Pine nematode ni karantine yimuka endoparasite izwiho guteza igihombo gikomeye mubukungu bwibinyabuzima byamashyamba. Ubu bushakashatsi busubiramo ibikorwa bya nematiside ya halogenated indoles irwanya pine nematode hamwe nuburyo bwo gukora. Ibikorwa bya nematiside ya 5-iodoindole na avermectine (kugenzura neza) kurwanya pine nematode byari bisa kandi biri hejuru cyane (10 μg / mL). 5-iodoindole yagabanije fecundity, ibikorwa byimyororokere, impfu zinda na lisiti, hamwe nimyitwarire ya moteri. Imikoreshereze ya molekulari ya ligande hamwe na glutamate-iterwa na chloride ya chloride itagira ubuzima bushyigikira igitekerezo kivuga ko 5-iodoindole, kimwe na avermectine, ihuza cyane na site ikora neza. 5-Iodoindole yateje kandi fenotipiki itandukanye muri nematode, harimo gusenyuka kw'ingingo zidasanzwe / kugabanuka no kongera vacuolisation. Ibisubizo byerekana ko vacuole ishobora kugira uruhare mu rupfu rwa nematode methylation. Icy'ingenzi, 5-iodoindole ntabwo yari uburozi bwubwoko bwibimera byombi (cabage na radish). Kubwibyo, ubu bushakashatsi bwerekana ko ikoreshwa rya iyode mu bihe by’ibidukikije rishobora kugenzura ibikomere bya pinusi.
Ibiti bya pinusi nematode (Bursaphelenchus xylophilus) ni iy'ibiti bya pinusi nematode (PWN), nematode yimuka ya endoparasitike izwiho kwangiza ibidukikije byangiza ibidukikije by’amashyamba1. Indwara ya pine wilt (PWD) iterwa nigiti cya pinusi nematode iragenda iba ikibazo gikomeye kumigabane myinshi, harimo Aziya n'Uburayi, no muri Amerika ya ruguru, nematode isenya amoko y'ibiti bya pinusi 1,2. Kugabanuka kw'ibiti bya pinusi nikibazo gikomeye cyubukungu, kandi ibyiringiro byo gukwirakwira kwisi yose biteye impungenge3. Ubwoko bw'inanasi bukurikira bwibasirwa cyane na nematode: Pinus densiflora, Pinus sylvestris, Pinus thunbergii, Pinus koraiensis, Pinus thunbergii, Pinus thunbergii, na Pinus radiata4. Pine nematode nindwara ikomeye ishobora kwica ibiti byinanasi mugihe cyibyumweru cyangwa ukwezi kwandura. Byongeye kandi, indwara ya pine nematode ikunze kugaragara mubinyabuzima bitandukanye, bityo hashyizweho urunigi rwanduye1.
Bursaphelenchus xylophilus ni karantine igihingwa-parasitike nematode ya Aphelenchoidea na claf 102.5. Nematode igaburira ibihumyo kandi ikororoka mubice byimbaho byibiti byinanasi, bikura mubice bine bitandukanye: L1, L2, L3, L4 numuntu mukuru1,6. Mugihe cyibura ryibiryo, pinusi nematode inyura mubyiciro byihariye - dauer, yangiza virusi yayo - inyenzi ya pinusi (Monochamus alternatus) ikoherezwa mubiti byinanasi bizima. Mubakira neza, nematode yimuka vuba binyuze mumyanya yibihingwa kandi igaburira ingirabuzimafatizo, ibyo bigatuma habaho reaction nyinshi ziterwa na hyperensitivite, kwinanura no gupfa mugihe cyumwaka umwe nyuma yo kwandura 1,7,8.
