan disaccharide su (MAL da SUC) sun haifar da ingantaccen tasirin tallafi akan ci gaban tsutsa, yana basu damar cimma ma'aunin nauyi na ƙarshe fiye da CEL. Ba kamar CEL mara narkewa ba, GLU, FRU, da SUC na iya ƙetare shingen hanji kuma suyi aiki azaman mahimman tushen abinci mai gina jiki a cikin tsarin abinci16,28. MAL ba shi da takamaiman masu jigilar dabbobi kuma ana tsammanin za a sanya shi cikin ruwa zuwa kwayoyin glucose guda biyu kafin assimilation15. Ana adana waɗannan kwayoyin a cikin jikin kwari azaman tushen makamashi kai tsaye ko azaman lipids18. Na farko, game da na ƙarshe, wasu bambance-bambancen intramodal da aka lura na iya kasancewa saboda ƙananan bambance-bambance a cikin ƙimar jima'i. Lallai, a cikin H. illucens, haifuwa na iya zama kwata-kwata: manya-manyan mata a zahiri suna da isassun ajiyar kwai kuma sun fi maza nauyi29. Koyaya, tarin lipid a cikin BSFL yana daidaitawa tare da abinci mai narkewa na CH2, kamar yadda aka gani a baya don GLU da xylose26,30. Alal misali, Li et al.30 ya lura cewa lokacin da aka ƙara 8% GLU zuwa abincin tsutsa, abun ciki na lipid na BSF tsutsa ya karu da 7.78% idan aka kwatanta da sarrafawa. Sakamakonmu ya yi daidai da waɗannan abubuwan lura, yana nuna cewa kitsen da ke cikin tsutsa da ke ciyar da sukari mai narkewa ya fi na tsutsa da ke ciyar da abincin CEL, idan aka kwatanta da karuwar 8.57% tare da ƙarin GLU. Abin mamaki, an lura da irin wannan sakamakon a cikin tsutsa da aka ciyar da GAL da LAC, duk da mummunan tasiri akan ci gaban tsutsa, nauyin jiki na ƙarshe, da kuma rayuwa. Larvae da ke ciyar da LAC sun yi ƙasa da waɗanda aka ciyar da abincin CEL, amma abin da ke cikin su ya yi kama da tsutsa da ke ciyar da sauran sikari mai narkewa. Wadannan sakamakon suna nuna tasirin rashin abinci mai gina jiki na lactose akan BSFL. Na farko, abincin ya ƙunshi babban adadin CH. Tsarin sha da tsarin hydrolysis na monosaccharides da disaccharides, bi da bi, na iya kaiwa jikewa, haifar da kwalabe a cikin tsarin assimilation. Amma ga hydrolysis, ana aiwatar da shi ta α- da β-glucosidases 31. Wadannan enzymes sun fi son substrates dangane da girman su da kuma haɗin gwiwar sunadarai (α ko β linkages) tsakanin su monosaccharides 15. Hydrolysis na LAC zuwa GLU da GAL ana aiwatar da su ta hanyar β-galactosidase, wani enzyme wanda aka nuna ayyukansa a cikin gut na BSF 32. Duk da haka, maganganunsa na iya rashin isa idan aka kwatanta da adadin LAC da tsutsa ke cinyewa. Sabanin haka, α-glucosidase maltase da sucrase 15, waɗanda aka san ana bayyana su sosai a cikin kwari, suna iya rushe yawancin MAL da sucrose SUC, don haka suna iyakance wannan tasirin satiating. Abu na biyu, illar rashin abinci mai gina jiki na iya kasancewa saboda raguwar kuzarin aikin amylase na hanji na ƙwari da raguwar halayen ciyarwa idan aka kwatanta da sauran jiyya. Lallai, an gano sukari mai narkewa a matsayin masu haɓaka aikin enzyme mai mahimmanci don narkewar kwari, kamar amylase, kuma azaman abubuwan da ke haifar da amsawar ciyarwa33,34,35. Matsayin ƙarfafawa ya bambanta dangane da tsarin kwayoyin halitta na sukari. A zahiri, disaccharides suna buƙatar hydrolysis kafin sha kuma suna haɓaka amylase fiye da monosaccharides34. Sabanin haka, LAC yana da tasiri mai sauƙi kuma an gano cewa ba zai iya tallafawa ci gaban kwari a cikin nau'o'in nau'i daban-daban33,35. Misali, a cikin kwaro Spodoptera exigua (Boddie 1850), babu wani aikin hydrolytic na LAC da aka gano a cikin tsantsa na caterpillar midgut enzymes36.
