I-pine nematode yi-endoparasite yokufuduka ehlala abantu abasuka kwenye indawo eyaziwa ngokuba ibangela ilahleko enkulu kwezoqoqosho kwiindawo eziphilayo zehlathi lephayini. Olu phononongo luphonononga umsebenzi wokubulala ii-indole ze-halogenated ngokuchasene nee-pine nematode kunye nendlela ezisebenza ngayo. Imisebenzi yokubulala ii-pine nematode ze-5-iodoindole kunye ne-avermectin (ulawulo oluhle) ngokuchasene nee-pine nematode yayifana kwaye iphezulu kumanqanaba aphantsi (10 μg/mL). I-5-iodoindole inciphise ubudoda, umsebenzi wokuzala, ukufa kwe-embryonic kunye ne-larval, kunye nokuziphatha kwe-locomotor. Ukusebenzisana kwee-molecular ze-ligands kunye nee-receptors ze-chloride channel ze-glutamate-gated ezingenamqolo zixhasa ingcamango yokuba i-5-iodoindole, njenge-avermectin, ibopha ngokuqinileyo kwindawo esebenzayo ye-receptor. I-5-Iodoindole ikwabangele ukuguquguquka okuhlukeneyo kwe-phenotypic kwi-nematode, kubandakanya ukuwa/ukuncipha okungaqhelekanga kwamalungu omzimba kunye nokwanda kwe-vacuolization. Ezi ziphumo zibonisa ukuba ii-vacuoles zinokudlala indima ekufeni okubangelwa yi-nematode methylation. Okubalulekileyo kukuba, i-5-iodoindole yayingenabungozi kuzo zombini iintlobo zezityalo (ikhaphetshu kunye neradish). Ngoko ke, olu phononongo lubonisa ukuba ukusetyenziswa kwe-iodoindole phantsi kweemeko zendalo kunokulawula ukwenzakala kwe-pine wilt.
I-pine wood nematode (iBursaphelenchus xylophilus) yeyama-pine wood nematodes (PWN), ama-nematodes afudukayo ahlala ephila aziwa ngokubanga umonakalo omkhulu kwindalo kwindalo kwi-pine forest ecosystems1. Isifo se-pine wilt (PWD) esibangelwa yi-pine wood nematode siba yingxaki enkulu kumazwekazi aliqela, kuquka i-Asia neYurophu, kwaye eMntla Melika, le pine itshabalalisa iintlobo ze-pine ezingenisiweyo1,2. Ukuncipha kwemithi ye-pine yingxaki enkulu yezoqoqosho, kwaye amathuba okusasazeka kwayo kwihlabathi liphela axhalabisa3. Ezi ntlobo zilandelayo ze-pine zihlaselwa kakhulu yi-nematode: i-Pinus densiflora, i-Pinus sylvestris, i-Pinus thunbergii, i-Pinus koraiensis, i-Pinus thunbergii, i-Pinus thunbergii, kunye ne-Pinus radiata4. I-pine nematode sisifo esibi esinokubulala imithi ye-pine kwiiveki okanye kwiinyanga emva kosulelo. Ukongeza, ukuqhambuka kwe-pine nematode kuqhelekile kwiindawo ezahlukeneyo zendalo, ngoko ke kuye kwasekwa iindlela zosulelo eziqhubekayo1.
I-Bursaphelenchus xylophilus yi-nematode yezityalo ezihlala zodwa ekwi-superfamily Aphelenchoidea kunye ne-clade 102.5. I-nematode itya i-fungi kwaye izala kwizicubu zomthi wemithi yephayini, ikhula ibe ngamanqanaba amane ahlukeneyo e-larval: L1, L2, L3, L4 kunye nomntu omdala1,6. Phantsi kweemeko zokunqongophala kokutya, i-pine nematode idlulela kwinqanaba elikhethekileyo le-larval - i-dauer, edla i-vector yayo - i-pine bark beetle (i-Monochamus alternatus) kwaye idluliselwe kwimithi yephayini esempilweni. Kwiindawo eziphilileyo, i-nematodes ifuduka ngokukhawuleza kwizicubu zezityalo kwaye itye iiseli ze-parenchymatous, nto leyo ekhokelela kwiimpendulo ezininzi ze-hypersensitivity, ukubuna kwephayini kunye nokufa kungakapheli unyaka emva kosulelo1,7,8.
