Impamvu ubururu bwa metilene bukoreshwa nk'irangi ryunganira mu isiga rigaragaza kwihanganira aside

Umutekinisiye asuzuma icyitegererezo cy'igikororwa cyasizwe ku kirahuri nyuma y'intambwe zose zo kugisiga amarangi. Bagiteri zitukura zimeze nk'uduti zigaragara neza mu buso bw'ubururu, maze raporo ikavuga ko habonetse basili zihanganira aside. Bukeye, ikindi kirahuri kigaragaza ubuso bwose bw'ubururu butarimo uduti dutukura, maze raporo ikavuga ko nta gituntu cyabonetse. Imwe muri izi raporo ivuga ibirenze ibyo ikirahuri gishobora kwemeza, kandi si iya mbere.

Uku kwitiranya guturuka ku buryo igitekerezo cyumvikanywe. Umutekinisiye yafashe irangi ry'ubururu nk'igice cy'ubutabire bwo kuvumbura bagiteri, nk'aho ubururu bwa metilene bwafashaga kubona mikobagiteri. Ntabwo bwabikoze. Igihe ubururu bwa metilene bugera ku kirahuri, ikizamini kiba cyararangiye. Iri rangi risiga gusa ibiri inyuma kugira ngo igisubizo gisomeke. Iyi nyandiko isobanura akamaro k'ubururu bwa metilene mu isiga rigaragaza kwihanganira aside, impamvu hakoreshwa ubururu aho gukoresha irindi bara, n'icyo umutuku ugaragara ku bururu ukwemerera kwemeza.

Igisubizo kigufi

Isiga rigaragaza kwihanganira aside ni ikizamini gitandukanya uturemangingo gishingiye ku bitekerezo bitatu. Carbol fuchsin, irangi ry'ibanze, yinjizwa ku ngufu muri buri karemangingo, harimo na mikobagiteri zifite igifuniko kimeze nk'igishashara. Aside-alukolo, ikuraho irangi, ikura irangi ritukura mu turemangingo dusanzwe ariko ntishobore kurikura mu twihanganira aside. Hanyuma ubururu bwa metilene, irangi ryunganira, bugasiga ibyo ikuraho irangi yari yambuye ibara byose, bityo uturemangingo tutihanganira aside, utw'uwatanze icyitegererezo n'ibisigazwa bikagaragara ari ubururu, naho basili zihanganira aside zigakomeza kuba umutuku. Irangi ryunganira ritanga itandukaniro ry'amabara. Nta ruhare rifite mu butabire butuma akaremangingo kihanganira aside.

Icyo buri ntambwe ikora

Igishushanyo gikurikira cyerekana uko imirimo igabanywa. Gisome uva ibumoso ugana iburyo nk'uko intambwe zikurikirana, kandi usome umurongo wo hasi nk'icyibutsa ko buri tsinda ry'uburyo bwo gusiga amarangi risubiza ikibazo gitandukanye.

Kwinjira kw'irangi ni ko kubanza. Mikobagiteri zibuza amarangi asanzwe avanze n'amazi kwinjira kubera ko igifuniko cyazo kitavangwa n'amazi ku rugero rudasanzwe, bityo amabwiriza y'ubu buryo agatuma irangi ryinjira ku ngufu. Mu buryo bwa Ziehl-Neelsen, gushyushya kugeza habaye umwuka bituma carbol fuchsin irimo phenol yinjira mu rukuta rumeze nk'igishashara. Mu buryo bukonje bwa Kinyoun, ubushyuhe busimburwa n'uruvange rw'irangi ry'ibanze rukomeye cyane kurushaho, kandi uburyo bumwe bwahinduwe budakoresha ubushyuhe bwongeramo ibinyabutabire bigabanya ubukana bwo ku buso. Amabwiriza ya American Society for Microbiology ku isiga rigaragaza kwihanganira aside ashyira izi mvange zombi iruhande kugira ngo zigereranywe, kandi itandukaniro ry'ubwinshi bw'irangi riragaragara cyane: hafi 0.3 g ya fukusine y'ibanze kuri 100 mL mu buryo bukoresha ubushyuhe, ugereranyije na hafi 4 g mu buryo bukonje.

Gutandukanya uturemangingo ni byo bikurikiraho. Aside-alukolo, akenshi igizwe na 3 mL za aside idorokilorike ikomeye muri 97 mL za etanolu ya 95 ku ijana, yoza irangi ritukura ikarikura muri buri karemangingo gafite urukuta rudatanga ubwirinzi bwihariye. Uturemangingo dufite igifuniko cya mikobagiteri kigizwe n'ibinure kandi kitangiritse tugumana irangi. Iyi ni yo ntambwe ubu buryo bwose bushingiyeho, kandi ni na yo yibasirwa cyane n'uburyo imirimo ikorwa, nk'uko bisobanurwa hepfo.