Game da bakan FA, sakamakonmu yana nuna mahimman tasirin daidaitawa na CH da aka gwada. Musamman ma, kodayake lauric acid (C12: 0) ya ƙidaya ƙasa da 1% na jimlar FA a cikin abincin, ya mamaye duk bayanan martaba (duba Ƙarin Teburin S1). Wannan ya yi daidai da bayanan da suka gabata cewa lauric acid an haɗa shi daga abinci mai gina jiki CH a cikin H. illumins ta hanyar hanyar da ta ƙunshi acetyl-CoA carboxylase da FA synthase19,27,37. Sakamakonmu ya tabbatar da cewa CEL ba shi da narkewa kuma yana aiki a matsayin "wakili mai girma" a cikin abincin sarrafa BSF, kamar yadda aka tattauna a cikin binciken BSFL da yawa38,39,40. Maye gurbin CEL tare da monosaccharides da disaccharides ban da LAC ya haɓaka rabon C12: 0, yana nuna ƙarar CH ta hanyar tsutsa. Abin sha'awa shine, disaccharides MAL da SUC suna haɓaka haɓakar acid lauric da inganci fiye da monosaccharides ɗin su, suna ba da shawarar cewa duk da babban matakin polymerization na GLU da FRU, kuma tunda Drosophila shine kawai jigilar sucrose wanda aka gano a cikin nau'ikan furotin na dabba, masu jigilar kayayyaki na disaccharide. bazai kasance a cikin hanjin H. illucens larvae15 ba, yin amfani da GLU da FRU shine ya karu. Duk da haka, kodayake GLU da FRU suna da sauƙin daidaita su ta hanyar BSF, amma ana samun sauƙin daidaita su ta hanyar ƙwayoyin cuta da ƙwayoyin cuta na hanji, wanda zai iya haifar da saurin lalacewa da raguwar amfani da tsutsa idan aka kwatanta da disaccharides.
A kallo na farko, abun ciki na lipid na tsutsa da ke ciyar da LAC da MAL ya yi kama da juna, yana nuna kwatankwacin kasancewar waɗannan sikari. Koyaya, abin mamaki, bayanin martabar FA na LAC ya fi aukaka a cikin SFA, musamman tare da ƙananan abun ciki na C12: 0, idan aka kwatanta da MAL. Hasashe ɗaya don bayyana wannan bambance-bambancen shine LAC na iya haɓaka haɓakar ƙwayoyin abinci na FA ta hanyar acetyl-CoA FA synthase. Taimakawa wannan hasashe, LAC larvae yana da mafi ƙarancin decanoate (C10: 0) rabo (0.77 ± 0.13%) fiye da abincin CEL (1.27 ± 0.16%), yana nuna raguwar FA synthase da ayyukan thioesterase19. Na biyu, ana ɗaukar fatty acids na abinci a matsayin babban abin da ke tasiri tsarin SFA na H. illucens27. A cikin gwaje-gwajenmu, linoleic acid (C18: 2n6) ya ƙunshi 54.81% na fatty acids, tare da rabo a cikin LAC larvae shine 17.22 ± 0.46% kuma a cikin MAL 12.58 ± 0.67%. Oleic acid (cis + trans C18: 1n9) (23.22% a cikin abinci) ya nuna irin wannan yanayin. Rabon α-linolenic acid (C18: 3n3) kuma yana goyan bayan hasashe na bioaccumulation. Wannan fatty acid an san yana tarawa a cikin BSFL akan wadatar ƙasa, kamar ƙari na kek ɗin flaxseed, har zuwa 6-9% na jimlar fatty acid a cikin tsutsa19. A cikin wadataccen abinci, C18: 3n3 na iya lissafin har zuwa 35% na jimillar fatty acids. Koyaya, a cikin bincikenmu, C18: 3n3 ya lissafta 2.51% kawai na bayanan fatty acid. Kodayake adadin da aka samo a cikin yanayi ya kasance ƙasa a cikin tsutsanmu, wannan rabo ya fi girma a cikin LAC larvae (0.87 ± 0.02%) fiye da MAL (0.49 ± 0.04%) (p <0.001; duba Ƙarin Table S1). Abincin CEL yana da matsakaicin matsakaici na 0.72 ± 0.18%. A ƙarshe, rabon palmitic acid (C16: 0) a cikin tsutsa na CF yana nuna gudummawar hanyoyin roba da abinci na FA19. Hoton et al. 19 ya lura cewa an rage haɗin C16: 0 lokacin da aka wadatar da abinci tare da abinci na