Kungani i-methylene blue isetshenziswa njengodayi ophikisanayo ekudayeni okumelana ne-asidi

Uchwepheshe uhlola isikhwehlela esisabalele engilazini yesibonakhulu ngemva kokulandelana okuphelele kwezinyathelo zokudaya. Amagciwane abomvu amise okwezinduku agqama endaweni eluhlaza okwesibhakabhaka, bese kukhishwa umbiko othi kubonakale ama-bacilli amelana nokususwa kombala nge-asidi. Ngosuku olulandelayo, enye ingilazi yesibonakhulu ibonisa indawo eluhlaza okwesibhakabhaka ngokulinganayo, engenazo izinduku ezibomvu, bese kubikwa ukuthi ayinalo ufuba. Omunye wale mibiko usho okungaphezu kwalokho okusekelwa yingilazi yesibonakhulu, futhi akuwona owokuqala.

Ukudideka kusekuqondeni umqondo. Uchwepheshe wathatha udayi oluhlaza okwesibhakabhaka njengengxenye yekhemistri yokuthola amagciwane, sengathi i-methylene blue yasiza ekutholeni ama-mycobacteria. Ayizange. Ngesikhathi i-methylene blue ithinta ingilazi yesibonakhulu, ukuhlolwa kusuke sekwenzekile. Udayi udwebela ingemuva ngombala kuphela ukuze umphumela ufundeke. Lesi sihloko sichaza inhloso ye-methylene blue ekudayeni okumelana ne-asidi, ukuthi kungani ikhethwa iluhlaza okwesibhakabhaka kunokuba ibe ngomunye umbala, nokuthi okubomvu okuqhathaniswa nokuluhlaza okwesibhakabhaka kukuvumela ukuba uphethe ngokuthini.

Impendulo emfushane

Ukudaya okumelana ne-asidi kuwukuhlolwa kokuhlukanisa okwakhelwe phezu kwemibono emithathu. I-carbol fuchsin, engudayi oyinhloko, ifakwa ngenkani kuwo wonke amaseli, kuhlanganise nama-mycobacteria anolwelwesi olunjenge-wax. I-asidi notshwala, okuyisusambala, kususa udayi obomvu kumaseli avamile kodwa kwehluleke ukuwukhipha kulawo amelana nokususwa kombala nge-asidi. I-methylene blue, engudayi wombala ophikisanayo, ibe isidaya konke isusambala esikususe umbala, ngakho amaseli angamelani nokususwa kombala nge-asidi, amaseli omzimba ophethe amagciwane, nezinsalela kubonakala kuluhlaza okwesibhakabhaka, kuyilapho ama-bacilli amelana nokususwa kombala nge-asidi ehlala ebomvu. Udayi ophikisanayo unikeza ukuhlukana kwemibala. Awubambi qhaza kukhemistri eyenza iseli limelane nokususwa kombala nge-asidi.

Okwenziwa yisinyathelo ngasinye

Umdwebo ongezansi ubonisa ukuhlukaniswa kwemisebenzi. Wufunde usuka kwesobunxele uye kwesokudla njengenqubo, bese ufunda umugqa ongezansi njengesikhumbuzo sokuthi iqembu ngalinye lezindlela zokudaya libuza umbuzo ohlukile.

Ukungena kukadayi kuza kuqala. Ama-mycobacteria amelana nodayi abavamile abancibilikiswe emanzini ngoba ulwelwesi lwawo luwaxosha kakhulu amanzi, ngakho inqubo iphoqa ukungena kukadayi. Endleleni ye-Ziehl-Neelsen, ukushisisa kuze kuphume umhwamuko kungenisa i-carbol fuchsin equkethe i-phenol odongeni olunjenge-wax. Endleleni ebandayo ye-Kinyoun, ukushisa kuthathelwa indawo yingxube kadayi oyinhloko enamandla kakhulu, kanti ezinye izinhlobo eziguquliwe ezingasebenzisi ukushisa zengeza izinto ezinciphisa ukudonselana ebusweni boketshezi. Izinqubo zokudaya okumelana ne-asidi ze-American Society for Microbiology zichaza zombili izingxube zihambisana, futhi umehluko wokugxila kwazo mkhulu: cishe 0.3 g we-basic fuchsin ku-100 mL ngayinye endleleni eshisisiwe, uma kuqhathaniswa cishe no-4 g endleleni ebandayo.