Ulawulo lwebhayoloji lwee-nematodes zephayini belusoloko lungumceli mngeni, kunye namanyathelo okuvalelwa edwaleni aqala kwinkulungwane yama-20. Amaqhinga angoku okulawula ii-nematodes zephayini abandakanya unyango lweekhemikhali, kubandakanya ukufutha imithi kunye nokufakelwa kwee-nematicides kwiziqu zemithi. Ezona nematicides zisetyenziswa kakhulu yi-avermectin kunye ne-avermectin benzoate, ezikusapho lwe-avermectin. Ezi khemikhali zibiza kakhulu zisebenza kakhulu kwiintlobo ezininzi zee-nematode kwaye zithathwa njengezikhuselekileyo kwindalo9. Nangona kunjalo, ukusetyenziswa ngokuphindaphindiweyo kwezi nematicides kulindeleke ukuba kudale uxinzelelo lokukhetha oluya kukhokelela ekuveleni kwee-nematodes zephayini ezichaseneyo, njengoko kubonisiwe kwizinambuzane ezininzi, ezifana neLeptinotarsa decemlineata, iPlutella xylostella kunye nee-nematodes iTrichostrongylus colubriformis kunye ne-Ostertagia circumcincta, eziye zakhula kancinci kancinci ukuxhathisa ii-avermectins10,11,12. Ke ngoko, iipateni zokuxhathisa kufuneka zifundwe rhoqo kwaye ii-nematicides zihlolwe rhoqo ukuze kufunyanwe amanye amanyathelo, angabizi kakhulu kwaye anobuhlobo nokusingqongileyo okulawula i-PVD. Kwiminyaka yakutshanje, ababhali abaninzi bacebise ukusetyenziswa kwezicatshulwa zezityalo, iioyile ezibalulekileyo kunye ne-volatiles njengeearhente zokulawula i-nematode13,14,15,16.
Kutshanje sibonise umsebenzi wokubulala iintsholongwane we-indole, imolekyuli yesignali ephakathi kweeseli kunye ne-interkingdom, kwi-Caenorhabditis elegans 17. I-Indole sisignali esaziwayo ngaphakathi kweeseli kwi-microbial ecology, elawula imisebenzi emininzi echaphazela i-microbial physiology, ukwakheka kwe-spore, uzinzo lwe-plasmid, ukumelana namayeza, ukwakheka kwe-biofilm, kunye ne-virulence 18, 19. Umsebenzi we-indole kunye nezinto eziphuma kuyo ngokuchasene nezinye ii-nematode ezibangela izifo awukafundwa. Kolu phononongo, siphande umsebenzi wokubulala iintsholongwane we-indole ezingama-34 ngokuchasene nee-pine nematode kwaye sacacisa indlela yokusebenza kwe-5-iodoindole enamandla kakhulu sisebenzisa i-microscopy, i-time-lapse photography, kunye novavanyo lwe-molecular docking, kwaye savavanya iziphumo zayo ezinobuthi kwizityalo sisebenzisa uvavanyo lokuhluma kwembewu.
Uxinzelelo oluphezulu (>1.0 mM) lwe-indole ngaphambili luxelwe ukuba lunefuthe lokubulala iintsholongwane kwi-nematodes17. Emva konyango lwe-B. xylophilus (amanqanaba obomi adibeneyo) nge-indole okanye i-33 ezahlukeneyo ze-indole derivatives kwi-1 mM, ukufa kwe-B. xylophilus kwalinganiswa ngokubala iintsholongwane eziphilayo nezifileyo kumaqela olawulo nawanyangwayo. Ii-indoles ezintlanu zibonakalise umsebenzi obalulekileyo wokubulala iintsholongwane; ukusinda kweqela lolawulo elinganyangwanga kwakuyi-95 ± 7% emva kweeyure ezingama-24. Kwi-indoles ezingama-34 ezivavanyiweyo, i-5-iodoindole kunye ne-4-fluoroindole kwi-1 mM zibangele ukufa kwe-100%, ngelixa i-5,6-difluoroindigo, i-methylindole-7-carboxylate, kunye ne-7-iodoindole zibangele ukufa okumalunga ne-50% (Itheyibhile 1).