Itandukaniro ry'amabara riza ku musozo. Ikirahuri kiba noneho kiriho uturemangingo dutukura twihanganira aside ku buso bw'inyuma bugizwe n'uturemangingo twambuwe irangi hafi yo kubura ibara burundu. Ubururu bwa metilene bwa 0.3 ku ijana w/v butuma ibyo biri inyuma byongera kugaragara, mu ibara ritandukanye cyane n'umutuku. Gusiga irangi ryunganira bimara igihe gito, ubusanzwe amasegonda 20 kugeza kuri 30 mu buryo bukoresha ubushyuhe, kandi bishobora kumara iminota mike mu buryo bumwe bukonje bwahinduwe; igihe gishingira ku mabwiriza y'uburyo bukoreshwa, si ku miterere bwite y'irangi.

Impamvu y'iri rangi, n'impamvu y'ubururu

Ubururu bwa metilene ni irangi ry'ibanze rifite iyoni nziza, bivuze ko iyoni yaryo ifite ibara itwaye amashanyarazi meza kandi ikifatisha ku buso bufite amashanyarazi mabi bwa bagiteri zambuwe irangi, uturemangingo tw'uwatanze icyitegererezo n'ibisigazwa. OpenStax Microbiology isobanura iyi mikoranire y'amashanyarazi nk'ihame rusange amarangi y'ibanze ashingiyeho. Ariko ubururu bwa metilene nta mwihariko w'ubutabire bufite hano. Crystal violet na safranin na byo ni amarangi y'ibanze. Guhitamo bishingira ku buryo amabara agaragara, si ku butabire.

Uduti dutukura dukeneye ko ibiri inyuma bigira ibara ritandukanye na two, kandi ubururu butandukanye n'umutuku mu buryo bufasha cyane mu gukoresha mikorosikopi ifite ubuso bumurikiwe. Safranin yaba irangi ryunganira ridakwiriye kubera ko ibara ryayo rya nyuma ry'iroza-umutuku ryegereye cyane irya carbol fuchsin yasigaye mu turemangingo, bityo rigasibanganya itandukaniro ubu buryo bwari bwashyizeho. Crystal violet yatuma ibiri inyuma biba ibara ry'umuyugubwe wijimye rirushanwa na basili nto zitukura mu gukurura amaso. Ikimenyetso gikomeye kurusha ibindi cyerekana ko umurimo ari ugutanga itandukaniro ry'amabara aho kuba uw'ubutabire ni uko irangi rishobora gusimburwa: amarangi ryunganira y'icyatsi nka malachite green na brilliant green akoreshwa mu mabwiriza amwe y'ubuvuzi bw'amatungo no mu buryo bwahinduwe, kandi ikizamini kigakomeza gukora, kubera ko ibara iryo ari ryo ryose ritandukanye neza rikora uwo murimo.

Ibi bitanga imyanzuro ibiri ifatika. Mbere na mbere, uruvange rusanzwe ntirutegeka guhindura pH; ntukitiranye uyu muti woroshye uvanze n'amazi wa 0.3 ku ijana n'imvange za alikali nka ubururu bwa metilene bwa Loeffler, zishingiye ku gitekerezo gitandukanye cyo gusiga amarangi. Icya kabiri, gukoresha irangi birenze urugero birangiza. Inyandiko zerekeye iyigwa ry'uturemangingo ziraburira by'umwihariko ko ubururu bwinshi cyane mu biri inyuma bushobora guhisha basili nke, bityo gusiga ikirahuri gitwikiriwe n'irangi igihe kirenze icyateganyijwe mu mabwiriza bigabanya uburyo gisomeka nta nyungu bitanze.