flaxseed, wanda aka danganta da raguwar samar da ƙwayar acetyl-CoA saboda raguwa a cikin rabon CH. Abin mamaki, ko da yake duka abinci suna da irin wannan abun ciki na CH kuma MAL sun nuna mafi girma bioavailability, MAL larvae ya nuna mafi ƙasƙanci C16: 0 rabo (10.46 ± 0.77%), yayin da LAC ya nuna mafi girma rabo, lissafin 12.85 ± 0.27% (p <0.05; duba). Ƙarin Tebura S1). Wadannan sakamakon suna nuna tasirin tasirin abubuwan gina jiki akan BSFL narkewa da metabolism. A halin yanzu, bincike kan wannan batu ya fi dacewa a cikin Lepidoptera fiye da Diptera. A cikin caterpillars, an gano LAC a matsayin mai rauni mai motsa halayen ciyarwa idan aka kwatanta da sauran masu narkewa kamar su SUC da FRU34,35. Musamman, a cikin Spodopteralittoralis (Boisduval 1833), amfani da MAL ya motsa ayyukan amylolytic a cikin hanji zuwa mafi girma fiye da LAC34. Irin wannan tasirin a cikin BSFL na iya yin bayanin haɓakar haɓakar hanyar C12: 0 synthetic a cikin MAL larvae, wanda ke da alaƙa da haɓakar CH na hanji, ciyar da tsawan lokaci, da aikin amylase na hanji. Karancin kuzarin kari na ciyarwa a gaban LAC na iya yin bayanin raguwar ci gaban LAC. Bugu da ƙari, Liu Yanxia et al. 27 ya lura cewa rayuwar rayuwar lipids a cikin H. illukens substrates ya fi tsayi fiye da na CH. Saboda haka, larvae na LAC na iya dogaro da yawa akan lipids na abinci don kammala ci gaban su, wanda zai iya ƙara abun ciki na lipid ɗin su na ƙarshe kuma ya canza bayanin martabar fatty acid ɗin su.
Ga iyakar iliminmu, ƙananan binciken ne kawai suka gwada tasirin monosaccharide da disaccharide ƙari ga abincin BSF akan bayanan martaba na FA. Na farko, Li et al. 30 ya kimanta tasirin GLU da xylose kuma an lura da matakan lipid kamar namu a ƙimar kari na 8%. Ba a cika bayanin bayanan FA ba kuma ya ƙunshi galibi na SFA, amma ba a sami bambance-bambance tsakanin sukarin biyu ba ko lokacin da aka gabatar da su lokaci guda30. Har ila yau, Cohn et al. 41 bai nuna wani tasiri na 20% GLU, SUC, FRU da GAL ƙari ga ciyarwar kaza akan bayanan bayanan FA daban-daban. An samo waɗannan bakan daga fasaha maimakon kwafin halittu, wanda, kamar yadda mawallafa suka bayyana, na iya iyakance ƙididdigar ƙididdiga. Bugu da ƙari, rashin sarrafa iso-sugar (ta amfani da CEL) yana iyakance fassarar sakamakon. Kwanan nan, binciken biyu na Nugroho RA et al. An nuna rashin daidaituwa a cikin FA spectra42,43. A cikin binciken farko, Nugroho RA et al. 43 sun gwada tasirin ƙara FRU zuwa abincin kwayan dabino. Bayanan FA na sakamakon larvae ya nuna matakan PUFA da yawa, fiye da 90% wanda aka samo daga abincin da ke dauke da 10% FRU (kama da bincikenmu). Kodayake wannan abincin ya ƙunshi pellet ɗin kifin PUFA mai wadata, ƙimar bayanin martabar FA da aka ruwaito na tsutsa akan tsarin sarrafa abinci wanda ya ƙunshi 100% PCM mai ƙima bai dace da duk bayanan da aka ruwaito a baya ba, musamman ma matakin mara kyau na C18: 3n3 na 17.77 ± 1.67% da 26.08 ± 0.20% don conjugated linoleic acid (C18: 2n6t), mai ƙarancin isomer na linoleic acid. Nazarin na biyu ya nuna irin wannan sakamako da suka haɗa da FRU, GLU, MAL da SUC42 a cikin abincin ƙwaya na dabino. Waɗannan karatun, kamar namu, suna nuna matsaloli masu tsanani wajen kwatanta sakamako daga gwajin abinci na tsutsa na BSF, kamar zaɓin sarrafawa, hulɗa tare da sauran hanyoyin gina jiki, da hanyoyin bincike na FA.