Kurwanya ibinyabuzima bya pinusi nematode kuva kera byabaye ingorabahizi, hamwe n’ingamba zo gushyira mu kato guhera mu kinyejana cya 20. Ingamba zigezweho zo kurwanya pine nematode zirimo cyane cyane kuvura imiti, harimo gutera ibiti no gushyira nematiside mu biti by'ibiti. Nematicide ikoreshwa cyane ni avermectin na avermectin benzoate, ikomoka mumuryango wa avermectin. Iyi miti ihenze cyane irwanya amoko menshi ya nematode kandi ifatwa nkibidukikije9. Nyamara, gukoresha inshuro nyinshi iyi nematiside biteganijwe ko bizatera umuvuduko wo gutoranya bizatuma rwose havuka indwara ya pine nematode irwanya, nkuko byagaragaye ku byonnyi byinshi by’udukoko, nka Leptinotarsa Decemlineata, Plutella xylostella na nematode Trichostrongylus colubriformis na Ostertagia Circincta, bigenda byiyongera buhoro buhoro10,12. Niyo mpamvu, uburyo bwo kurwanya bugomba kwigwa buri gihe kandi nematiside ikagenzurwa ubudahwema gushakisha ubundi buryo, buhendutse kandi bwangiza ibidukikije bwo kurwanya PVD. Mu myaka ya vuba aha, abanditsi benshi basabye ko hakoreshwa ibimera bivamo ibimera, amavuta yingenzi hamwe n’ibihindagurika nkibikoresho byo kugenzura nematode13,14,15,16.
Duherutse kwerekana ibikorwa bya nematicidal ya indole, intercellular intercelldom and interkingdom yerekana molekile, muri Caenorhabditis elegans 17. Indole ni ikimenyetso gikwirakwira cyane muri ecologiya ya mikorobe, igenzura imikorere myinshi igira ingaruka kuri physiologiya ya mikorobe, imiterere ya spore, ituze rya plasmide, kurwanya ibiyobyabwenge, ibinyabuzima, na virusi 18, 19. Igikorwa cya indole n'ibiyikomokaho birwanya izindi nematode zitera indwara ntabwo byigeze byigwa. Muri ubu bushakashatsi, twasesenguye ibikorwa bya nematicidal ya indoles 34 zirwanya pine nematode kandi tunasobanura uburyo bwo gukora bwa 5-iodoindole ikomeye cyane dukoresheje microscopi, gufotora igihe, hamwe nubushakashatsi bwa docking molekile, tunasuzuma ingaruka z’ubumara ku bimera dukoresheje uburyo bwo kumera imbuto.
Ubushuhe bwinshi (> 1.0 mM) bwa indole byavuzwe mbere ko bigira ingaruka mbi kuri nematode17. Nyuma yo kuvurwa B. xylophilus (ibyiciro byubuzima bivanze) hamwe na indole cyangwa 33 zikomoka kuri indole kuri mM 1, impfu za B. xylophilus zapimwe no kubara nematode nzima kandi yapfuye mugenzuzi no kuvura amatsinda. Indoles eshanu zagaragaje ibikorwa byingenzi byo kuvura indwara; kubaho kw'itsinda rishinzwe kutavurwa byari 95 ± 7% nyuma ya 24 h. Muri indoles 34 zapimwe, 5-iodoindole na 4-fluoroindole kuri mM 1 byateje impfu 100%, mu gihe 5,6-difluoroindigo, methylindole-7-carboxylate, na 7-iodoindole byateje impfu zigera kuri 50% (Imbonerahamwe 1).
Ingaruka ya 5-iodoindole kumiterere ya vacuole na metabolism yibiti bya pinusi nematode. . Imyambi itukura yerekana kubyimba vacuole, imyambi yubururu yerekana guhuza vacuole naho imyambi yumukara yerekana vacuole nini. Umwanya munini = 50 mm.