Isiphumo se-5-iodoindole ekwakhiweni kwe-vacuole kunye ne-metabolism ye-pine wood nematode. (A) Isiphumo se-avermectin kunye ne-5-iodoindole kwi-nematodes yamadoda amadala, (B) amaqanda e-L1 stage nematode kunye (C) metabolism ye-B. xylophilus, (i) ii-vacuole azibonwanga kwi-0 h, unyango lubangele (ii) ii-vacuole, (iii) ukuqokelelwa kwee-vacuole ezininzi, (iv) ukudumba kwee-vacuole, (v) ukudibana kwee-vacuole kunye (vi) ukwakheka kwee-vacuole ezinkulu. Iintolo ezibomvu zibonisa ukudumba kwee-vacuole, iintolo eziluhlaza okwesibhakabhaka zibonisa ukudibana kwee-vacuole kwaye iintolo ezimnyama zibonisa ii-vacuole ezinkulu. Ibha yesikali = 50 μm.
Ukongeza, olu phononongo lukwachaze inkqubo elandelelanayo yokufa okubangelwa yi-methane kwii-pine nematodes (Umfanekiso 4C). Ukufa kwe-methanogenic luhlobo lokufa kweeseli olungengolwa-apoptotic olunxulunyaniswa nokuqokelelwa kwee-cytoplasmic vacuoles ezibalaseleyo27. Iziphene ze-morphological ezibonwe kwii-pine nematodes zibonakala zinxulumene kakhulu nendlela yokufa okubangelwa yi-methane. Uvavanyo lwe-Microscopic ngamaxesha ahlukeneyo lubonise ukuba ii-giant vacuoles zenziwe emva kweeyure ezingama-20 zokuvezwa kwi-5-iodoindole (0.1 mM). Ii-microscopic vacuoles zabonwa emva kweeyure ezi-8 zonyango, kwaye inani lazo landa emva kweeyure ezili-12. Ii-vacuoles ezininzi ezinkulu zabonwa emva kweeyure ezili-14. Ii-fused vacuoles ezininzi zabonakala ngokucacileyo emva kweeyure ezili-12-16 zonyango, nto leyo ebonisa ukuba i-vacuole fusion sisiseko se-methanogenic death mechanism. Emva kweeyure ezingama-20, ii-giant vacuoles ezininzi zafunyanwa kulo lonke i-worm. Ezi ngqwalasela zimele ingxelo yokuqala ye-metuosis kwi-C. elegans.
Kwiimpethu eziphathwe nge-5-iodoindole, ukuhlangana kwe-vacuole kunye nokuqhekeka kwayo kwabonwa (Umzobo 5), njengoko kungqinwa kukugoba kwempethu kunye nokukhululwa kwe-vacuole kwindawo esingqongileyo. Ukuphazamiseka kwe-vacuole kwabonwa nakwi-membrane yegobolondo leqanda, edla ngokugcinwa ingaphazanyiswa yi-L2 ngexesha lokuqanduselwa (Umzobo Ongezelelweyo S2). Ezi ngqwalasela zixhasa ukubandakanyeka kokuqokelelana kolwelo kunye nokusilela kwe-osmoregulatory, kunye nokulimala kweseli okuguquguqukayo (RCI), kwinkqubo yokwakheka kwe-vacuole kunye ne-suppuration (Umzobo 5).
Sicinga ngendima ye-iodine ekwakhiweni kwe-vacuole okubonweyo, siphande umsebenzi wokubulala i-sodium iodide (NaI) kunye ne-potassium iodide (KI). Nangona kunjalo, kumanqanaba (0.1, 0.5 okanye 1 mM), azizange zichaphazele ukusinda kwe-nematode okanye ukwakheka kwe-vacuole (Umzobo oNcedisayo S5), nangona i-1 mM KI yayinefuthe elincinci lokubulala i-nematic. Kwelinye icala, i-7-iodoindole (1 okanye 2 mM), njenge-5-iodoindole, yabangela ii-vacuoles ezininzi kunye nokuguquguquka kwesakhiwo (Umzobo oNcedisayo S6). Ii-iodoindoles ezimbini zibonise iimpawu ezifanayo ze-phenotypic kwi-pine nematodes, ngelixa i-NaI kunye ne-KI azizange zibonise. Okubangel 'umdla kukuba, i-indole ayizange ibangele ukwakheka kwe-vacuole kwi-B. xylophilus kumanqanaba avavanyiweyo (idatha ayiboniswanga). Ke ngoko, iziphumo ziqinisekisile ukuba i-indole-iodine complex inoxanduva lwe-vacuolization kunye ne-metabolism ye-B. xylophilus.