A lokacin gwaje-gwajen, mun lura cewa launi da warin substrate sun bambanta dangane da abincin da aka yi amfani da su. Wannan yana nuna cewa ƙananan ƙwayoyin cuta na iya taka rawa a cikin sakamakon da aka lura a cikin substrate da tsarin narkewa na tsutsa. A gaskiya ma, monosaccharides da disaccharides suna samun sauƙin narkewa ta hanyar mallakan ƙwayoyin cuta. Saurin amfani da sikari mai narkewa ta hanyar ƙwayoyin cuta na iya haifar da sakin adadi mai yawa na samfuran rayuwa na ƙwayoyin cuta kamar ethanol, lactic acid, fatty acid gajere (misali acetic acid, propionic acid, butyric acid) da carbon dioxide44. Wasu daga cikin waɗannan mahadi na iya kasancewa da alhakin tasirin guba mai guba akan tsutsa wanda Cohn et al.41 ya lura a ƙarƙashin yanayin haɓaka irin wannan. Misali, ethanol yana da illa ga kwari45. Yawan iskar iskar carbon dioxide na iya haifar da taruwar ta a kasan tankin, wanda zai iya hana iskar iskar oxygen idan yanayin iska bai bari a sake shi ba. Game da SCFAs, tasirin su akan kwari, musamman H. illucens, sun kasance marasa fahimta, kodayake lactic acid, propionic acid, da butyric acid an nuna su zama masu mutuwa a cikin Callosobruchus maculatus (Fabricius 1775)46. A cikin Drosophila melanogaster Meigen 1830, waɗannan SCFAs alamun ƙamshi ne waɗanda ke jagorantar mata zuwa wuraren oviposition, suna ba da shawara mai fa'ida ga ci gaban tsutsa47. Koyaya, acetic acid an rarraba shi azaman abu mai haɗari kuma yana iya hana haɓakar tsutsa sosai. Sabanin haka, kwanan nan an gano lactate na microbially yana da tasirin kariya daga ƙwayoyin cuta masu haɗari a cikin Drosophila48. Bugu da ƙari, ƙananan ƙwayoyin cuta a cikin tsarin narkewa suna taka rawa a cikin narkewar CH a cikin kwari49. Abubuwan da ke tattare da ilimin lissafi na SCFAs akan microbiota na gut, irin su yawan ciyarwa da maganganun kwayoyin halitta, an bayyana su a cikin 50 na kashin baya. Hakanan suna iya samun tasirin trophic akan H. illucens larvae kuma suna iya ba da gudummawa a wani ɓangare ga tsarin bayanan bayanan FA. Nazarin kan illolin sinadirai na waɗannan samfuran haifuwa na ƙananan ƙwayoyin cuta za su fayyace tasirin su akan H. yana haskaka abinci mai gina jiki kuma ya ba da tushe don nazarin nan gaba akan ƙwayoyin cuta masu fa'ida ko masu lahani dangane da haɓaka su da ƙimar fa'idodin FA. Dangane da haka, ana ƙara yin nazari kan rawar da ƙwayoyin cuta ke takawa a cikin hanyoyin narkewar ƙwayoyin ƙwayoyin cuta. An fara kallon kwari a matsayin bioreactors, samar da pH da yanayin oxygenation wanda ke taimakawa ci gaban ƙwayoyin cuta na musamman a cikin lalata ko lalata kayan abinci mai gina jiki wanda ke da wuya ga kwari su narke 51. Kwanan nan, Xiang et al.52 ya nuna cewa, alal misali, ƙaddamar da sharar gida tare da cakuda kwayan cuta yana ba CF damar jawo hankalin kwayoyin da suka ƙware a cikin lalata lignocellulose, inganta lalacewarsa a cikin substrate idan aka kwatanta da substrates ba tare da tsutsa ba.