Ukuhlukanisa kuza okwesibili. I-asidi notshwala, ngokuvamile okuyi-3 mL ye-hydrochloric acid egxilile ku-97 mL we-ethanol engamaphesenti angu-95, kugeza udayi obomvu kuwukhiphe kuwo wonke amaseli odonga lwawo olungenakho ukumelana okukhethekile. Amaseli anolwelwesi lwama-lipid lwama-mycobacteria olungonakele abambelela kudayi. Lesi yisinyathelo yonke indlela encike kuso, futhi yisona esithinteka kakhulu endleleni yokusebenza, njengoba kuxoxwa ngezansi.

Ukuhlukanisa imibala kuza ekugcineni. Ingilazi yesibonakhulu manje inamaseli abomvu amelana nokususwa kombala nge-asidi phezu kwengemuva lamaseli asuswe umbala, acishe angabi nambala. I-methylene blue engu-0.3 percent w/v ibuyisela ukubonakala kwalelo ngemuva ngombala ohluke kakhulu kobomvu. Ukufaka udayi ophikisanayo kuthatha isikhathi esifushane, ngokuvamile imizuzwana engu-20 kuya kwengu-30 endleleni eshisisiwe, kuze kufike emizuzwini embalwa kwezinye izinhlobo zendlela ebandayo; isikhathi sincike enqubweni, hhayi esicini sikadayi uqobo.

Kungani kukhethwa lo dayi, futhi kungani kuluhlaza okwesibhakabhaka

I-methylene blue iwudayi we-basic one-ion ene-charge ephozithivu, okusho ukuthi i-ion yayo enombala ithwala i-charge ephozithivu futhi inamathela ezindaweni ezine-charge enegethivu zamagciwane asuswe umbala, amaseli omzimba ophethe amagciwane, nezinsalela. I-OpenStax Microbiology ichaza lo mqondo wama-charge njengomgomo ojwayelekile wokusebenza kodayi be-basic. Kodwa i-methylene blue ayihlukile ngokukhethekile ngokwekhemistri lapha. I-crystal violet ne-safranin nazo zingodayi be-basic. Ukukhetha kusekelwe ekubonakaleni, hhayi kukhemistri.

Izinduku ezibomvu zidinga ingemuva elinombala ophikisanayo, futhi okuluhlaza okwesibhakabhaka kuphikisana nokubomvu ngendlela ewusizo kakhulu ekusebenziseni isibonakhulu senkundla ekhanyayo. I-safranin ingaba udayi ophikisanayo ongafanele ngoba umbala wayo wokugcina obomvana-obomvu usondelene kakhulu ne-carbol fuchsin egciniwe, ufiphaze wona kanye umehluko owakhiwe yinqubo. I-crystal violet inganikeza ingemuva elinsomi elimnyama elincintisana nama-bacilli abomvu azacile ekudonseni amehlo. Ubufakazi obuqine kakhulu bokuthi umsebenzi ukuhlukanisa imibala kunokuba kube yikhemistri bubonakala ekushintshweni kukadayi: odayi ophikisanayo abaluhlaza, njenge-malachite green ne-brilliant green, bayasetshenziswa kwezinye izinqubo zezilwane neziguquliwe, futhi ukuhlolwa kusasebenza, ngoba noma yimuphi umbala ohluke ngokucacile wenza umsebenzi ofanayo.

Kulokhu kulandela imiphumela emibili ephathekayo. Okokuqala, indlela evamile yokuxuba ayisho ukuthi kufanele kulungiswe i-pH; ungadidanisi lesi sixazululo esilula samanzi esingamaphesenti angu-0.3 nezingxube ezine-alkali ezifana ne-methylene blue ka-Loeffler, ezingezomunye umqondo wokudaya. Okwesibili, ukusebenzisa ngokweqile kuyalimaza. Izincwadi zesayensi yezicubu zixwayisa ngokucacile ukuthi ingemuva eliluhlaza okwesibhakabhaka eliminyene kakhulu lingafihla ama-bacilli ambalwa, ngakho ukushiya ingilazi igcwele udayi isikhathi eside kunesivunyelwe yinqubo kwehlisa ukufundeka ngaphandle kwenzuzo.