Phakathi kwee-indoles ezivavanyiweyo ukuba azinazo iintsholongwane, i-5-iodoindole yayine-slip index ephezulu kakhulu ye--5.89 kcal/mol, ilandelwa yi-7-iodoindole (-4.48 kcal/mol), i-4-fluoroindole (-4.33), kunye ne-indole (-4.03) (Umfanekiso 6). Ukubopha okuqinileyo kwe-hydrogen ye-5-iodoindole kwi-leucine 218 kuzinzisa ukubopha kwayo, ngelixa zonke ezinye izinto eziphuma kwi-indole zibopha kwi-serine 260 nge-side chain hydrogen bonds. Phakathi kwezinye ii-iodoindoles ezimodeliweyo, i-2-iodoindole inexabiso lokubopha eliyi--5.248 kcal/mol, elibangelwa kukudityaniswa kwayo okuphambili kwe-hydrogen ne-leucine 218. Ezinye izibophelelo ezaziwayo ziquka i-3-iodoindole (-4.3 kcal/mol), i-4-iodoindole (-4.0 kcal/mol), kunye ne-6-fluoroindole (-2.6 kcal/mol) (Umfanekiso ongezelelweyo S8). Uninzi lwee-indole ezine-halogenated kunye ne-indole ngokwazo, ngaphandle kwe-5-iodoindole kunye ne-2-iodoindole, zenza i-bond ne-serine 260. Inyani yokuba i-hydrogen bonding ne-leucine 218 ibonisa ukuba i-receptor-ligand binding iyasebenza, njengoko kubonwe kwi-ivermectin (Umzobo oNcedisayo S7), iqinisekisa ukuba i-5-iodoindole kunye ne-2-iodoindole, njenge-ivermectin, zibopha ngokuqinileyo kwindawo esebenzayo ye-GluCL receptor nge-leucine 218 (Umzobo 6 kunye nomzobo oNcedisayo S8). Sicebisa ukuba oku kubopha kuyimfuneko ukugcina isakhiwo se-pore evulekileyo ye-GluCL complex kwaye ngokubopha ngokuqinileyo kwindawo esebenzayo ye-GluCL receptor, i-5-iodoindole, i-2-iodoindole, i-avermectin kunye ne-ivermectin ngaloo ndlela zigcina umjelo we-ion uvulekile kwaye uvumele ukufunxwa kolwelo.
Ukufakwa kwemolekyuli ye-indole kunye ne-halogenated indole kwi-GluCL. Ukudibana kwe-(A) indole, (B) 4-fluoroindole, (C) 7-iodoindole, kunye (D) 5-iodoindole ligands kwindawo esebenzayo ye-GluCL. Iproteni imelwe yiribhoni, kwaye i-backbone hydrogen bonds iboniswa njengemigca etyheli enamachaphaza. (A′), (B′), (C′), kunye (D′) zibonisa ukusebenzisana kwee-ligands ezihambelanayo nee-amino acid residues ezijikelezileyo, kwaye i-side-chain hydrogen bonds iboniswa ziintolo ezinamachaphaza apinki.
Kwenziwe uvavanyo lokuvavanya impembelelo enetyhefu ye-5-iodoindole ekukhuleni kwembewu yekhaphetshu kunye neradish. I-5-iodoindole (0.05 okanye 0.1 mM) okanye i-avermectin (10 μg/mL) yayinempembelelo encinci okanye ayizange ibe nayo ekukhuleni kokuqala kunye nokuvela kwezityalo (Umfanekiso 7). Ukongeza, akukho mahluko ubalulekileyo ofunyenweyo phakathi kwesantya sokuhluma kwezityalo ezilawulwayo ezinganyangwanga kunye nembewu ephathwe nge-5-iodoindole okanye i-avermectin. Impembelelo ekukhuleni kweengcambu ezithambileyo kunye nenani leengcambu ezisecaleni ezenziweyo yayingabalulekanga, nangona i-1 mM (ixesha eliphindwe kalishumi uxinzelelo lwayo olusebenzayo) ye-5-iodoindole ilibazise kancinci uphuhliso lweengcambu ezisecaleni. Ezi ziphumo zibonisa ukuba i-5-iodoindole ayinabungozi kwiiseli zezityalo kwaye ayiphazamisi iinkqubo zophuhliso lwezityalo kumanqanaba afundwe.