A ƙarshe, game da amfani da sharar gida ta hanyar H. illucens, abincin CEL da SUC sun samar da mafi yawan adadin tsutsa a kowace rana. Wannan yana nufin cewa duk da ƙananan nauyin ƙarshe na ɗaiɗaikun daidaikun mutane, jimillar nauyin tsutsa da aka samar akan abin da ke kunshe da CH mara narkewa yana kama da wanda aka samu akan abincin homosaccharide mai ɗauke da monosaccharides da disaccharides. A cikin bincikenmu, yana da mahimmanci a lura cewa matakan sauran abubuwan gina jiki sun isa don tallafawa haɓakar yawan tsutsa kuma ya kamata a iyakance ƙarin CEL. Duk da haka, tsarin ƙarshe na tsutsa ya bambanta, yana nuna muhimmancin zabar dabarun da ya dace don ƙarfafa kwari. CEL larvae da ake ciyar da abinci gabaɗaya sun fi dacewa don amfani da su azaman abincin dabbobi saboda ƙananan abun ciki da ƙananan matakan lauric acid, yayin da tsutsa da ake ciyar da su tare da abincin SUC ko MAL suna buƙatar lalata ta hanyar danna don ƙara darajar mai, musamman a cikin biofuel. sashen. Ana samun LAC a cikin samfuran kiwo irin su whey daga samar da cuku. Kwanan nan, amfani da shi (3.5% lactose) ya inganta nauyin jikin tsutsa na ƙarshe53. Koyaya, tsarin sarrafa abinci a cikin wannan binciken ya ƙunshi rabin abun ciki na lipid. Sabili da haka, tasirin rage abinci mai gina jiki na LAC ƙila an magance shi ta hanyar tsutsa bioaccumulation na lipids na abinci.
Kamar yadda aka nuna ta hanyar binciken da ya gabata, kaddarorin monosaccharides da disaccharides suna tasiri sosai ga ci gaban BSFL kuma suna daidaita bayanin martabar FA. Musamman ma, LAC da alama tana taka rawar hana abinci mai gina jiki yayin haɓakar tsutsa ta hanyar iyakance wadatar CH don shayarwar lipid na abinci, ta haka yana haɓaka haɓakar UFA bioaccumulation. A cikin wannan mahallin, zai zama mai ban sha'awa don gudanar da bioassays ta amfani da abincin da ke haɗa PUFA da LAC. Bugu da ƙari, rawar ƙananan ƙwayoyin cuta, musamman rawar ƙananan ƙwayoyin cuta (irin su SCFAs) waɗanda aka samo daga hanyoyin haifuwa na sukari, ya kasance batun bincike wanda ya cancanci bincike.
An samo kwari daga yankin BSF na Laboratory of Functional and Evolutionary Entomology da aka kafa a 2017 a Agro-Bio Tech, Gembloux, Belgium (don ƙarin cikakkun bayanai game da hanyoyin kiwon lafiya, duba Hoc et al. 19). Don gwaje-gwajen gwaji, 2.0 g na qwai BSF an tattara bazuwar yau da kullun daga kejin kiwo kuma an sanya su cikin kilogiram 2.0 na 70% rigar abincin kaza (Aveve, Leuven, Belgium). Kwanaki biyar bayan ƙyanƙyashe, an raba tsutsa daga cikin ƙasa kuma an ƙidaya su da hannu don dalilai na gwaji. An auna nauyin farko na kowane rukuni. Matsakaicin nauyin kowane mutum shine 7.125 ± 0.41 MG, kuma ana nuna matsakaicin kowane jiyya a cikin Ƙarin Tebura S2.
An daidaita tsarin abincin abinci daga binciken da Barragan-Fonseca et al. 38 . A taƙaice, an sami daidaituwa tsakanin ingancin abinci iri ɗaya don kajin tsutsa, irin abubuwan busassun busassun (DM) abun ciki, babban CH (10% dangane da sabon abinci) da rubutu, tun da sauƙi sugars da disaccharides ba su da kaddarorin rubutu. Dangane da bayanan masana'anta (Ciyarwar Chicken, AVEVE, Leuven, Belgium), CH da aka gwada (watau mai narkewa sugar) an ƙara shi daban azaman maganin ruwa mai narkewa (15.9%) zuwa abinci wanda ya ƙunshi furotin 16.0%, 5.0% jimlar lipids, 11.9% abincin kajin ƙasa wanda ya ƙunshi ash da 4.8% fiber. A cikin kowane kwalban 750 ml (17.20 × 11.50 × 6.00 cm, AVA, Tempsee, Belgium), 101.9 g na autoclaved CH an haɗe shi da 37.8 g na abincin kaza. Ga kowane abinci, abubuwan busassun busassun sun kasance 37.0%, gami da furotin na kama (11.7%), lipids masu kama da juna (3.7%) da sukari iri ɗaya (26.9% na ƙara CH). CH da aka gwada sune glucose (GLU), fructose (FRU), galactose (GAL), maltose (MAL), sucrose (SUC) da lactose (LAC). Abincin sarrafawa ya ƙunshi cellulose (CEL), wanda aka yi la'akari da rashin narkewa ga H. illucens larvae 38. An sanya tsutsa mai kwanaki dari 5 a cikin tire da aka sanya murfi mai ramin diamita 1 cm a tsakiya kuma an rufe shi da gidan sauro na filastik. An maimaita kowane abinci sau hudu.
