Nooc cusub oo ah Gram-negative, aerobic, cusbo-dulqaad leh, firfircoon, ul qaabaysan, iyo bakteeriyada ugaarsada ASxL5T ayaa laga soocay balli saxaro lo'aad ah oo ku taal Nottinghamshire, England, waxayna u isticmaashay Campylobacter sidii ay ugaarsan lahayd. Ka dib, noocyada kale ee Campylobacter iyo xubnaha qoyska Enterobacteriaceae ayaa la ogaaday inay yihiin ugaadh. Ka dib dhaqanka hoose ee aan lahayn unugyada martida loo yahay, koritaanka aseptic daciifka ah ayaa lagu gaaray faleebo Wadnaha Maskaxda Agar. Xaaladaha korriinka ugu wanaagsani waa 37 °C iyo pH waa 7. Mikroskoobyada elektarooniga ah ee gudbinta ayaa shaaca ka qaaday qaar ka mid ah sifooyinka qaab-dhismeedka aan caadiga ahayn ee la xiriira helitaanka ugaadha. Falanqaynta Phylogenetic iyadoo la adeegsanayo taxanaha hiddo-wadaha 16S rRNA waxay muujisay in go'doominta ay la xiriirto xubin ka tirsan qoyska Marine Spirulina, laakiin aan si cad loogu kala saari karin xubin ka mid ah cirrid kasta oo la yaqaan. Isku xigxiga guud-genome-ga ee ASxL5T ayaa xaqiijiyay xidhiidhka xubnaha spirochetes badda. Baadhitaan xog ururin ah ayaa daaha ka qaaday in dhowr ASxL5Ts ay wadaagaan 16S rRNA taxane hiddo-wade ah oo ay la socdaan bakteeriyayaal badan oo aan dhaqan lahayn oo ka yimid badda, oogada dhulka iyo biyaha dhulka hoostiisa. Waxaan soo jeedinaynaa in cadaadiska ASxL5T uu matalo nooc cusub oo hidde cusub ah. Waxaan ku talinaynaa magaca Venatorbacter cucullus gen. November, sp. Bishii Nofembar, ASxL5T waxaa loo adeegsaday nooca nooca.
Bakteeriyada ugaadhsiga ahi waa bakteeriya muujisa awoodda ay ku ugaarsato oo ay disho bakteeriyada kale ee nool si ay u hesho agabka biosynthetic iyo tamar. Tani waxay ka duwan tahay ka soo kabashada guud ee nafaqeeyayaalka yar yar ee dhintay, sidoo kale waxay ka duwan tahay isdhexgalka parasitic, taas oo bakteeriyada ay la sameysato xiriir dhow iyada oo aan la dilin. Bakteeriyada wax ugaadhsiga ahi waxay soo saartay wareegyo nololeed oo kala duwan si ay uga faa'iidaysato ilo cunto oo badan meelaha laga helo (sida deegaanka badda). Waa koox canshuur ahaan kala duwan, kuwaas oo kaliya ay ku xiran yihiin meertodooda nololeed ee jeermiska ee gaarka ah1. Tusaalooyinka bakteeriyada ugaadhsiga ah ayaa laga helay dhowr phyla oo kala duwan, oo ay ku jiraan: Proteobacteria, Bacteroides, iyo Chlorella.3. Si kastaba ha ahaatee, bakteeriyada ugaadhsiga aadka loo bartay waa Bdellovibrio iyo Bdellovibrio-iyo noole la mid ah (BALOs4). Bakteeriyada ugaadhsiga ahi waa il rajo leh oo laga helo xeryahooda cusub ee bayooloji firfircoon iyo wakiilada bakteeriyada5.
Bakteeriyada ugaadhsiga ah ayaa la aaminsan yahay inay kor u qaaddo kala duwanaanta microbial waxayna saameyn togan ku leedahay caafimaadka nidaamka deegaanka, wax soo saarka iyo xasilloonida6. Inkasta oo sifooyinkan wanaagsani ay jiraan, waxaa jira daraasado yar oo ku saabsan bakteeriyada cusub ee ugaadhsiga ah sababtoo ah dhibaatada dhaqanka bakteeriyada iyo baahida loo qabo in si taxadar leh loo ilaaliyo isdhexgalka unugyada si loo fahmo wareegyada nololeed ee adag. Macluumaadkan ma fududa in laga helo falanqaynta kombayutarka.
Xilliga sii kordhaya ee iska caabbinta antimicrobial, xeelado cusub oo lagu beegsanayo cudurada bakteeriyada ayaa la daraaseeyay, sida isticmaalka bakteeriyada iyo bakteeriyada ugaadhsiga7,8. Bakteeriyada ASxL5T waa la go'doomiyay sanadka 2019 iyadoo la adeegsanayo tignoolajiyada go'doominta fage ee saxarada lo'da ee laga soo ururiyay Xarunta Caanaha ee Jaamacadda Nottingham, Nottinghamshire. Ujeedada baaritaanku waa in la go'doomiyo noolaha awooda u leh inay yihiin wakiilada xakamaynta noolaha. Campylobacter hyointestinalis waa cudur-sidaha zoonotic, kaas oo si isa soo taraya ula xiriira cudurrada mindhicirka aadanaha10. Waa meel walba oo serum ah waxaana loo isticmaalaa martigaliyaha bartilmaameedka ah.
Bakteeriyada ASxL5T waxay ka go'day jelly lo'da sababtoo ah waxaa la arkay in taarikada ay ku samaysay cawska C. hyointestinalis ay la mid yihiin kuwa ay soo saarto bakteeriyada. Tani waa helitaan lama filaan ah, sababtoo ah qayb ka mid ah habka go'doominta phage waxay ku lug leedahay shaandhaynta 0.2 µm filter, kaas oo loogu talagalay in laga saaro unugyada bakteeriyada. Baadhitaan aan yarayn oo lagu sameeyay walxaha laga soo saaray huurada ayaa daaha ka qaaday in bakteeriyada yar ee garaam-negative-ka ah ee usha u samaysan aanay ururin polyhydroxybutyrate (PHB). Dhaqanka aseptic ee ka madax banaan unugyada ugaadhsiga waxaa lagu gartaa dhexdhexaad adag oo qani ah (sida faleebo wadnaha maskaxda agar (BHI) iyo agar dhiiga (BA)), koritaankeeduna waa daciif. Waxaa la helaa ka dib subculture oo leh inoculum culus oo hagaagsan. Waxay si isku mid ah u kortaa hoos microaerobic (7% v/v oxygen) iyo xaaladaha ogsijiinta ee jawiga, laakiin kuma jirto jawi anaerobic ah. 72 saacadood ka dib, dhexroorka gumaysigu wuxuu ahaa mid aad u yar, wuxuu gaadhay 2 mm, wuxuuna ahaa beige, translucent, wareegsan, convex iyo dhalaalaya. Tijaabada caadiga ah ee kiimikaad baayooloji waa caqabad sababtoo ah ASxL5T laguma dhaqmi karo si la isku halleyn karo warbaahinta dareeraha ah, taas oo soo jeedinaysa inay ku tiirsan tahay wareegga nolosha ee kakan ee samaynta biofilm. Si kastaba ha ahaatee, joojinta saxanka ayaa muujisay in ASxL5T ay tahay aerobic, oo u fiican oxidase iyo catalase, waxayna u dulqaadan kartaa 5% NaCl. ASxL5T waxay u adkaysataa 10 µg streptomycin, laakiin waxay u nugul tahay dhammaan antibiyootiga kale ee la tijaabiyey. Unugyada bakteeriyada ASxL5T waxaa baadhay TEM (Jaantuska 1). Marka la koray iyada oo aan lahayn unugyo ugaadhsi ah oo ku yaal BA, unugyada ASxL5T waa