Byongeye kandi, ubu bushakashatsi bwanasobanuye inzira ikurikiranye y’urupfu rwa metani muri nematode (Ishusho 4C). Urupfu rwa Methanogenic ni ubwoko butari apoptotique y'urupfu rw'utugingo ngengabuzima bifitanye isano no kwegeranya vacuole ikomeye ya cytoplasme27. Inenge ya morphologiya igaragara muri pine nematode isa nkaho ifitanye isano rya bugufi nuburyo bwurupfu rwa metani. Isuzuma rya Microscopique mu bihe bitandukanye ryerekanye ko vacuole nini zakozwe nyuma ya 20 h zihura na 5-iodoindole (0.1 mM). Vacuole ya microscopique yagaragaye nyuma ya 8 h yo kuvurwa, kandi umubare wabo wiyongereye nyuma ya 12 h. Imyuka myinshi nini yagaragaye nyuma ya 14 h. Imyuka myinshi yahujwe yagaragaye neza nyuma ya 12-16 h yo kuvurwa, byerekana ko guhuza vacuole ari ishingiro ryuburyo bwurupfu rwa methanogene. Nyuma yamasaha 20, habonetse vacuole nini nini zose. Ibyo kureba byerekana raporo yambere ya metuose muri C. elegans.
Muri inyo zivuwe na 5-iyode, hagaragaye no guteranya vacuole no guturika (Ishusho 5), nkuko bigaragazwa no kunama inyo no kurekura vacuole mu bidukikije. Ihungabana rya Vacuole ryagaragaye no mu gishishwa cy’amagi, ubusanzwe kibikwa neza na L2 mugihe cyo gutera (Ishusho yinyongera. S2). Izi nyigisho zishyigikira uruhare rwo kwegeranya amazi no kunanirwa kwa osmoregulatory, ndetse no gukomeretsa ingirabuzimafatizo (RCI), mugihe cyo gukora vacuole no kwiyongera (Ishusho 5).
Twibanze ku ruhare rwa iyode mu iyerekwa rya vacuole yagaragaye, twakoze iperereza ku bikorwa bya nematiside ya sodium iode (NaI) na potasiyumu iyode (KI). Nyamara, kuri concentration (0.1, 0.5 cyangwa 1 mM), ntabwo byagize ingaruka ku kubaho kwa nematode cyangwa kubaho kwa vacuole (Ishusho yinyongera. S5), nubwo 1 mM KI yagize ingaruka nkeya ya nematiside. Ku rundi ruhande, 7-iodoindole (1 cyangwa 2 mM), nka 5-iodoindole, yateje vacuole nyinshi no guhindura imiterere (Ishusho yinyongera. S6). Iyode ebyiri zerekanaga ibintu bisa na fenotipiki muri pine nematode, naho NaI na KI ntabwo. Igishimishije, indole ntabwo yatumye habaho vacuole muri B. xylophilus kuri concentration zageragejwe (amakuru aterekanwa). Rero, ibisubizo byemeje ko uruganda rwa indole-iyode rufite inshingano zo gukurura no guhindagurika kwa B. xylophilus.
Muri indoles zapimwe kubikorwa bya nematiside, 5-iodoindole yari ifite igipimo kinini cyo kunyerera cya -5.89 kcal / mol, ikurikirwa na 7-iodoindole (-4.48 kcal / mol), 4-fluoroindole (-4.33), na indole (-4.03) (Ishusho 6). Umugozi ukomeye wa hydrogène uhuza 5-iodoindole na leucine 218 bikomeza guhuza kwayo, mugihe izindi nkomoko zose za indole zihuza serine 260 binyuze mumurongo wa hydrogen. Mubindi byitegererezo bya iyode, 2-iodoindole ifite agaciro kangana na -5.248 kcal / mol, ibyo bikaba biterwa numubano wingenzi wa hydrogène hamwe na leucine 218. Ibindi bizwi bizwi harimo 3-iodoindole (-4.3 kcal / mol), 4-iodoindole (-4.0 kcal / mol), na 6-fluoroindole (-2). Indogene nyinshi ya halogene kandi irigaragaza ubwayo, usibye 5-iodoindole na 2-iodoindole, ikora isano na serine 260. Kuba hydrogène ihuza na leucine 218 byerekana guhuza neza reseptor-ligand, nkuko bigaragara kuri ivermectin (Ishusho yinyongera. GluCL reseptor ikoresheje leucine 218 (Igishusho 6 hamwe ninyongera. S8). Turasaba ko ibyo bisabwa bisabwa kugirango habeho imiterere ya pore ifunguye yikigo cya GluCL kandi ko muguhuza cyane kurubuga rukora rwa reseptor ya GluCL, 5-iodoindole, 2-iodoindole, avermectin na ivermectin bityo bigatuma umuyoboro wa ion ufunguka kandi bikemerera gufata amazi.