Ikhemistri yodonga eyenza ukuhlolwa kwenzeke

Isizathu sokuthi i-asidi notshwala kuhlukanise amaseli sisolwelwesini lwama-mycobacteria. Isisekelo sesakhiwo salo siyinhlanganisela ye-mycolyl-arabinogalactan-peptidoglycan: i-peptidoglycan exhunywe ku-arabinogalactan, emaphethelweni ayo angaphandle ethwala ama-mycolic acid anamaketanga amade kakhulu, kwakheke ulwelwesi lwangaphandle lwe-mycobacteria olunama-lipid amaningi futhi olungavumeli izinto ukuba zidlule kalula. Ukubuyekezwa kolwelwesi lwangaphandle lwama-mycobacteria kuchaza lesi sakhiwo ngokuningiliziwe. Lesi sivimbelo sisebenza kuzo zombili izinhlangothi. Sivimbela ukungena kukadayi, yingakho ukungena kudinga ukushisa noma udayi oyinhloko oqinisiwe, futhi sivimbela ukukhishwa kukadayi, yingakho i-asidi notshwala kwehluleka ukususa i-carbol fuchsin uma isingaphakathi. Amagciwane avamile awanalo lolu lwelwesi futhi alahlekelwa udayi. Ngakho ukumelana nokususwa kombala nge-asidi kuqondakala kangcono njengomphumela wesivimbelo sama-lipid, hhayi njengokubopha kwe-fuchsin ku-mycolic acid ngokwekhemistri.

Le khemistri iphinde ichaze indida evamile: ama-mycobacteria adaya kabi nangendlela engabikezeleki ngendlela ye-Gram, kwesinye isikhathi abonakale eyizimo ezifiphele ezinjengobuhlalu noma eyizithunzi ezingadayekile. Udonga lwawo oluxosha amanzi lunciphisa ukungena kwe-crystal violet. Ukudaya okumelana ne-asidi akukona ukudaya kwe-Gram okuguquliwe. Kungumgomo ohlukile wokuhlukanisa owakhiwe phezu kwesici esihlukile sodonga, njengoba ukuqhathaniswa kokudaya okulula nokudaya kwe-Gram kubonisa kwezinye izindlela ezimbili.

Izindlela ezintathu, imibuzo emithathu

Ukuze ufunde ngeqembu elibanzi lezindlela zokudaya amagciwane ngodayi owodwa, bheka i-atlas yokudaya ngesibonakhulu.

Lokho okubomvu phezu kokuluhlaza okwesibhakabhaka okungeke kukufakazele

Izinduku ezibomvu zisho ukuthi izinto eziphilayo ezincane zigcine i-carbol fuchsin ngesikhathi sokugezwa nge-asidi notshwala. Lokho okutholakele kuhambisana ne-Mycobacterium tuberculosis, kodwa isiqondiso se-CDC sokuxilonga ufuba siphawula ngokucacile ukuthi namanye ama-mycobacteria amelana nokususwa kombala nge-asidi, ngakho ukuhlonza uhlobo kudinga ukukhuliswa kwamagciwane nokuhlolwa kwe-nucleic acid, hhayi umbala.

Amaphutha endlela yokusebenza agxila ekugezweni okubili. Ukususwa kombala okunganele kushiya izinto ezingemuva zibomvu futhi kuholele ekufundeni imiphumela ephozithivu engamanga. Ukususwa kombala ngokweqile kukhipha udayi oyinhloko obanjwe ngokubuthaka futhi kube nomthelela emiphumeleni enegethivu engamanga. Udayi ophikisanayo ofakwe ngokweqile ufihla ama-bacilli ambalwa kokuluhlaza okwesibhakabhaka. Amasampula asabalele awugqinsi, izinto zokuhlola ezakhe inzika, namanzi okugeza angcolisiwe kunezela okwako ukubonakala okudukisayo.

Ikhono lokuthola amagciwane likhawulelwa ubuningi bawo, kungakhathaliseki ukuthi indlela yokusebenza yenziwe kahle kangakanani. Ukuhlolwa okuvamile kwesampula esisabalele ngesibonakhulu kudinga cishe ama-bacilli angu-5,000 kuya kwangu-10,000 ngemililitha yesikhwehlela, futhi ukubuyekezwa okudala kobufakazi kwe-CDC kwabeka ikhono lokuthola ngesampula esisabalele cishe kumaphesenti angu-45 kuya kwangu-80 phakathi kwezimo zofuba lwamaphaphu eziqinisekiswe ngokukhuliswa kwamagciwane. Ngakho isampula esisabalele enomphumela onegethivu ayikaze ikhiphe ngokuphelele ukuthi kungaba khona ufuba. Isiqondiso samanje se-CDC sigcina ukuhlolwa ngokukhulisa amagciwane kuhambisana nayo yonke isampula esisabalele, futhi sincoma ukuhlolwa kokwandiswa kwe-nucleic acid okungenani esampuleni eyodwa yomgudu wokuphefumula lapho kusolwa ufuba lwamaphaphu futhi umphumela ungashintsha indlela yokuphatha isiguli.