Campylobacter yar, oo leh celcelis ahaan dhererka 1.63 μm (± 0.4), ballac ah 0.37 μm (± 0.08), iyo hal dheer oo dheer (ilaa 5 μm). Calan galmo. Qiyaastii 1.6% unugyadu waxay u muuqdaan inay leeyihiin ballac ka yar 0.2 μm, taas oo u oggolaan doonta inay dhex maraan qalabka shaandhada. Kordhinta qaabdhismeed aan caadi ahayn ayaa lagu arkay korka unugyada qaar, oo la mid ah caddaaladda (Latin cucullus) (eeg fallaadhaha 1D, E, G). Tani waxay u muuqataa inay ka kooban tahay xuubka sare ee xad-dhaafka ah, taas oo laga yaabo inay sabab u tahay hoos u dhaca degdega ah ee xajmiga baqshadda periplasmic, halka xuubka dibedda uu weli yahay mid sax ah, oo muujinaya muuqaal "dabcan". Ku-dhaqanka ASxL5T ee maqnaanshaha nafaqooyinka (PBS) muddo dheer 4 ° C waxay keentay inta badan (laakiin ma aha dhammaan) unugyada muujinaya qaab-dhismeedka coccal (Jaantus 1C). Marka ASxL5T ula korto Campylobacter jejuni sida ugaarsi ahaan 48 saacadood, celceliska cabbirka unuggu aad ayuu uga dheer yahay oo wuu ka cidhiidhsan yahay unugyada koray marti-geliyaha la'aan (Shaxda 1 iyo Jaantuska 1E). Taas bedelkeeda, marka ASxL5T ay u koraan E. coli sida ugaadhsiga 48 saacadood, celceliska cabbirka unuggu wuu ka dheer yahay oo wuu ka badan yahay marka uu korayo ugaadh la'aan (Shaxda 1), dhererka unugguna waa isbeddel, badanaa wuxuu muujiyaa fiilo (Jaantus 1F). Marka lagu beero Campylobacter jejuni ama E. coli sidii ugaadh 48 saacadood ah, unugyada ASxL5T ma muujin wax calan ah haba yaraatee. Shaxda 1 waxay soo koobaysaa indha-indhaynta isbeddelada cabbirka unugga ee ku salaysan joogitaanka, maqnaanshaha, iyo nooca ugaadhsiga ee ASxL5T.
Muujinta TEM ee ASx5LT: (A) ASx5LT waxay muujinaysaa karbaash dheer; (B) batari ASx5LT caadiga ah; (C) cocci unugyada ASx5LT ka dib markii muddo dheer la kariyey oo aan nafaqo lahayn; (D) koox ka mid ah unugyada ASx5LT waxay muujinayaan aan caadi ahayn (E) kooxda unugyada ASx5LT ee lagu dhex beeray Campylobacter prey waxay muujisay dhererka unugyada oo kordhay marka la barbardhigo kuwa aan lahayn korriinka ugaadhsiga (D) ayaa sidoo kale muujiyay qaab-dhismeedka apical; (F) Calanka Filamentous Large, unugyada ASx5LT, ka dib marka lagu beero E. coli ugaadha; (G) Hal unug oo ASx5LT ah ka dib marka lagu beero E. coli, oo muujinaya qaab-dhismeed sare oo aan caadi ahayn. Baarku waxa uu ka dhigan yahay 1 μm.
Go'aaminta taxanaha hiddo-wadaha 16S rRNA (lambarka gelitaanka MT636545.1) waxay awood u siinaysaa raadinta xogta xogta si loo sameeyo taxane la mid ah kuwa fasalka Gammaproteobacteria, waxayna ugu dhow yihiin bakteeriyada badda ee qoyska spirillum badda (Jaantuska 2), waxayna xubno ka yihiin Thalassolituus genus. Qoyska ugu dhow ee Marine Bacillus. Taxanaha hidda-wadaha 16S rRNA waxa uu si cad uga duwan yahay bakteeriyada ugaarsiga ah ee ka tirsan qoyska Bdelvibrionaceae (Deltaproteobacteria). Isbarbardhigga labada dhinac ee B. bacteriovorus HD100T (nooca nooca, DSM 50701) iyo B. bacteriovorus DM11A waxay ahaayeen 48.4% iyo 47.7%, iyo B. exovorus JSS waxay ahayd 46.7%. Bakteeriyada ASxL5T waxay leedahay 3 koobi oo ah hidda-wadaha 16S rRNA, laba ka mid ah waa isku mid, tan saddexaadna waa 3 saldhig. Laba jeer oo kale oo bakteeriya ugaadha ah (ASx5S iyo ASx5O; 16S rRNA tirooyinka gelitaanka hidda-wadaha waa MT636546.1 iyo MT636547.1, siday u kala horreeyaan) oo leh astaamo muuqaal iyo muuqaal ah oo isku mid ah oo isla meel ah isku mid maaha, laakiin way ka duwan yihiin ASxL5T iyo bakteeriyada aan dhaqan lahayn. taxanaha xogta xogta ayaa lagu soo ururiyay jiilal kale Oceanospirillaceae (Jaantuska 2). Dhammaan taxanaha genome ee ASxL5T waa la go'aamiyay oo lagu kaydiyay xogta NCBI, lambarka gelitaankana waa CP046056. Genome-ka ASxL5T wuxuu ka kooban yahay koromosoom wareeg ah oo ah 2,831,152 bp oo leh saamiga G + C ee 56.1%. Taxanaha genome-ku waxa uu ka kooban yahay 2653 CDS (wadarta), kuwaas oo 2567 ka mid ah la saadaaliyay in ay codeeyan borotiinada, kuwaas oo 1596 ka mid ah loo qoondayn karo hawl qabad (60.2%). Genome-ku waxa uu ka kooban yahay 67 hiddo-sidaha RNA, oo ay ku jiraan 9 rRNAs (3 midkiiba 5S, 16S, iyo 23S) iyo 57 tRNAs. Astaamaha genomic ee ASxL5T ayaa la barbardhigay genome-yada la heli karo ee noocyada qaraabada ugu dhow ee laga aqoonsaday taxanaha hiddo-wadaha 16S rRNA (Shaxda 2). Isticmaal aqoonsiga amino acid (AAI) si aad u barbar dhigto dhammaan genomeyada Thalassolituus ee jira iyo ASxL5T. Sida ugu dhow ee la heli karo (aan dhammaystirnayn) taxanaha genome ee ay go'aamisay AAI waa Thalassolituus sp. C2-1 (ku dar NZ_VNIL01000001). Cadaadiskan ayaa ka go'doonsan qulqulka qoto dheer ee badda Mariana Trench, laakiin hadda ma jiraan wax macluumaad ah oo ku saabsan culeyskan marka la barbardhigo. Marka la barbar dhigo ASxL5T's 2.82 Mb, genome-ka noolaha ayaa ka weyn 4.36 Mb. Celceliska cabbirka genome ee spirochetes badda waxay ku saabsan tahay 4.16 Mb (± 1.1; n = 92 genomes tixraaceed dhammaystiran oo laga baaray https://www.ncbi.nlm.nih.gov/assembly), markaa genome-ka ASxL5T wuxuu la jaan qaadayaa nidaamka Marka loo eego xubnaha kale, aad bay u yar tahay. Adeegso GToTree 1.5.54 si aad u soo saarto genome-ku-salaysan qiyaasta ugu badan ee suurtogalka ah ee geed phylogenetic ah (Jaantuska 3A), addoo isticmaalaya isku xigxiga iyo isku xidhka amino acid taxanaha 172 hal nuqul oo gaar ah Gammaproteobacteria 11,12,13,14,15,16, 17,18. Falanqaynta ayaa muujisay in ay si dhow ula xiriirto Thalassolituus, Bakteeriyada Diyaaradda, iyo Bakteeriyada badda. Si kastaba ha ahaatee, xogtani waxay tilmaamaysaa in ASxL5T ay ka duwan tahay qaraabadeeda ku jira Marine Spirulina iyo xogta isku xigxiga genome-ga waa la heli karaa.