Molecular docking ya indole na halogenated indole kuri GluCL. Icyerekezo cyo guhuza (A) indole, (B) 4-fluoroindole, (C) 7-iodoindole, na (D) 5-iodoindole ligands kurubuga rukora rwa GluCL. Poroteyine igereranwa na lente, kandi imigozi ya hydrogène yumugongo yerekanwa nkumurongo utudomo wumuhondo. .
Hakozwe ubushakashatsi bwo gusuzuma ingaruka z'ubumara bwa 5-iodoindole ku kumera kw'imyumbati n'imbuto za radis. 5-iodoindole (0.05 cyangwa 0.1 mM) cyangwa avermectine (10 μg / mL) ntacyo byagize cyangwa ntigire ingaruka ku kumera kwambere no kuvuka kw'ibimera (Ishusho 7). Byongeye kandi, nta tandukaniro rikomeye ryabonetse hagati yikigereranyo cyo kumera kwimbuto zitavuwe nimbuto zavuwe na 5-iodoindole cyangwa avermectine. Ingaruka zo kurambura taproot n'umubare w'imizi yaturutseho ntibyari bike, nubwo mM 1 (inshuro 10 yibikorwa byayo) ya 5-iodoindole yatinze gato gukura kumizi. Ibisubizo byerekana ko 5-iodoindole idafite ubumara bwo gutera ingirabuzimafatizo kandi ntibibangamira iterambere ry’ibihingwa ku bushakashatsi bwakozwe.
Ingaruka ya 5-iodoindole kumera kwimbuto. Kumera, kumera no gushinga imizi ya B. oleracea na R. raphanistrum imbuto kuri Murashige na Skoog agar medium hamwe cyangwa idafite avermectine cyangwa 5-iodoindole. Kumera byanditswe nyuma yiminsi 3 yubushakashatsi kuri 22 ° C.
Ubu bushakashatsi butangaza ibibazo byinshi byica nematode by indoles. Icy'ingenzi, iyi niyo raporo yambere ya iodoindole itera methylation (inzira iterwa no kwegeranya vacuole nto zigenda zivanga buhoro buhoro zinjira mu cyuho kinini, amaherezo zikaviramo guturika no gupfa) mu nshinge za pinusi, hamwe na iodoindole yerekana ibintu byingenzi bya nematiside bisa n’ubucuruzi bwa nematicide avermectin.
Indoles zavuzwe mbere zikora ibikorwa byinshi byerekana ibimenyetso muri prokaryote na eukaryote, harimo kubuza biofilm kubuza / gukora, kubaho kwa bagiteri, no gutera indwara 19,32,33,34. Vuba aha, ingaruka zishobora kuvura indwara ya halogene, indole alkaloide, hamwe nimbuto zikomoka kuri semisynthetic indole zashishikaje ubushakashatsi 35,36,37. Kurugero, indogene ya halogene yerekanwe kwica Escherichia coli idahoraho hamwe na selile Staphylococcus aureus37. Byongeye kandi, ni inyungu za siyanse kwiga ku mikorere ya indogene ya halogene irwanya ayandi moko, genera, n’ubwami, kandi ubu bushakashatsi ni intambwe iganisha kuri iyi ntego.