An auna ma'aunin larval kwanaki uku bayan fara gwajin. Ga kowane ma'auni, an cire larvae 20 daga ma'auni ta amfani da ruwa mai dumi da karfi, bushe, da auna (STX223, Ohaus Scout, Parsippany, Amurka). Bayan an auna, an mayar da tsutsa zuwa tsakiyar substrate. Ana ɗaukar ma'auni akai-akai sau uku a mako har sai prepupa na farko ya bayyana. A wannan gaba, tattara, ƙidaya, kuma auna duk tsutsa kamar yadda aka bayyana a baya. Rarrabe mataki 6 tsutsa (watau farar tsutsa daidai da matakin tsutsa da ke gaban matakin prepupal) da prepupae (watau matakin tsutsa na ƙarshe lokacin da tsutsa BSF ta zama baki, dakatar da ciyarwa, da neman yanayin da ya dace da metamorphosis) kuma adana a - - 18 ° C don nazarin abun ciki. An ƙididdige yawan amfanin ƙasa a matsayin rabon jimlar yawan kwari (larvae da prepupae na mataki na 6) da aka samu kowace tasa (g) zuwa lokacin haɓaka (d). Duk ma'anar ma'ana a cikin rubutu an bayyana su azaman: ma'anar ± SD.
Duk matakan da suka biyo baya ta hanyar amfani da abubuwan kaushi (hexane (Hex), chloroform (CHCl3), methanol (MeOH)) an yi su a ƙarƙashin murfin hayaƙi kuma ana buƙatar sanye da safofin hannu na nitrile, aprons da gilashin aminci.
An bushe fararen tsutsa a cikin injin daskarewa na FreeZone6 (Labconco Corp., Kansas City, MO, Amurka) na tsawon sa'o'i 72 sannan a kasa (IKA A10, Staufen, Jamus). An fitar da jimlar lipids daga ± 1 g na foda ta amfani da hanyar Folch 54. An ƙayyade ragowar danshi na kowane samfurin lyophilized a cikin kwafi ta yin amfani da mai nazarin danshi (MA 150, Sartorius, Göttiggen, Jamus) don gyara jimlar lipids.
An canza jimlar lipids a ƙarƙashin yanayin acidic don samun methyl esters mai kitse. A taƙaice, kusan 10 mg lipids/100 µl CHCl3 maganin (100 µl) an kwashe shi da nitrogen a cikin bututun Pyrex© 8 ml (SciLabware – DWK Life Sciences, London, UK). An sanya bututun a cikin Hex (0.5 ml) (PESTINORM®SUPRATRACE n-Hexane> 95% don nazarin binciken kwayoyin halitta, VWR Chemicals, Radnor, PA, Amurka) da Hex/MeOH/BF3 (20/25/55) bayani (0.5) ml) a cikin wanka na ruwa a 70 ° C na 90 min. Bayan sanyaya, an ƙara 10% mai ruwa H2SO4 bayani (0.2 ml) da cikakken NaCl bayani (0.5 ml). Mix da bututu kuma cika cakuda tare da Hex mai tsabta (8.0 ml). An canza wani yanki na babban lokaci zuwa vial kuma an bincika ta hanyar chromatography gas tare da mai gano ionization na harshen wuta (GC-FID). An yi nazarin samfurori ta amfani da Trace GC Ultra (Thermo Scientific, Waltham, MA, Amurka) sanye take da injector mai tsaga / rarrabuwa (240 ° C) a cikin yanayin tsaga (raguwa: 10 mL / min), shafi na Stabilwax®-DA ( 30 m, 0.25 mm id, 0.25 μm, Restek Corp., Bellefonte, PA, Amurka) da FID (250) °C). An saita tsarin zafin jiki kamar haka: 50 ° C don 1 min, yana ƙaruwa zuwa 150 ° C a 30 ° C / min, yana ƙaruwa zuwa 240 ° C a 4 ° C / min kuma ya ci gaba a 240 ° C na 5 min. An yi amfani da Hex azaman fanko da ma'aunin tunani mai ɗauke da 37 fatty acid methyl esters (Supelco 37-bangaren FAMEmix, Sigma-Aldrich, Overijse, Belgium) an yi amfani dashi don ganewa. An tabbatar da gano acid fatty acid (UFAs) ta cikakkiyar GC mai girma biyu (GC × GC-FID) kuma kasancewar isomers an ƙaddara daidai ta hanyar ɗan daidaitawa na hanyar Ferrara et al. 55. Ana iya samun cikakkun bayanai na kayan aiki a cikin Ƙarin Tebura S3 da sakamakon a Ƙarin Hoto S5.