Ubutabire bw'urukuta butuma ikizamini gishoboka

Impamvu aside-alukolo ishobora gutandukanya uturemangingo iri mu gifuniko cya mikobagiteri. Igice cy'ingenzi cyubaka icyo gifuniko ni urusobe rwa mycolyl-arabinogalactan-peptidoglycan: peptidoglycan ihujwe na arabinogalactan, ifite ku mpera zayo z'inyuma aside mikolike zifite iminyururu miremire cyane, bigakora igihu cyo hanze cya mikobagiteri gikungahaye ku binure kandi kitanyurwamo ku rugero rudasanzwe. Isesengura ry'igihu cyo hanze cya mikobagiteri risobanura iyi myubakire mu buryo burambuye. Iyi nzitizi ikora mu byerekezo byombi. Ibuza irangi kwinjira, ari yo mpamvu kwinjiza irangi bisaba ubushyuhe cyangwa irangi ry'ibanze ryongerewe imbaraga, kandi ikabuza irangi gukurwamo, ari yo mpamvu aside-alukolo idashobora gukuramo carbol fuchsin iyo yamaze kwinjira. Bagiteri zisanzwe ntizifite iki gifuniko kandi zitakaza irangi. Bityo kwihanganira aside byumvikana neza nk'ingaruka y'inzitizi y'ibinure, aho kuba gufatana kwa fukusine na aside mikolike binyuze mu butabire.

Ubu butabire busobanura kandi ikibazo gikunze gutera urujijo: mikobagiteri ntizisigwa neza kandi ntizitanga ibisubizo byizewe mu buryo bwa Gram, rimwe na rimwe zikagaragara nk'imirongo y'uduce tumeze nk'amasaro dufite ibara ridakomeye cyangwa nk'ibishusho bitasizwe irangi. Urukuta rwazo rutavangwa n'amazi rugabanya uko crystal violet yinjira. Isiga rigaragaza kwihanganira aside si isiga rya Gram ryahinduwe. Ni ihame ritandukanye ryo gutandukanya uturemangingo rishingiye ku wundi mwihariko w'urukuta, nk'uko igereranya ry'isiga ryoroshye n'isiga rya Gram ribigaragaza ku bundi buryo bubiri.

Uburyo butatu, ibibazo bitatu

Ku itsinda ryagutse ry'uburyo bwo gusiga bagiteri hakoreshejwe irangi rimwe, reba igitabo cy'amashusho y'uburyo bwo gusiga amarangi mu gukoresha mikorosikopi.

Icyo umutuku ugaragara ku bururu udashobora kwemeza

Uduti dutukura dusobanura ko izo bagiteri zagumanye carbol fuchsin nubwo zakorewe aside-alukolo. Icyo cyabonetse gihuye na Mycobacterium tuberculosis, ariko amabwiriza ya CDC yo gusuzuma igituntu avuga yeruye ko izindi mikobagiteri na zo zihanganira aside, bityo kumenya ubwoko bwihariye bigasaba kuzikuza muri laboratwari no gupima aside nyukileyike, aho gushingira ku ibara.

Amakosa mu mikorere akunze kubaho mu ntambwe ebyiri zo koza. Kudakuraho irangi bihagije bituma ibiri inyuma biguma ari umutuku, bikaba byatuma hasomwa igisubizo cyemeza kitari cyo. Kurikuraho birenze urugero bikuraho irangi ry'ibanze ryari ryafashwe buhoro, bikagira uruhare mu bisubizo bihakana bitari byo. Irangi ryunganira ryakoreshejwe birenze urugero rihisha basili nke mu bururu. Icyitegererezo cyasizwe ku kirahuri ari kinini cyane, imiti y'ikizamini yibumbiye hamwe n'amazi yanduye yo kwoza na byo byongeramo ibigaragara bitari iby'icyitegererezo ubwacyo.

Ubushobozi bwo kubona bagiteri bugengwa n'ubwinshi bwazo, nubwo tekinike yaba yakozwe neza gute. Gusuzuma icyitegererezo cyasizwe ku kirahuri hakoreshejwe mikorosikopi mu buryo busanzwe bisaba hafi basili 5,000 kugeza kuri 10,000 muri buri mililitiro y'igikororwa, kandi amasesengura ya kera ya CDC ku bimenyetso yashyiraga ubushobozi bw'ubu buryo bwo kubona bagiteri hagati ya 45 na 80 ku ijana mu barwayi b'igituntu cy'ibihaha cyemejwe no kuzikuriza muri laboratwari. Bityo igisubizo cy'icyitegererezo cyasizwe ku kirahuri kitagaragaza bagiteri ntigishobora na rimwe guhakana igituntu. Amabwiriza ya CDC y'ubu asaba gukomeza gukuriza bagiteri muri laboratwari ku ruhande rwa buri cyitegererezo gisuzumwe ku kirahuri, kandi agasaba ikizamini cyo kongera aside nyukileyike nibura ku cyitegererezo kimwe cyo mu myanya y'ubuhumekero igihe hakekwa igituntu cy'ibihaha kandi igisubizo gishobora guhindura uko umurwayi yitabwaho.