Hano, turasaba uburyo bwokwica 5-iodoindole yatewe na C. elegans hashingiwe ku gukomeretsa ingirabuzimafatizo (RCI) na methylation (Ishusho 4C na 5). Impinduka zikomeye nka puffness na vacuolar degeneration ni ibipimo bya RCI na methylation, bigaragarira nka vacuole nini muri cytoplazme 48,49. RCI ibangamira umusaruro w'ingufu igabanya umusaruro wa ATP, itera kunanirwa kwa pompe ya ATPase, cyangwa guhagarika ingirabuzimafatizo no gutera umuvuduko mwinshi wa Na +, Ca2 +, n'amazi 50,51,52. Vacuole ya Intracytoplasmic ituruka mu ngirabuzimafatizo zinyamaswa bitewe no kwegeranya amazi muri cytoplazme bitewe no kwinjira kwa Ca2 + n’amazi53. Igishimishije, ubu buryo bwo kwangirika kwakagari burashobora guhinduka mugihe ibyangiritse ari byigihe gito kandi selile zigatangira kubyara ATP mugihe runaka, ariko niba ibyangiritse bikomeje cyangwa bikabije, selile zirapfa.54 Ibyo twabonye byerekana ko nematode ivurwa na 5-iodoindole idashobora kugarura biosynthesis isanzwe nyuma yo guhura nibibazo byikibazo.
Methylation phenotype yatewe na 5-iodoindole muri B. xylophilus irashobora guterwa no kuba hariho iyode no gukwirakwiza molekile yayo, kubera ko 7-iodoindole itagize ingaruka nke zo kubuza B. xylophilus kurenza 5-iodoindole (Imbonerahamwe 1 n’ishusho yinyongera S6). Ibisubizo birahuye nibice byubushakashatsi bwa Maltese nibindi. . Imikoranire hagati ya indole cyangwa halogenated indoles hamwe na reseptor ya GluCL yagaragaye muri ubu bushakashatsi nayo ishyigikira iki gitekerezo, kuko wasangaga 5- na 2-iodoindole ihuza abayakira ba GluCL cyane kurusha izindi indoles zasuzumwe (Igicapo cya 6 n’inyongera S8). Iyode iri ku mwanya wa kabiri cyangwa uwa gatanu wa indole wasangaga ihuza leucine 218 ya reseptor ya GluCL ikoresheje imigozi ya hydrogène ya hydrogène, mu gihe izindi indogene za halogene hamwe na indole ubwazo zigira intege nke za hydrogène ya hydrogène hamwe na serine 260 (Ishusho 6). Turakeka rero ko kuba halogen igira uruhare runini mu kwinjiza kwangirika kwa vacuolar, mu gihe guhuza cyane 5-iodoindole bituma umuyoboro wa ion ufunguka, bityo bigatuma amazi yinjira vuba kandi akavunika. Nyamara, uburyo burambuye bwibikorwa bya 5-iodoindole buracyamenyekana.
Mbere yo gushyira mubikorwa 5-iodoindole, ingaruka zayo zubumara ku bimera zigomba gusesengurwa. Ubushakashatsi bwacu bwo kumera kwimbuto bwerekanye ko 5-iodoindole itagize ingaruka mbi kumera kwimbuto cyangwa inzira ziterambere ryakurikiyeho (ubushakashatsi 7). Niyo mpamvu, ubu bushakashatsi butanga urufatiro rwo gukoresha 5-iodoindole mu bidukikije kugira ngo hirindwe ingaruka mbi za pine nematode ku biti bya pinusi.
Raporo zabanje zerekanye ko ubuvuzi bushingiye kuri indole bugaragaza uburyo bushoboka bwo gukemura ikibazo cyo kurwanya antibiyotike no gutera kanseri55. Byongeye kandi, indoles zifite antibacterial, anticancer, antioxidant, anti-inflammatory, antidiabete, antiviral, antiproliferative na antituberculose kandi zishobora kuba umusingi utanga ikizere cyo guteza imbere ibiyobyabwenge 56,57. Ubu bushakashatsi bwerekana ku nshuro ya mbere uburyo bwo gukoresha iyode nka antiparasitike na anthelmintic.
Avermectin yavumbuwe mu myaka mirongo itatu ishize kandi yegukana igihembo cyitiriwe Nobel mu 2015, kandi ikoreshwa nka anthemmintique iracyakomeza. Ariko, kubera iterambere ryihuse ryo kurwanya avermectine muri nematode nudukoko twangiza udukoko, hakenewe ubundi buryo, buhendutse, kandi bwangiza ibidukikije kugira ngo hirindwe ubwandu bwa PWN mu biti bya pinusi. Ubu bushakashatsi kandi butangaza uburyo 5-iodoindole yica pine nematode kandi ko 5-iodoindole ifite uburozi buke ku ngirabuzimafatizo, ibyo bikaba bitanga amahirwe menshi yo gukoreshwa mu bucuruzi.