Umsebenzi owenziwayo udinga izinqubo ezigunyaziwe

Izincazelo zokufundisa, kuhlanganise nalesi sihloko, azigunyazi umsebenzi wokuxilonga iziguli. Emsebenzini waselabhorethri, landela inqubo ejwayelekile yokusebenza yelabhorethri eqinisekisiwe futhi ehambisana nemibhalo yamanje ye-CDC, APHL, noma CLSI; umhlahlandlela we-CDC wokudaya okumelana ne-asidi ogcinwe enqolobaneni yayo ye-Stacks ubonisa ukuthi lezi zinqubo zizicacisa kanjani izinto zokuhlola nokufundwa kwemiphumela. Ukusebenza ngamasampula ofuba kudinga ukuvinjelwa kokusabalala kwezinto eziphilayo: amasampula asabalele alungiswa ekhabetheni lokuphepha kwezinto eziphilayo ngoba ukuwalungisa kudala izinhlayiyana ezintanta emoyeni namaconsi afafazekayo, futhi isiqondiso se-CDC sokuphepha kwelabhorethri sithi ngisho namasampula asabalele aqiniswe ngokushisa angase abe nama-bacilli ofuba asaphila futhi kusafanele aphathwe njengangcolisiwe. Umbhalo we-CDC Public Health Image Library wesampula esisabalele edaywe nge-carbol fuchsin ne-methylene blue yahlolwa ukuze kutholakale i-M. leprae uyisikhumbuzo esiwusizo sokusetshenziswa kwale ndlela nangaphandle kofuba: ukuhlukana okufanayo kokubomvu phezu kokuluhlaza okwesibhakabhaka kusiza nomaphi lapho kufunwa khona izinto eziphilayo ezimelana nokususwa kombala nge-asidi, futhi ukufunda ngokucophelela kungatholakali lutho nakho kuwumphumela okufanele ubhalwe.

Ukufunda ingilazi yesibonakhulu esiqediwe

Udayi ophikisanayo ubonisa ukubaluleka kwawo ngesikhathi sokuhlola kokuqala. Umhloli ngesibonakhulu ohlola ngokukhulisa okuphansi uqala asebenzise ingemuva eliluhlaza okwesibhakabhaka ukuze ahlole ikhwalithi yesampula esisabalele: ukusabalala kwamaseli ngokulinganayo, amaseli omzimba ophethe amagciwane angonakele okusiza ekucijiseni isithombe, nokungabikho kwenzika eningi. Izinto ezibomvu ezingase zibe ama-bacilli zibe sezidonsa iso ngoba akukho okunye okubomvu engilazini yesibonakhulu. Ukuqinisekisa kwenzeka kusetshenziswa ilensi ecwiliswa emafutheni, lapho ama-bacilli angempela ebonisa izinduku ezilindelekile ezizacile, kwesinye isikhathi ezigoqeke kancane noma ezinjengobuhlalu, kunokuba zibe izinsalela zikadayi ezifanayo.

Ukudaywa kwengemuva kuphinde kunqume amagama asetshenziswa embikweni. Ingilazi yesibonakhulu enama-bacilli abomvu ibikwa ngokuthi kubonakale ama-bacilli amelana nokususwa kombala nge-asidi, ngokuvamile ihambisane nebanga elilinganisa ubuningi ngokwesikali selabhorethri. Ingilazi engenazo izinto eziphilayo ezibomvu ibikwa ngokuthi awekho ama-bacilli amelana nokususwa kombala nge-asidi abonakele, hhayi ngokuthi ayinalo ufuba. Lo mehluko ubalulekile ngoba indawo eluhlaza okwesibhakabhaka ifakazela ukuthi ukudaya kusebenzile, amaseli athathe udayi ophikisanayo futhi izimo zokubuka bezanele, kodwa ayisho lutho ngezinto eziphilayo ezingaphansi komkhawulo wokutholwa. Indawo eluhlaza okwesibhakabhaka engenazo izinduku ezibomvu iwumphumela ovumelekile ophathelene nengilazi yesibonakhulu, hhayi isinqumo ngesiguli.

Khumbula ukuhlukaniswa kwemisebenzi, bese imibiko iba lula ukuyibhala. I-carbol fuchsin iveza izinto ezingase zibe ama-bacilli. I-asidi notshwala kwenza ukuhlolwa. I-methylene blue yenza impendulo ifundeke kuphela: okubomvu kugcine udayi naphezu kwe-asidi, okuluhlaza okwesibhakabhaka kuwulahlekele. Konke okunye, uhlobo lwamagciwane, ubuningi bawo, nencazelo ephathelene nesiguli, kutholakala emsebenzini ongaphezu kokuhlola ingilazi yesibonakhulu.