Geedka phylogenetic ee isticmaalaya taxanaha hiddo-wadaha 16S rRNA wuxuu muujinayaa booska ASxL5T, ASxO5, iyo ASxS5 (oo leh xiidmaha) marka loo eego noocyada bakteeriyada aan la beerin iyo kuwa badda ee ku jira Marine Spirulinaceae. Lambarka gelitaanka Genbank wuxuu raacayaa magaca cidhiidhiga ah ee qawladaha. U adeegso ClustalW si aad isugu toosiso taxanaha, oo aad isticmaasho habka suurtogalka ah ee ugu badan iyo qaabka Tamura-Nei si aad u fahanto xidhiidhada phylogenetic, oo aad u sameyso 1000 dib u habayn oo la hagayo barnaamijka MEGA X. Nambarka laanta ayaa tilmaamaya in qiimaha koobiga la hagayay uu ka badan yahay 50%. Escherichia coli U/541T ayaa loo isticmaalay sidii koox ka baxsan.
(A) Geed phylogenetic ah oo ku salaysan genome, oo muujinaya xidhiidhka ka dhexeeya bakteeriyada Spirospiraceae ee bakteeriyada ASxL5T iyo qaraabada dhow, E. coli U 5/41T sida koox ka baxsan. (B) Marka la barbar dhigo T. oleivorans MIL-1T, qaybinta qaybta shaqaynaysa ee hiddo-wadaha ayaa la saadaaliyay iyadoo lagu salaynayo kooxda orthologues (COG) ee borotiinka ASx5LT. Jaantuska bidix wuxuu muujinayaa tirada hidda-sidaha ee qayb kasta oo COG shaqaynaysa ee genome kasta. Jaantuska dhinaca midig wuxuu muujinayaa boqolkiiba genome-yada ku jira koox kasta oo COG ah oo shaqaynaysa. (C) Marka la barbar dhigo T. oleiverans MIL-1T, falanqaynta KEGG oo dhamaystiran
Isticmaalka xogta KEGG si loo baadho hiddo-wadaha ka kooban ee ku jira genome-ka ASxL5T ayaa shaaca ka qaaday dariiqa dheef-shiid kiimikaadka caadiga ah ee gamma aerobic Proteus. ASxL5T waxay ka kooban tahay wadarta 75 hiddo-wadaha loo qoondeeyay borotiinnada dhaqdhaqaaqa bakteeriyada, oo ay ku jiraan hiddo-wadaha ku lug leh chemotaxis, kulanka calanka, iyo nooca IV habka fimbriae. Qeybta ugu dambeysa, 9 ka mid ah 10-kii hidde-sideyaasha ayaa mas'uul ka ah dhaqdhaqaaqa qulqulaya ee noocyada kale ee noolaha. Genome-ka ASxL5T wuxuu ka kooban yahay tetrahydropyrimidine biosynthetic pathway kaas oo ka qaybqaata jawaabta ilaalinta osmotic stress20, sida la filayo halophiles. Genome-ku waxa kale oo uu ka kooban yahay wadooyin badan oo dhamaystiran oo loogu talagalay cofactors iyo vitamins, oo ay ku jiraan dariiqooyinka isku-dhafka riboflavin. Inkasta oo hidda-wadaha alkane 1-monooxygenase (alkB2) uu ku jiro ASxL5T, dariiqa isticmaalka hydrocarbon ma dhammaystirna. Marka la eego taxanaha genome ee ASxL5T, homologues of genes loo aqoonsaday in ay inta badan masuulka ka yihiin xaalufka hydrocarbons ee T. oleiverans MIL-1T21, sida TOL_2658 (alkB) iyo TOL_2772 (khamriga dehydrogenase) ayaa si cad u maqan. Jaantuska 3B wuxuu muujinayaa isbarbardhigga qaybinta hiddo-wadaha ee qaybta COG ee u dhaxaysa ASxL5T iyo saliid saytuun MIL-1T. Guud ahaan, genome-ka ASxL5T yari wuxuu ka kooban yahay hiddo-sideyaal ka yar qayb kasta oo COG ah marka loo eego genome-ka weyn ee la xidhiidha. Marka tirada hiddo-sidaha ee qayb kasta oo shaqeynaya lagu muujiyo sida boqolkiiba genome-ga, kala duwanaanshiyaha ayaa lagu xusay boqolleyda hiddo-wadaha ee tarjumaada, qaab-dhismeedka ribosomal iyo qaybaha biogenesis, iyo qaybaha wax soo saarka tamarta iyo beddelka, taas oo ka kooban ASxL5T weyn. genome Boqolkiiba marka la barbar dhigo isla kooxda ku jirta genome-ga T. oleiverans MIL-1T. Taas bedelkeeda, marka la barbardhigo ASxL5T genome, T. oleivorans MIL-1T waxay leedahay boqolkiiba sare ee hiddo-wadaha ku jira soo-celinta, dib-u-habaynta iyo hagaajinta, iyo qaybaha qoraalka. Waxa xiisaha lihi leh, faraqa ugu weyn ee ka kooban qayb kasta oo shaqaynaya ee labada genome waa tirada hiddo-sideyaasha aan la garanayn ee jooga ASxL5T (Jaantuska 3B). Falanqaynta kobcinta cutubyada KEGG ayaa la sameeyay, halkaas oo cutub kasta oo KEGG uu ka dhigan yahay cutubyo hawleed gacanta lagu qeexay oo loogu talagalay tafsiir iyo tafsiir bayooloji ee xogta taxanaha genome. Isbarbardhigga qaybinta hiddo-wadaha ee dariiqa moduleka KOG oo dhammaystiran ee ASxL5T iyo saytuunka MIL-1T ayaa lagu muujiyey Jaantuska 3C. Falanqayntani waxay muujinaysaa in inkasta oo ASxL5T ay leedahay dariiqa dheef-shiid kiimikaadka baaruudda iyo nitrogen oo dhamaystiran, T. oleiverans MIL-1T ma laha. Taas bedelkeeda, T. oleiverans MIL-1T waxay leedahay cysteine ​​​​iyo methionine dhamaystiran dariiqa dheef-shiid kiimikaadka, laakiin waa mid aan dhamaystirnayn ASxL5T. Sidaa darteed, ASxL5T waxay leedahay qaab sifo ah oo loogu talagalay assimilation sulfate (oo lagu qeexay sida hidde-sideyaasha loo isticmaali karo calaamadaha phenotypic, sida awoodda dheef-shiid kiimikaadka ama cudur-sidaha; https://www.genome.jp/kegg/module.html) gudaha T T Oleiverans MIL-1T. Isbarbardhigga nuxurka hidda-sidaha ASxL5T iyo liiska hiddo-wadaha ee soo jeedinaya qaab nololeedka ugaarsiga waa mid aan natiijo lahayn. Inkasta oo hidda-wadaha waaL ee ku dhejinaya ligase la xidhiidha O antigen polysaccharide ilaa xuddunta ay ku jirto ASxL5T genome (laakiin waxay ku badan tahay bakteeriyada Gram-negative badan), tryptophan 2,3-dioxygenase (TDO) Hidde-yaasha waxaa ku jiri kara 60 amino gobollada aysiidhka sida caadiga ah laga helo bakteeriyada ugaadhsiga ah ee aan joogin. Ma jiraan hidde-sideyaal kale oo sifo ugaadhsi ah oo ku jira genome-ka ASxL5T, oo ay ku jiraan kuwa ensaymes-ka dhigaya ee ku lug leh biosynthesis isoprenoid ee waddada mevalonate. Ogsoonow in aysan jirin hiddo-wadaha hab-nololeedka ah ee gntR ee kooxda ugaarsada ee la baaray, laakiin saddex hidde-side gntR ayaa lagu aqoonsan karaa ASxL5T.