Isiphumo se-5-iodoindole ekukhuleni kwembewu. Ukuhluma, ukuhluma kunye nokuhluma kweengcambu ecaleni kwembewu ye-B. oleracea kunye ne-R. raphanistrum kwi-Murashige kunye ne-Skoog agar medium ene-avermectin okanye i-5-iodoindole okanye engenayo. Ukuhluma kwarekhodwa emva kweentsuku ezi-3 zokufukama kwi-22°C.
Olu phononongo lubika amatyala aliqela okubulawa kwe-nematode yi-indoles. Okubalulekileyo kukuba, le yingxelo yokuqala ye-iodoindole ebangela i-methylation (inkqubo ebangelwa kukuqokelelwa kwe-vacuoles ezincinci ezidibana kancinci kancinci zibe yi-vacuoles ezinkulu, ekugqibeleni zikhokelela ekuqhekekeni kwe-membrane nokufa) kwiinaliti zephayini, i-iodoindole ibonisa iipropati ezibalulekileyo zokubulala i-nematic ezifana nezo ze-avermectin ye-nematicide yorhwebo.
Ii-Indoles ziye zaxelwa ngaphambili ukuba zinemisebenzi emininzi yokubonisa kwi-prokaryotes nakwi-eukaryotes, kubandakanya ukuthintela/ukwakheka kwe-biofilm, ukusinda kwebhaktheriya, kunye ne-pathogenicity19,32,33,34. Kutshanje, iziphumo zonyango ezinokubakho ze-halogenated indoles, i-indole alkaloids, kunye ne-semisynthetic indole derivatives ziye zatsala umdla omkhulu wophando35,36,37. Umzekelo, ii-halogenated indoles zibonakalisiwe ukuba zibulala iiseli ze-Escherichia coli eziqhubekayo kunye ne-Staphylococcus aureus37. Ukongeza, kunomdla wesayensi ukufunda ukusebenza kwe-halogenated indoles ngokuchasene nezinye iintlobo, izizukulwana, kunye nezikumkani, kwaye olu phononongo linyathelo eliya ekufezekiseni le njongo.
Apha, sicebisa indlela yokwenza i-5-iodoindole-induced lethality kwi-C. elegans ngokusekelwe kwingozi yeseli eguqukayo (RCI) kunye ne-methylation (Imifanekiso 4C kunye no-5). Utshintsho oluvela ekufubeni olufana nokudumba kunye nokuwohloka kwe-vacuolar zizibonakaliso ze-RCI kunye ne-methylation, ezibonakaliswa njenge-vacuoles ezinkulu kwi-cytoplasm48,49. I-RCI iphazamisa ukuveliswa kwamandla ngokunciphisa ukuveliswa kwe-ATP, ibangela ukungasebenzi kwepompo ye-ATPase, okanye iphazamise ii-membranes zeseli kwaye ibangele ukungena ngokukhawuleza kwe-Na+, Ca2+, kunye ne-water50,51,52. Ii-vacuoles ze-intracytoplasmic zivela kwiiseli zezilwanyana ngenxa yokuqokelelana kolwelo kwi-cytoplasm ngenxa yokungena kwe-Ca2+ kunye ne-water53. Okunomdla kukuba, le ndlela yokwenza umonakalo weseli iyakwazi ukutshintshwa ukuba umonakalo ngowexeshana kwaye iiseli ziqala ukuvelisa i-ATP kangangexesha elithile, kodwa ukuba umonakalo uyaqhubeka okanye uba mandundu, iiseli ziyafa.54 Izinto esiziqapheleyo zibonisa ukuba ii-nematodes ezinyangwe nge-5-iodoindole azikwazi ukubuyisela i-biosynthesis eqhelekileyo emva kokuchaphazeleka kwiimeko zoxinzelelo.