An gabatar da bayanan a cikin tsarin maƙunsar bayanai na Excel (Microsoft Corporation, Redmond, WA, Amurka). An yi nazarin ƙididdiga ta amfani da R Studio (version 2023.12.1+402, Boston, USA) 56. Bayanai akan nauyin tsutsa, lokacin haɓakawa da yawan aiki an kiyasta ta amfani da samfurin layi (LM) (umurnin "lm", R kunshin "stats" 56) kamar yadda suka dace da rarraba Gaussian. An ƙididdige ƙimar rayuwa ta amfani da ƙididdigar ƙirar ƙirar binomial ta hanyar amfani da ƙirar layin gabaɗaya (GLM) (umarni "glm", kunshin R "lme4" 57). An tabbatar da al'ada da homoscedasticity ta amfani da gwajin Shapiro (umarni "shapiro.test", R kunshin "stats" 56) da kuma nazarin bambance-bambancen bayanai (umurnin betadisper, R kunshin "vegan" 58). Bayan nazarin nau'i-nau'i na mahimmanci p-darajar (p <0.05) daga gwajin LM ko GLM, an gano bambance-bambance tsakanin kungiyoyi ta amfani da gwajin EMM (umurnin "emmeans", R kunshin "emmeans" 59).
An kwatanta cikakkun sifofin FA ta amfani da nazarin bambance-bambance daban-daban na bambance-bambance (watau permMANOVA; umarni "adonis2", R kunshin "vegan" 58) ta amfani da matrix nesa na Euclidean da 999 permutations. Wannan yana taimakawa wajen gano fatty acids waɗanda ke tasiri ga yanayin carbohydrates na abinci. An ƙara yin nazarin bambance-bambance masu mahimmanci a cikin bayanan FA ta amfani da kwatance biyu. Bayan haka an hango bayanan ta hanyar amfani da mahimman abubuwan bincike (PCA) (umarni "PCA", kunshin R "FactoMineR" 60). FA da ke da alhakin waɗannan bambance-bambance an gano su ta hanyar fassara da'irar alaƙa. An tabbatar da waɗannan 'yan takarar ta hanyar yin amfani da bincike na hanya ɗaya na bambance-bambance (ANOVA) (umarni "aov", R kunshin "stats" 56) sannan Tukey's post hoc test (umurnin TukeyHSD, R kunshin "stats" 56). Kafin bincike, an yi la'akari da al'ada ta amfani da gwajin Shapiro-Wilk, an duba homoscedasticity ta amfani da gwajin Bartlett (umurnin "bartlett.test", R kunshin "stats" 56), kuma an yi amfani da hanyar da ba ta dace ba idan ba a hadu da tunanin biyu ba. . An kwatanta bincike (umurni "kruskal.test", R kunshin "stats" 56), sa'an nan kuma Dunn ta post hoc gwaje-gwaje da aka yi amfani da (umurni dunn.test, R kunshin "dunn.test" 56).
An duba sigar ƙarshe ta rubutun ta amfani da Editan Grammarly azaman mai karantawa na Ingilishi (Grammarly Inc., San Francisco, California, Amurka) 61 .
Rubutun bayanan da aka ƙirƙira kuma aka bincika yayin binciken na yanzu suna samuwa daga mawallafin da ya dace bisa buƙatun da ya dace.
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Lokacin aikawa: Dec-25-2024