Astaamaha phenotypic ee ASxL5T waxaa lagu soo koobay Jadwalka 3 waxaana la barbardhigay astaamaha phenotypic ee genera 23, 24, 25, 26, iyo 27 ee lagu soo sheegay suugaanta. Go'doominta T. marinus, T. olevorans, B. sanyensis, iyo Oceanobacter kriegii waa firfircoon, milix u dulqaadan, oxidase-positive usha qaabaysan, laakiin ma laha ku dhawaad ​​sifooyin kale oo phenotypic la ASxL5T. Celceliska pH ee baddu waa 8.1 kriegii. ASxL5T waxay ku habboon tahay tirada pH ee weyn (4-9) ee noocyada aan badda ahayn. Astaamaha phenotypic ee Thalassoltuus sp. C2-1. Lama garanayo. Heerkulka korriinka ee ASxL5T guud ahaan wuu ka ballaaran yahay kan noocyada badda (4-42 °C), inkastoo qaar laakiin aan dhammaan T. marinus go'doomin ay yihiin kuwa u adkeysan kara kulaylka. Awood la'aanta in ay ku koraan ASxL5T ee warbaahinta maraq ayaa ka hortagtay sifooyin kale oo ifafaale ah. Isticmaal API 20E si aad u tijaabiso walxaha laga soo xoqay saxanka BA, ONPG, arginine dihydrolase, lysine decarboxylase, ornithine decarboxylase, ka faa'iidaysiga citrate, urease, tryptophan deaminase, gelatin hydrolysis Enzyme, natiijooyinka imtixaanku waxay ahaayeen dhammaan taban, laakiin ma laha indole, acetoin iyo H2S ayaa la soo saaray. Karbohaydraytyada aan la khamiirin waxaa ka mid ah: glucose, mannose, inositol, sorbitol, rhamnose, sucrose, melibiose, amygdalin iyo arabinose. Marka la barbar dhigo noocyada tixraaca ee laxiriira ee la daabacay, astaamaha dufanka gacanta ee nooca ASxL5T ayaa lagu muujiyay shaxda 4. Asiidhyada dufanka gacanta ee ugu muhiimsan waa C16: 1ω6c iyo / ama C16: 1ω7c, C16: 0 iyo C18: 1ω9. Hydroxy fatty acids C12:0 3-OH iyo C10:0 3-OH sidoo kale way jiraan. Saamiga C16:0 ee ASxL5T wuu ka sarreeyaa qiimaha la soo sheegay ee hiddaha la xidhiidha. Taas bedelkeeda, marka la barbardhigo T. marinus IMCC1826TT, saamiga C18: 1ω7c iyo / ama C18: 1ω6c ee ASxL5T waa la dhimay. oleivorans MIL-1T iyo O. kriegii DSM 6294T, laakiin aan laga helin B. sanyensis KCTC 32220T. Isbarbardhigga astaamaha aashitada dufanka leh ee ASxL5T iyo ASxLS waxay daaha ka qaaday kala duwanaansho dahsoon oo ah qaddarka asiidhka dufanka leh ee shakhsi ahaaneed ee u dhexeeya labada nooc, kuwaas oo la jaan qaadaya taxanaha DNA-da ee isku nooc. Ma jiraan qaybo ka mid ah poly-3-hydroxybutyrate (PHB) oo la ogaaday iyadoo la isticmaalayo baaritaanka madow ee Suudaan.
Dhaqdhaqaaqa ugaadhsiga bakteeriyada ASxL5T waa la daraaseeyay si loo go'aamiyo tirada ugaadhsiga. Bakteeriyadani waxay samayn kartaa huurada noocyada Campylobacter, oo ay ku jiraan: Campylobacter suis 11608T, Campylobacter jejuni PT14, Campylobacter jejuni 12662, Campylobacter jejuni NCTC 11168T; Escherichia coli NCTC 12667; C. helveticus NCTC 12472; C lari NCTC 11458 iyo C. upsaliensis NCTC 11541T. Adeegso dhaqamada ku taxan qaybta go'aaminta kala duwanaanshaha martida loo yahay ee habka si aad u tijaabiso noocyo balaadhan oo ah bakteeriyada Gram-negative iyo Gram-positive. Natiijooyinka waxay muujinayaan in ASxL5T sidoo kale loo isticmaali karo Escherichia coli NCTC 86 iyo Citrobacter freundii NCTC 9750T. Plaques laga sameeyay Klebsiella oxytoca 11466. Isdhexgalka TEM ee E. coli NCTC 86 ayaa lagu muujiyay Jaantuska 4A-D, iyo isdhexgalka Campylobacter jejuni PT14 iyo Campylobacter suis S12 ayaa lagu muujiyay Jaantuska 4E-H dhexe. Habka weerarku wuxuu u muuqdaa inuu ka duwan yahay noocyada ugaadhsiga ee la tijaabiyay, iyadoo hal ama in ka badan oo E. coli ah ay ku xiran yihiin unug kasta oo ASxL5T ah oo la dhigay dhinaca dambe ee unugga fidsan ka hor intaan la dhejin. Taas bedelkeeda, ASxL5T waxay u muuqataa inay ku xiran tahay Campylobacter iyada oo loo marayo hal dhibic oo xiriir ah, badiyaa xiriir la leh cirifka unugyada ugaadhsiga iyo meel u dhow cirifka unugyada Campylobacter (Jaantus 4H).
TEM oo muujinaysa isdhexgalka u dhexeeya ASx5LT iyo ugaadha: (AD) iyo E. coli ugaadha; (EH) iyo C. jejuni ugaadhsi. (A) Unug caadi ah oo ASx5LT ah oo ku xidhan hal unug E. coli (EC); (B) ASx5LT fiilo leh oo ku dheggan hal unug oo EC ah; (C) Unug fiilo leh oo ASx5LT ah oo ku xidhan unugyo EC oo badan; (D) Lifaaqa yaryar ASx5LT unug E. coli (EC); (E) unug keliya oo ASx5LT ah oo ku xidhan unugga Campylobacter jejuni (CJ); (F) ASx5LT waxay weerartaa unugyada C. hyointestinalis (CH); (G) laba hal unug oo ASx5LT ah ayaa weeraray unug CJ ah; (H) Muuqaal dhow oo ah barta ku-xidhka ASx5LT, ee u dhow cirifka unugyada CJ (bar 0.2 μm). Barku waxa uu ka dhigan yahay 1 μm gudaha (A-G).