Ubushakashatsi bwose bwemejwe na komite ishinzwe imyitwarire ya kaminuza ya Yeungnam, Gyeongsan, muri Koreya, kandi uburyo bwakozwe hakurikijwe amabwiriza ya komite ishinzwe imyitwarire ya kaminuza ya Yeungnam.
Igeragezwa ryamagi ryakozwe hakoreshejwe inzira zashyizweho43. Kugira ngo hamenyekane igipimo cy’imyororokere (HR), nematode yumunsi wumunsi 1 (hafi 100 wigitsina gore nigitsina gabo 100) yimuriwe mubiryo bya Petri birimo ibihumyo hanyuma yemererwa gukura kuri 24 h. Amagi yahise yitandukanya hanyuma avurwa na 5-iodoindole (0,05 mM na 0.1 mM) cyangwa avermectine (10 μg / ml) nk'ihagarikwa mu mazi meza. Izi mpagarike (500 μl; hafi amagi 100) zimuriwe ku mariba y’isahani y’umuco 24 yuzuye kandi yashyizwe kuri 22 ° C. Ibarura rya L2 ryakozwe nyuma ya 24 h ya incububasi ariko byafatwaga nkuwapfuye niba selile zitimutse mugihe zashizwemo insinga nziza ya platine. Ubu bushakashatsi bwakozwe mubyiciro bibiri, buri kimwe gisubiramo bitandatu. Amakuru yavuye mubushakashatsi bwombi yarahujwe kandi aratangwa. Ijanisha rya HR ribarwa kuburyo bukurikira:
Impfu nini zapimwe hakoreshejwe uburyo bwateguwe mbere. Amagi ya Nematode yakusanyirijwe hamwe insoro zahujwe no kuvoma mumazi meza atoboye kugirango habeho lisiti ya L2. Ibinyomoro bisanisha (hafi 500 nematode) byavuwe hamwe na 5-iodoindole (0,05 mM na 0.1 mM) cyangwa avermectine (10 μg / ml) hanyuma bikorerwa kuri plaque B. cinerea Petri. Nyuma ya 48 h ya incububasi kuri 22 ° C, nematode yakusanyirijwe mumazi meza ya sterile hanyuma basuzumwa ko hari L2, L3, na L4. Kubaho kwa L3 na L4 byerekanaga ihinduka ryinshi, mugihe kuba icyiciro cya L2 byerekanaga ko nta gihinduka. Amashusho yabonetse ukoresheje iRiS System Sisitemu ya Digital Cell Imaging Sisitemu. Ubu bushakashatsi bwakozwe mubyiciro bibiri, buri kimwe gisubiramo bitandatu. Amakuru yavuye mubushakashatsi bwombi yarahujwe kandi aratangwa.
Uburozi bwa 5-iodoindole na avermectin ku mbuto bwasuzumwe hifashishijwe ibizamini byo kumera ku masahani ya agara ya Murashige na Skoog.62 B. oleracea na R. raphanistrum imbuto zabanje kwibizwa mu mazi ya sterile umunsi umwe, zogejwe na ml 1% 100% ya etanol, hamwe na ml 1% ya sodium ya 15% hamwe na mitiweli ya 3%. Imbuto zashizwemo noneho zashyizwe ku isahani yimeza irimo 0.86 g / l (0.2X) Murashige na Skoog hamwe na 0.7% ya bacteriologiya hamwe na 5-iodoindole cyangwa avermectine. Amasahani yahise ashyirwa kuri 22 ° C, kandi amashusho yafashwe nyuma yiminsi 3 yubushakashatsi. Ubu bushakashatsi bwakozwe mu byiciro bibiri, buri kimwekimwe cyari gifite inshuro esheshatu.
Igihe cyo kohereza: Gashyantare-26-2025