Imirimo ifatika igomba kugengwa n'amabwiriza yemewe

Ibisobanuro bigenewe kwigisha, harimo n'iyi nyandiko, ntibitanga uburenganzira bwo gukora imirimo yo gusuzuma abarwayi. Mu mirimo yo muri laboratwari, kurikiza uburyo busanzwe bwemejwe na laboratwari, buhuye n'inyandiko za CDC, APHL cyangwa CLSI zigezweho; inyandiko ngenderwaho ya CDC ku isiga rigaragaza kwihanganira aside ibitswe mu bubiko bwayo bwa Stacks yerekana uko ayo mabwiriza agaragaza imiti y'ikizamini n'uko ibisubizo bisomwa. Gukora ku byitegererezo by'igituntu bisaba ingamba zo gukumira ikwirakwira: ibyitegererezo bisigwa ku birahuri bitegurirwa mu kabati kabugenewe k'umutekano w'ibinyabuzima kubera ko kubitegura bitera udutonyanga tuguruka mu mwuka no kumeneka kw'amatonyanga; amabwiriza ya CDC y'umutekano muri laboratwari avuga ko n'ibyitegererezo byafashishijwe ku birahuri hakoreshejwe ubushyuhe bishobora kuba birimo basili z'igituntu zikiri nzima kandi bigomba gukomeza gufatwa nk'ibihumanye. Inyandiko yo mu Isomero ry'Amashusho y'Ubuzima Rusange rya CDC ku cyitegererezo cyasizwe carbol-fuchsin n'ubururu bwa metilene, cyasuzumwe hashakwa M. leprae yibutsa neza ko ubu buryo bukoreshwa no ku bindi bitari igituntu: iri tandukaniro ry'umutuku ku bururu rikoreshwa aho hose hashakishwa ibinyabuzima byihanganira aside, kandi igisubizo gihakana cyasomwe witonze na cyo ni igisubizo gikwiye kwandikwa.

Gusoma ikirahuri cyarangije gutegurwa

Irangi ryunganira rigaragaza akamaro karyo mu isuzuma rya mbere. Usuzuma akoresheje mikorosikopi, abanje kureba ku rugero ruto rwo gutubura ishusho, akoresha ubururu bw'ibiri inyuma kugira ngo amenye ubuziranenge bw'icyitegererezo cyasizwe: uturemangingo tugomba kuba dutatanye ku buryo bungana, uturemangingo tw'uwatanze icyitegererezo tutangiritse tugafasha gutyaza ishusho, kandi ntihabe imiti myinshi yibumbiye hamwe. Hanyuma ibisa na basili bitukura bikurura amaso kubera ko nta kindi kintu ku kirahuri gitukura. Kwemeza bikorwa hakoreshejwe uburyo bwo kwibiza mu mavuta, aho basili nyazo zigaragaza uduti duto twitezwe, rimwe na rimwe tugoramye buhoro cyangwa tumeze nk'amasaro, aho kuba ibisigazwa by'irangi bifite ishusho imwe.

Gusiga ibiri inyuma irangi na byo bigena amagambo akoreshwa muri raporo. Ikirahuri kiriho basili zitukura gitangwaho raporo ivuga ko habonetse basili zihanganira aside, akenshi hakongerwaho urwego rugereranya ubwinshi bwazo hakurikijwe igipimo cya laboratwari. Ikirahuri kitarimo ibinyabuzima bitukura gitangwaho raporo ivuga ko nta basili zihanganira aside zabonetse, ntikigomba na rimwe kuvugwaho ko nta gituntu cyabonetse. Iri tandukaniro ni ingenzi kubera ko ubuso bw'ubururu bwemeza ko isiga ryagenze neza, uturemangingo twafashe irangi ryunganira kandi uburyo bwo kureba bwari bukwiye, ariko ntibugire icyo buvuga ku binyabuzima biri munsi y'urugero rushobora kuboneka. Ubuso bw'ubururu butarimo ibigaragara ni igisubizo gifite agaciro ku byerekeye ikirahuri, si umwanzuro ku murwayi.

Wibuke uko imirimo igabanywa, maze kwandika raporo bikorohe. Carbol fuchsin igaragaza ibishobora kuba ari byo bishakishwa. Aside-alukolo ikora ikizamini. Ubururu bwa metilene butuma gusa igisubizo gisomeka: umutuku wagumanye irangi nubwo hakoreshejwe aside, ubururu bwararitakaje. Ibindi byose, ari byo ubwoko bwihariye, ubwinshi n'igisobanuro ku buzima bw'umurwayi, bituruka ku mirimo irenze kureba ikirahuri gusa.