Bakteeriyada wax ugaadhsiga ah ayaa u kobcay si ay uga faa'iideysato ilo badan oo ugaarsi ah. Sida iska cad, waxay si weyn uga jiraan deegaanno badan oo kala duwan. Cabbirka cidhiidhiga ah ee xubnaha dadweynaha awgeed, waxa suurtogal ah in bakteeriyada ASxL5T laga sooco slurry-ka iyadoo la adeegsanayo habka kala-soocidda phage-ga. Xidhiidhka genomic ee ASxL5T xubnaha qoyska oceanospirillaceae ee bakteeriyada badda waa wax lala yaabo, in kasta oo nooluhu uu u dulqaadan karo milix oo uu ku kori karo dhexdhexaad ka kooban 5% milix ah. Falanqaynta tayada biyaha ee slurry waxay muujisay in maadada sodium chloride ay ka yar tahay 0.1%. Sidaa darteed, dhoobada ayaa aad uga fog deegaanka badda - juqraafi ahaan iyo kiimiko ahaan labadaba. Joogitaanka saddex go'doomin oo isku mid ah laakiin kala duwan oo ka yimid isla isha ayaa caddaynaysa in ugaarsadahan ay ku koraan deegaankan aan badda ahayn. Intaa waxaa dheer, falanqaynta microbiome (faylalka xogta laga heli karo https://www.ebi.ac.uk/ena/browser/view/PRJEB38990) waxay muujisay in isla 16S rRNA taxanaha hiddo-wadaha uu ku yaal 50ka ugu sarreeya cashuuraha hawlgelinta (OTU) ) Dhawr muunad oo dhooqo ah. Dhowr jeer oo bakteeriya ah ayaa laga helay xogta Genbank, kuwaas oo leh 16S rRNA taxane ah oo la mid ah bakteeriyada ASxL5T. Taxanahan, oo ay weheliyaan taxanaha ASxL5T, ASxS5, iyo ASxO5, waxay u muuqdaan inay metelaan lakabyo kala duwan oo ka soocay Thalassolituus iyo Oceanobacter (Jaantus 2). Saddex nooc oo bakteeriya ah oo aan dhaqan lahayn (GQ921362, GQ921357 iyo GQ921396) ayaa laga soocay biyaha fiska ah oo qoto dheer 1.3 kiiloomitir macdanta dahabka ee Koonfur Afrika 2009, iyo labada kale (DQ256320 iyo DQ337006) waxay ka yimaadeen South Africa sanadkii 2005). Taxanaha hidda-wadaha 16S rRNA ee aadka ugu dhow ASxL5T waa qayb ka mid ah taxanaha hiddo-wadaha 16S rRNA ee laga helay dhaqanka kobcinta ee sediments bacaadka ah ee laga helay xeebaha waqooyiga Faransiiska 2006 (lambarka gelitaanka AM29240828). Isku xigxiga hiddo-wadayaasha 16S rRNA ee kale ee xidhiidh dhow la leh bakteeriyada aan la dhaqanin ee HQ183822.1 ayaa laga helay haamo ururineed oo ka soo caarisay qashinka dawladda hoose ee Shiinaha. Sida iska cad, bakteeriyada ASxL5T maaha mid aad uga wakiil ah kaydka taxonomic-ga, laakiin taxanahan bakteeriyada aan la dhaqanin waxay u badan tahay inay metelaan noolaha la midka ah ASxL5T, kuwaas oo lagu qaybiyo adduunka oo dhan, badanaaba jawi adag. Laga soo bilaabo dhammaan falanqaynta genome phylogenetic, qaraabada ugu dhow ASxL5T waa Thalassolituus sp. C2-1, T. marinus, T. oleivorans. iyo O. kriegii 23, 24, 25, 26, 27. Thalassolituus waa xubin ka mid ah bakteeriyada kala qaybsanaanta hydrocarbon ee badda (OHCB), taas oo ku baahsan badda iyo bey'ada dhulka, oo badanaa waxay noqotaa xukunka ka dib dhacdooyinka wasakhowga hydrocarbon30,31. Bakteeriyada badda ma aha xubno ka tirsan kooxda OHCB, laakiin waxay ka go'doonsan tahay deegaanka badda.
Xogta Phenotypic waxay tilmaamaysaa in ASxL5T ay tahay nooc cusub iyo xubin ka mid ah cirifka hore ee aan hore loo aqoonsan ee qoyska spirospiraceae badda. Hadda ma jiro halbeeg cad oo lagu kala saarayo noocyada cusub ee go'doonsan ee nooc cusub. Waxaa la isku dayay in la go'aamiyo xuduudaha guud ee guud, tusaale ahaan, iyadoo lagu salaynayo boqolleyda genome ee borotiinka muxaafidka ah (POCP), waxaa lagu talinayaa in qiimaha goyntu uu yahay 50% la mid ah cadaadiska tixraaca33. Kuwa kale waxay soo jeedinayaan isticmaalka qiyamka AAI, kuwaas oo faa'iidooyin ka leh POCP sababtoo ah waxaa laga heli karaa genomes34 aan dhamaystirnayn. Qoraagu wuxuu aaminsan yahay in haddii qiimaha AAI uu ka yar yahay 74% marka la barbardhigo nooca moodeelka noocyada moodeelka, cadaadiska ayaa wakiil ka ah noocyo kala duwan. Qaabka hiddaha ee spirillaceae marine waa spirillum badda, nooca moodeelkuna waa O. linum ATCC 11336T. Qiimaha AAI ee u dhexeeya ASxL5T iyo O. linum ATCC 11336T waa 54.34%, iyo qiimaha AAI ee u dhexeeya ASxL5T iyo T. oleivorans MIL-1T (nooca nooca genus) waa 67.61%, taas oo muujinaysa in ASxL5T ay u taagan tahay hidde cusub oo ka duwan Thalassolituus. Isticmaalka taxanaha hiddo-wadaha 16S rRNA sida heerka kala soocida, soohdinta xadaynta hiddaha la soo jeediyay waa 94.5%35. ASxL5T waxaa laga yaabaa in lagu dhejiyo genus Thalassolituus, oo muujinaya 95.03% 16S aqoonsiga rRNA oo leh T. oleivorans MIL-1T iyo 96.17%. marinus IMCC1826T. Si kastaba ha ahaatee, waxaa sidoo kale lagu dhejin doonaa Bacteroides genus kaas oo leh 94.64% 16S rRNA aqoonsiga hidda-wadaha B. sanyensis NV9, taas oo muujinaysa in isticmaalka hal hidde sida 16S rRNA hiddaha ay u horseedi karto kala saarid iyo meelayn aan sabab lahayn. Hab kale oo la soo jeediyay ayaa isticmaala ANI iyo Genome alignment Score (AF) si ay u baaraan ururinta dhibcaha xogta ee dhammaan noocyada iyo noocyada aan nooca ahayn ee hiddaha jira. Qoraagu waxa uu ku talinayaa in la isku daro soohdinta cirifka iyo bar-rogid ee cirifka la qiyaasay ee u gaarka ah cashuurta la lafaguray. Si kastaba ha ahaatee, haddii aysan jirin taxane dhammaystiran oo genome ah oo ka yimid go'doominta Thalassolituus, suurtagal maaha in la go'aamiyo in ASxL5T ay ka tirsan tahay hiddaha Thalassolituus habkan. Sababtoo ah helitaanka xaddidan ee taxanaha genome ee dhamaystiran ee falanqaynta, dhammaan geedka genome phylogenetic geed waa in si taxadar leh loo fasiraa. Marka labaad, hababka isbarbardhigga genome-ga oo dhan kuma xisaabtami karaan kala duwanaansho la taaban karo ee cabbirka genome-yada la barbardhigay. Waxay cabbireen isku ekaanshaha hiddo-wadaha hal koobiga ee xudunta u ah ee u dhexeeya hidde-sideyaasha la xidhiidha, laakiin kuma xisaabtamin tirada tirada badan ee hiddo-wadaha aan ku jirin genome-ka aadka u yar ee ASxL5T. Sida iska cad, ASxL5T iyo kooxaha ay ka mid yihiin Thalassolituus, Oceanobacter, iyo Bacterioplanes waxay leeyihiin awoowe wadaag ah, laakiin horumarku wuxuu qaaday waddo kale, taasoo keentay hoos u dhaca genome, kaas oo laga yaabo inuu noqdo inuu la qabsado qaab nololeedka ugaadhsiga. Tani waxay ka duwan tahay T. oleivorans MIL-1T, kaas oo 28% ka weyn oo ku soo kordhay cadaadisyo deegaan oo kala duwan si loo isticmaalo hydrocarbons23,30. Isbarbardhig xiiso leh ayaa lagu samayn karaa dulinnada gacanta ku jira ee waajibka ah iyo astaamaha, sida Rickettsia, Chlamydia, iyo Buchnera. Cabbirkooda genome-ku waa ilaa 1 Mb. Awoodda isticmaalka dheef-shiid kiimikaadka unugga martida loo yahay waxay horseedaa luminta hidda-wadaha, sidaas darteed waxaa la soo maray hoos u dhac weyn oo kobcin ah. Isbeddellada korriinka ee ka imanaya noolaha nafaqada kiimikada badda iyo hab-nololeedyada ugaadhsiga waxay keeni karaan hoos u dhac la mid ah cabbirka genome-ga. Falanqaynta COG iyo KEGG waxay muujinaysaa tirada hidde-sideyaasha loo isticmaalo hawlo gaar ah iyo kala duwanaanshaha caalamiga ah ee waddooyinka genomic ee u dhexeeya ASxL5T iyo T. oleivorans MIL-1T, kuwaas oo aan sabab u ahayn helitaanka baahsan ee unugyada hidde-socodka guurguura. Farqiga u dhexeeya saamiga G + C ee dhammaan genome-ka ASxL5T waa 56.1%, kan T. oleivorans MIL-1T waa 46.6%, taas oo sidoo kale tilmaamaysa in la kala soocay.