I-methylation phenotype ebangelwa yi-5-iodoindole kwi-B. xylophilus isenokuba ibangelwa bubukho be-iodine kunye nokusasazwa kwayo kweemolekyuli, kuba i-7-iodoindole yayinefuthe elincinci lokuthintela kwi-B. xylophilus kune-5-iodoindole (Itheyibhile 1 kunye noMfanekiso oNcedisayo S6). Ezi ziphumo zihambelana ngokuyinxenye nezifundo zikaMaltese et al. (2014), ababike ukuba ukudluliselwa kwe-pyridyl nitrogen moiety kwi-indole ukusuka kwi-para- ukuya kwi-meta-position kuphelise i-vacuolization, ukuthintelwa kokukhula, kunye ne-cytotoxicity kwiiseli ze-U251, nto leyo ebonisa ukuba ukusebenzisana kwemolekyuli nendawo ethile esebenzayo kwiproteni kubalulekile27,44,45. Ukusebenzisana phakathi kwe-indole okanye i-halogenated indoles kunye nee-GluCL receptors ezibonwe kolu phononongo nazo ziyayixhasa le ngcamango, njengoko i-5- kunye ne-2-iodoindole zifunyenwe zibopha kwi-GluCL receptors ngamandla ngakumbi kunezinye ii-indoles ezihlolwe (Umfanekiso 6 kunye noMfanekiso oNcedisayo S8). I-iodine kwindawo yesibini okanye yesihlanu ye-indole ifunyenwe ibopha kwi-leucine 218 ye-GluCL receptor ngee-backbone hydrogen bonds, ngelixa ezinye ii-indole ze-halogenated kunye ne-indole ngokwazo zenza ii-weak side-chain hydrogen bonds kunye ne-serine 260 (Umfanekiso 6). Ngoko ke sicinga ukuba indawo ye-halogen idlala indima ebalulekileyo ekuqaliseni i-vacuolar degeneration, ngelixa ukubopha okuqinileyo kwe-5-iodoindole kugcina umjelo we-ion uvulekile, ngaloo ndlela kuvumela ukungena kolwelo ngokukhawuleza kunye nokuqhekeka kwe-vacuole. Nangona kunjalo, indlela eneenkcukacha yokusebenza kwe-5-iodoindole isaza kugqitywa.
Ngaphambi kokuba kusetyenziswe i-5-iodoindole, kufuneka kuhlalutywe impembelelo yayo enobuthi kwizityalo. Uvavanyo lwethu lokuhluma kwembewu lubonise ukuba i-5-iodoindole ayinayo impembelelo embi ekuhlumeni kwembewu okanye kwiinkqubo zophuhliso ezilandelayo kumanqanaba afundwe (Umfanekiso 7). Ngoko ke, olu phononongo lubonelela ngesiseko sokusetyenziswa kwe-5-iodoindole kwimeko-bume yendalo ukulawula ubungozi bee-pine nematodes kwimithi yephayini.
Iingxelo zangaphambili zibonise ukuba unyango olusekelwe kwi-indole lumele indlela enokwenzeka yokujongana nengxaki yokuxhathisa amayeza okubulala iintsholongwane kunye nokuqhubela phambili komhlaza55. Ukongeza, ii-indoles zinemisebenzi yokulwa neentsholongwane, yokulwa nomhlaza, i-antioxidant, i-anti-inflammatory, i-antidiabetic, i-antiviral, i-antiproliferative kunye ne-antituberculosis kwaye zinokusebenza njengesiseko esithembisayo sophuhliso lwamayeza56,57. Olu phononongo lubonisa okokuqala ukusetyenziswa kwe-iodine njenge-arhente yokulwa neentsholongwane kunye ne-anthelmintic.
I-Avermectin yafunyanwa kwiminyaka engamashumi amathathu eyadlulayo yaza yaphumelela iNobel Prize ngo-2015, kwaye ukusetyenziswa kwayo njenge-anthelmintic kusaqhubeka. Nangona kunjalo, ngenxa yophuhliso olukhawulezayo lokuxhathisa i-avermectins kwi-nematodes kunye nezinambuzane eziyingozi, kufuneka icebo elikhethekileyo, elingabizi kakhulu, nelinobuhlobo nokusingqongileyo ukulawula usulelo lwe-PWN kwimithi yephayini. Olu phononongo lukwaxela indlela i-5-iodoindole ebulala ngayo i-nematodes yephayini kwaye i-5-iodoindole inetyhefu ephantsi kwiiseli zezityalo, nto leyo evula amathuba amahle okusetyenziswa kwayo kwixesha elizayo kwezorhwebo.