Baaritaanka waxa ku jira koodh-samaynta ASxL5T genome-ku waxa uu bixiyaa aragtiyo shaqaynaya oo ku saabsan sifooyinka phenotypic. Joogitaanka hidde-sideyaasha nooca IV fimbriae (Tfp) ayaa ah mid xiiso gaar ah leh sababtoo ah waxay kor u qaadaan dhaqdhaqaaqa unugyada, oo loo yaqaan qulqulka bulshada ama gariir, iyada oo aan lahayn calanka dusha sare. Sida laga soo xigtay warbixinnada, Tfp waxay leedahay hawlo kale, oo ay ku jiraan ugaadhsiga, pathogenesis, samaynta biofilm, qaadashada DNA-da dabiiciga ah, isku-darka unugyada tooska ah iyo horumarinta38. genome-ka ASxL5T waxa uu ka kooban yahay 18 hidde-socod oo dejinaya diguanylate cyclase (enzyme ka talyaanaya beddelka 2 guanosine triphosphate galay guanosine 2 phosphate iyo cyclic diGMP) iyo 6 hiddo-sidaha u dhigma diguanylate cyclase fosfate diguanylate. Hiddaha esterase (catalying demation of cyclic di-GMP to guanosine monophosphate) waa mid xiiso leh sababtoo ah cycl-di-GMP waa farriin labaad oo muhiim ah oo ku lug leh horumarinta biofilm iyo kala-soocidda, dhaqdhaqaaqa, isku-xidhka unugyada iyo fayraska 39, 40 ee habka. Waa in sidoo kale la ogaadaa in Bdellovibrio bacteriovorus, GMP laba jibaaran ayaa lagu muujiyay inuu xakameynayo kala-guurka u dhexeeya nolosha xorta ah iyo qaab nololeedka ugaadhsiga41.
Inta badan cilmi-baarista bakteeriyada ugaadhsiga ah waxay diiradda saartay Bdellovibrio, noolaha Bdellovibrio-u eg, iyo noocyada Myxococcus. Tusaalooyinkan iyo kuwa kale ee la yaqaan ee bakteeriyada ugaarsada waxay sameeyaan koox kala duwan. Iyadoo ay jirto kala duwanaanshiyahan, koox ka mid ah qoysaska borotiinka sifada ah ee ka tarjumaysa noocyada 11 bakteeriyada ugaadhsiga ee la yaqaan ayaa la aqoonsaday3,22. Si kastaba ha ahaatee, kaliya hiddo-sideyaasha O antigen ligase (waaL) ayaa la aqoonsaday, taas oo si gaar ah ugu badan bakteeriyada Gram-negative. Qaabka falanqaynta ma aha mid waxtar leh in ASxL5T loo asteeyo sidii ugaarsade, malaha sababtoo ah waxay isticmaashaa xeelad weerar oo cusub. Helitaanka hiddo-wadaha bakteeriyada ugaadhsiga ah ee kala duwan ayaa gacan ka geysan doona horumarinta falanqaynta xallinta wanaagsan kuwaas oo tixgelinaya caddaynta kala duwanaanshaha shaqada iyo deegaanka ee u dhexeeya xubnaha kooxda. Tusaalooyinka bakteeriyada ugaadhsiga ah ee aan lagu darin falanqayntan waxaa ka mid ah xubnaha Cupriavidus necator42 iyo Bradymonabacteria43, sababtoo ah cilmi-baarayaashu waxay baarayaan bulshooyinka microbial kala duwan, canshuur ugaadhsi badan ayaa la sameeyay.
Tilmaamaha ugu caansan ee bakteeriyada ASxL5T ee ay qabatay sawirka TEM waa qaab-dhismeedkeeda gaarka ah oo dabacsan, kaas oo kor u qaadi kara isdhexgalka bakteeriyada ugaadhsiga. Nooca isdhexgalka lagu arkay wuu ka duwan yahay bakteeriyada kale ee ugaadhsiga ah oo aan hore loo helin ama aan la soo sheegin. Wareegga nolosha bahalnimada ee ASxL5T ee la soo jeediyay ayaa lagu muujiyay sawirka 5. Waxaa jira tusaalooyin yar oo suugaanta ah oo leh qaab dhismeedyo isku mid ah sida aan halkan ku soo sheegayno, laakiin tusaalahan waxaa ka mid ah Terasakiispira papahanaumokuakeensis, bakteeriyada spirillum badda oo leh marmarka qaarkood 44, iyo Alphaproteobacteria, Terasakiella pus , oo hore uga tirsanaan jiray genus Oceanospirillum, bandhiga Waxa loogu yeero "filimka cirifka" 45. Foomamka Cocci waxaa inta badan lagu arkaa dhaqamadii hore, gaar ahaan bakteeriyada leh foomamka qaloocan, sida Vibrio, Campylobacter, iyo Helicobacter 46, 47, 48, taas oo laga yaabo inay matasho xaalad hoos u dhacday. Shaqo dheeraad ah ayaa loo baahan yahay si loo caddeeyo wareegga nolosha saxda ah ee bakteeriyada ASxL5T. Si loo go'aamiyo sida ay u qabato oo u ugaarsato, iyo in genome-kiisu uu qeexayo isku-dhisyada noolaha firfircoon ee loo isticmaali karo ujeedooyin caafimaad ama bayooloji.
Sharaxaada Venatorbacter gen. Noofambar Venatorbacter (Ven.a.tor, ba'c.ter, L. waxay ka kooban tahay venators ka L. n. venator,'Ugaarsiga' iyo Gr. n. bakteeriyada,' rod'. Venatorbacter,'a Ugaadhsiga Ugaadhsiga' Unugyadu waa aerobic, dulqaad milix ah, qalooca Gram-negative, usha jimicsiga ee PHB kuma ururto Heerkulka inta u dhaxaysa 4 ilaa 42 ° C Koritaanka pH ee 4-9 waa mid aan caadi ahayn. C18:1ω9 ; C12:0 3-OH iyo C10:0 3-OH waxaa laga helaa dufanka hydroxy Asiidhka .
Sharaxaada Venatorbacter cucullus sp. Noofambar Venatorbacter cucullus (cu'cull.us.; L. n. cucullus macnaheedu waa cadaalad).
Intaa waxaa dheer, sifada qeexan ee hiddahan ayaa ah in marka lagu korayo BA ama BHI, unugyadu ay dhererkoodu yihiin 1.63 µm iyo 0.37 µm ballaaran. Deegaannada ku yaal agar BHI aad bay u yar yihiin, iyagoo gaadhaya dhexroor 2 mm 72 saacadood ka dib. Waxay yihiin beige, translucent, wareegsan, convex iyo dhalaalaya. Xubnaha noocaan ah waxay isticmaali karaan Escherichia coli iyo Klebsiella. Campylobacter iyo dhowr bakteeriyayaal kale oo Gram-negative ah ayaa u adeega sida ugaadhsiga.
Cadaadiska caadiga ah ee ASxL5T ayaa laga soocay caanaha lo'da ee Nottinghamshire, UK, waxaana lagu kaydiyay Ururinta Dhaqanka Nooca Qaranka (UK): lambarka gelitaanka NCTC 14397 iyo Ururka Dhaqanka Bakteeriyada Nederlaan (NCCB) lambarka gelitaanka NCCB 100775. Isku-xigta genome ee dhammaystiran ee ASxL5T waxaa lagu shubay Genbank iyadoo loo eegayo kordhinta CP046056.
Bakteeriyada ASxL5T ayaa laga soocay caanaha lo'da iyadoo la adeegsanayo tignoolajiyada go'doominta fage9,49. Soodhaha waxaa lagu qasi jiray 1: 9 (w/v) ee SM bakhaar (50mM Tris-HCl [pH 7.5], 0.1 M NaCl, 8 mM MgSO4.7H2O iyo 0.01% jelatin; Sigma Aldrich, Gillingham, UK), Kadibna ku shub at 4°C ilaa 24 saacadood, si tartiib ah u rogrogaya si ay ugaarsata ugaadhsiga u galaan kaydka. Joojinta ayaa la saaray 3000g 3 daqiiqo. Mashiinka sare ayaa la ururiyey oo la saaray 13,000g mar labaad muddo 5 daqiiqo ah. Ka-hortagga ayaa markaa la dhex mariyay shaandhada 0.45 µm xuubka (Minisart; Sartorius, Gottingen, Jarmalka) iyo 0.2 µm shaandhada xuubka (Minisart) si meesha looga saaro wixii unugyo bakteeriyada soo hadhay. ASxL5T way gudbi kartaa filtarradan. Cawska agar jilicsan ee Campylobacter enterosus S12 (lambarka gelitaanka NCBI ee CP040464) ee isla slurry ayaa la diyaariyey iyadoo la adeegsanayo farsamooyinka caadiga ah. Soodhaha la sifeeyay ayaa lagu qaybiyey mid kasta oo ka mid ah taarikada unugga martida loo yahay ee 10 µl dhibco ah oo saddex geesood ah waxaana loo oggolaaday inuu qallajiyo. Saxanka waxaa lagu dhex dajiyay taangiga microaerophilic at 37 ° C 48 saacadood xaaladaha microaerobic (5% O2, 5% H2, 10% CO2, iyo 80% N2). Qoryaha muuqda ee la helay ayaa lagu soo saaray SM baffer waxaana loo wareejiyay cawska cusub ee C. hyointestinalis S12 si loo sii faafiyo noolaha lysed. Marka la go'aamiyo in bakteeriyadu ay tahay sababta huurada lytic ee aysan ahayn phage, isku day inaad u koraan noolaha si madax-bannaan oo ka madax bannaan martigeliyaha oo aad si dheeraad ah u siisid. Dhaqanka aerobic-ka waxaa lagu sameeyay 37 °C oo leh 5% v/v dhiiga faraska defibrinated (TCS Biosciences Lt, Buckingham, UK, supplement). Marka loo eego tilmaamaha Guddiga Heerarka Caafimaadka Qaranka, habka faafinta saxanka waxaa loo isticmaalaa baaritaanka nuglaanshaha bakteeriyada. BHI agar waxaa lagu dhaqay 37 °C iyadoo la isticmaalayo saxan ka kooban antibiyootiga soo socda (Oxoid) ee dhaqanka hawada: amoxicillin iyo clavulanic acid 30 µg; cefotaxime 30 µg; streptomycin 10 µg; ciprofloxacin 5 µg; Ceftazidime 30 µg Nalidixic acid 30 µg; Imipenem 10 µg; Azithromycin 15 µg; Chloramphenicol 30 µg; Cefoxitin 30 µg; Tetracycline 30 µg; Nitrofurantoin 300 µg; Aztreonam 30 µg; Ampicillin 10 µg; Cefpodoxime 10 µg; Trimethoprim-Sulfamethoxazole 25 µg. Dulqaadka milixda waxaa lagu aasaasay kuleejka hawada ee taarikada agar BHI at 37 °C. NaCl dheeraad ah ayaa lagu daray taarikada agar ee BHI si ay u bixiso xaddi laxaad leh oo ah ilaa 10% w/v. Qiyaasta pH waxaa lagu go'aamiyaa dhaqanka aerobic ee taarikada agar ee BHI at 37 ° C, halkaasoo qiyaasta pH lagu hagaajiyay inta u dhaxaysa 4 iyo 9 oo leh HCl nadiif ah ama NaOH nadiif ah, iyo qiimaha pH ee bartilmaameedka ah ayaa la xaqiijiyay ka hor inta aan la shubin saxanka. Falanqaynta aashitada dufanka gacanta, ASxL5T waxaa lagu beeray agar BHI muddo 3 maalmood ah iyo aerobic at 37 °C. Marka loo eego MIDI (Nidaamka Aqoonsiga Sherlock Microbial, nooca 6.10) borotokoolka caadiga ah ee FERA Science Ltd, (York, UK), asiidhka dufanka unugyada ayaa la soo saaray, la diyaariyay oo la falanqeeyay.
Dhanka TEM, ASxL5T waxaa lagu dhaqay aerobic iyadoo si isku mid ah ugu fiday BA at 37°C mudo 24 saacadood ah, ka dibna waxaa la goostay 1 ml oo ah 3% (v/v) glutaraldehyde gudaha 0.1 M cacodylate baffer heerkulka qolka hagaajin 1 saac, ka dibna centrifuge 10,000 g 3 daqiiqo. Ka dib si tartiib ah dib ugu celi pelletka 600 μl 0.1 M cacodylate baffer. Ku wareeji ganaax go'an ASxL5T filimka Formvar/kaarboon ee shabaqyada naxaasta ah ee 200 mesh ah. Bakteeriyada waxaa lagu wasakheeyay 0.5% (w/v) uranyl acetate 1 daqiiqo waxaana baadhay TEM iyadoo la isticmaalayo TEI Tecnai G2 12 Biotwin mikroskoob. Sida kor ku xusan, isku dar tiro isku mid ah ugaadhsiga iyo ugaadhsiga ee maraq NZCYM (BD Difco ™, Fisher Scientific UK Ltd, Loughborough) oo ku dhaji saacadaha 48 ee xaaladaha microaerobic ee Campylobacter ama Campylobacter at 37 ° C , Isdhexgalka ugaadhsiga iyo ugaadhsiga waxaa sidoo kale baaray TEM. Xaaladaha hawada ee Escherichia coli. Si madaxbanaan u baadh ugaadhsiga iyo bakteeriyada ugaadhsiga si aad u go'aamiso isbeddel kasta oo ku yimaada qaab-dhismeedka unugyada ee ugaadhsiga awgeed. Habka madow ee Suudaan waxa loo isticmaalay mikroskoobyada indhaha ee ururinta PHB.