Zonke iimvavanyo zivunyiwe yiKomiti yeMithetho yokuziphatha yeYunivesithi yaseYeungnam, eGyeongsan, eKorea, kwaye iindlela zenziwe ngokuhambelana nezikhokelo zeKomiti yeMithetho yokuziphatha yeYunivesithi yaseYeungnam.
Uvavanyo lokufukama amaqanda lwenziwe kusetyenziswa iinkqubo ezimiselweyo43. Ukuvavanya amazinga okufukama (HR), ii-nematodes ezindala ezinosuku olunye (malunga ne-100 yeemazi kunye neenkunzi ezili-100) zidluliselwe kwizitya zePetri eziqulethe ifungus kwaye zavunyelwa ukuba zikhule iiyure ezingama-24. Amaqanda emva koko ahlukaniswa aze anyangwe nge-5-iodoindole (0.05 mM kunye ne-0.1 mM) okanye i-avermectin (10 μg/ml) njenge-suspension emanzini acocekileyo acocekileyo. Ezi suspensions (500 μl; malunga namaqanda ali-100) zidluliselwe kwimithombo yeplate ye-tissue culture ye-24-well kwaye zafakwa kwi-22 °C. Ubalo lwe-L2 lwenziwa emva kweeyure ezingama-24 zokufukama kodwa zathathwa njengezifileyo ukuba iiseli azishukumi xa zivuselelwa ngentambo yeplatinum entle. Olu vavanyo lwenziwe ngamanqanaba amabini, ngalinye liphindaphindwa kathandathu. Idatha evela kuzo zombini iimvavanyo yadityaniswa kwaye yaboniswa. Ipesenti ye-HR ibalwa ngolu hlobo lulandelayo:
Ukufa kweembungu kuhlolwe kusetyenziswa iinkqubo ezaziphuhlisiwe ngaphambili. Amaqanda e-Nematode aqokelelwa kwaye iimbungu zadityaniswa ngokuqanduselwa emanzini acocekileyo acocekileyo ukuze kuveliswe iimbungu zesigaba se-L2. Iimbungu ezihambelanayo (malunga nama-500 eembungu) zanyangwa nge-5-iodoindole (0.05 mM kunye ne-0.1 mM) okanye i-avermectin (10 μg/ml) zaza zakhuliswa kwi-B. cinerea Petri plates. Emva kweeyure ezingama-48 zokufunxwa kwi-22 °C, iimbungu zaqokelelwa emanzini acocekileyo acocekileyo zaza zahlolwa ukuba zikhona na izigaba ze-L2, L3, kunye ne-L4. Ubukho bezigaba ze-L3 kunye ne-L4 bubonise utshintsho lweembungu, ngelixa ubukho besigaba se-L2 bubonise ukuba akukho tshintsho. Imifanekiso ifunyenwe kusetyenziswa i-iRiS™ Digital Cell Imaging System. Olu vavanyo lwenziwe ngamanqanaba amabini, ngalinye liphindaphindwa kathandathu. Idatha evela kuzo zombini iimvavanyo yadityaniswa yaza yaboniswa.
Ubuthi be-5-iodoindole kunye ne-avermectin kwimbewu buhlolwe kusetyenziswa uvavanyo lokuhluma kwiipleyiti ze-agar zeMurashige kunye neSkoog.62 Imbewu ye-B. oleracea kunye ne-R. raphanistrum zaqala zacwiliswa emanzini acocekileyo acocekileyo usuku olunye, zahlanjwa nge-1 ml ye-ethanol eyi-100%, zacocwa nge-1 ml ye-50% ye-bleach yorhwebo (3% ye-sodium hypochlorite) imizuzu eli-15, zaza zahlanjwa kahlanu ngamanzi acocekileyo ayi-1 ml. Imbewu ecocekileyo yacinezelwa kwiipleyiti ze-agar ezivelisa i-0.86 g/l (0.2X) yeMurashige kunye neSkoog medium kunye ne-0.7% ye-bacteriological agar ene-5-iodoindole okanye i-avermectin okanye engenazo. Iipleyiti zaza zafakwa kwi-22 °C, kwaye imifanekiso yathathwa emva kweentsuku ezi-3 zokufunxa. Olu vavanyo lwenziwe ngamanqanaba amabini, ngalinye lazo laphindaphindwa izihlandlo ezintandathu.
Ixesha leposi: Februwari-26-2025