Ku kori ASxL5T dhaqamada habeen dhaxa adiga oo koriinka BHI ama saxanada BA ku swab nadiif ah. Ururi unugyada ASxL5T oo ka jooji MRD (CM0733, Oxoid), ka dibna dhig 4°C ilaa 7 maalmood si ay unugyadu u gaajoodaan. Tixraaca NCTC ama dhaqanka bakteeriyada kaydka shaybaadhka ayaa lagu tallaalay maraq BHI ama maraq nafaqeeyayaal lambar 2 (CM007, Oxoid), oo habeen dhaxa lagu dhex shubay, oo la saaray 13,000g dibna loogu celiyay MRD ilaa OD600 uu ahaa 0.4. Dhaqanka: Bacillus subtilis NCTC 3610T, Citrobacter freundii NCTC 9750T, Enterobacter aerogenes NCTC 10006T, Enterococcus faecalis NCTC 775T, Escherichia coli NCTC 86, Klebsiella 46ocalis NCTC 10817, Listeria Special Bakteeriyada NCTC 4885, Bacillus macerans NCTC 6355T, Providencia stuartsii NCTC 10318, Pseudomonas fluorescens SMDL, Rhodococcus submarine hamburger NCTC 1621T, Salmonella Moncilde NCTC bakteeriyada NCTC. 10861, Staphylococcus aureus NCTC 8532T, Streptococcus pneumoniae NCTC 7465T, Yersinia enterocolitica NCTC 10460. Martigeliyaha Campylobacter waxa si microaerobically ah loogu dhex galiyay saxanka BA at 37°C waxaana la hakiyay maraq N.ZY Xeriyayaasha Campylobacter ee la tijaabiyey waa: C. coli 12667 NCTC, C. jejuni 12662, C. jejuni PT14, C. jejuni NCTC 11168T, C. helveticus NCTC 12472, C. lari NCTC 11445NC, C.111niTC. PT14. Ku dar aliquot 0.5 ml oo ganaax ah 5 ml oo dhalaalay NZCYM sare agar (0.6% agar) oo ku shub saxanka hoose ee 1.2% NZCYM. Ka dib markii la daaweeyay oo la qalajiyey, ASxL5T si taxane ah loo badhxay ayaa loo qaybiyey sidii 20 µl dhibco ah oo loox kasta oo cawska ah iyadoo saddex jeer ah. Heerkulka dhaqanka iyo jawigu waxay ku xiran yihiin shuruudaha bakteeriyada baaritaanka.
Isticmaal GenElute™ Bakteeriyada Genomic DNA Kit (Sigma Aldridge) si aad DNA uga diyaariso go'doominta bakteeriyada. Hababka caadiga ah ayaa loo adeegsaday kor u qaadida PCR ee hiddo-wadaha 16S rRNA iyo go'aaminta taxanaha badeecada iyadoo la adeegsanayo kiimikada joojinta dheeha (Adeegga Akhriska Qiimaha Yurub, Jarmalka). Isticmaal barnaamijka BLAST-N si aad u barbar dhigto taxanahan iyo taxanaha hiddo-wadaha 16S rRNA si aad u aqoonsato oo aad u ururiso noocyada xidhiidhka dhow la leh. Kuwaas waxaa lagu toosiyay iyadoo la isticmaalayo ClustalW barnaamijka MEGA X. Geedka phylogenetic waxaa dib loo dhisay iyadoo la adeegsanayo MEGA X iyadoo la adeegsanayo habka suurtogalka ah ee ugu badan ee ku saleysan qaabka Tamura-Nei, oo leh 1000 nuqul oo la hagayo54. Isticmaal PureLink™ Genomic DNA Kit (Fisher Scientific, Loughborough, UK) si aad u soo saarto DNA-da isku xigxiga-genome-ga oo dhan. Taxanaha hiddo-wadaha ASxL5T waxaa lagu go'aamiyay iyadoo la adeegsanayo isku-dhafka Illumina MiSeq, kaas oo ka kooban 250 bp akhrin laba-dhammaan ah oo ka kooban maktabad la diyaariyay iyadoo la adeegsanayo xirmada calaamadaynta Nextera iyo 2 ilaa 20 kb dheer oo akhrin ah oo laga soo qaatay madal PacBio. Genomics DNA Sequencing Xarunta Cilmi-baarista ee Jaamacadda Sembia. genome-ka waxaa la isugu keenay iyadoo la isticmaalayo CLC Genomics Workbench 12.0.3 (Qiagen, Aarhus, Denmark). Dhaqamada ASxL5T waxa lagu kaydiyaa Ururinta Dhaqanka Nooca Qaranka (UK) iyo Ururka Dhaqanka Bakteeriyada Nederlaan (NCCB). Genomy-yada noolaha la xidhiidha ee loo isticmaalo isbarbardhigga waa: Thalassolituus oleivorans MIL-1T (lambarka gelitaanka HF680312, dhamaystiran); Bacterioplanes sanyensis KCTC 32220T (lambarka gelitaanka wuxuu01000001, aan dhamaystirnayn); Oceanobacter kriegii DSM 6294T (lambarka gelitaanka NZ_AUGV00000000, aan dhamaystirnayn); Bulshada Marinamonas DSM 5604T (ku daray ASM436330v1, aan dhamaystirnayn), Oceanospirullum linum ATCC 11336T (ku daray MTSD02000001, aan dhamaystirnayn) iyo Thalassolituus sp. C2-1 (ku dar NZ_VNIL01000001, aan dhamaystirnayn). Adeegso JGI Genome Portal36 at https://img.jgi.doe.gov//cgi-bin/mer/main.cgi?section=ANI&page= si aad u go'aamiso dhibcaha toosinta (AF) iyo celceliska aqoonsiga nucleic acid (ANI). Laba-labo. Habka Rodriguez-R & Konstantinidis55 ayaa loo isticmaalay si loo go'aamiyo aqoonsiga amino acid (AAI). Isticmaal GToTree 1.5.5411,12,13,14,15,16,17,18 si aad u dhaliso qiyaasta ugu badan ee suurtogalka ah ee geedka phylogenetic. Genom-ga wax-soo-gelinta ee matalaya hiddo-wadaha tixraaca ee la heli karo waxa laga soo xushay hidde-raac tixraaceedka loo aqoonsaday inay xidhiidh la leeyihiin ASxL5T oo ka socda phylogeny 16S rRNA. Waxaa lagu sharraxay geedka iyadoo la adeegsanayo geedka is-dhexgalka ee aaladda onlaynka ah (https://itol.embl.de/). Sharaxaada shaqada iyo falanqaynta genome-ka ASxL5T waxaa lagu fuliyaa iyadoo la adeegsanayo BlastKOALA KEGG aaladda khadka tooska ah iyadoo la adeegsanayo qaybinta kobcinta moduleka KEGG (Kyoto Encyclopedia of Genes and Genomes). Qaybinta qaybaha COG (kooxaha orthologue) waxaa lagu go'aamiyaa iyadoo la isticmaalayo eggNOG-mapper aaladda khadka tooska ah.
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