Ụdị ọhụrụ nke Gram-negative, aerobic, salt-tolerant, arụ ọrụ, mkpara, na nje bacteria ASxL5T dịpụrụ adịpụ na ọdọ mmiri nsị ehi na Nottinghamshire, England, ma jiri Campylobacter mee anụ. Mgbe nke ahụ gasịrị, a chọtara ụdị Campylobacter ndị ọzọ na ndị ezinụlọ Enterobacteriaceae dị ka anụ oriri. Mgbe subculture na-enweghị mkpụrụ ndụ nnabata, enwetara uto aseptic na-adịghị ike na Brain Heart Infusion Agar. Ọnọdụ uto kacha mma bụ 37 Celsius C yana pH bụ 7. Nleba anya nyocha eletrọn ekpughere ụfọdụ atụmatụ morphological pụrụ iche metụtara nnweta anụ oriri. Nyocha phylogenetic site na iji usoro mkpụrụ ndụ ihe nketa 16S rRNA gosiri na mwepu ahụ na-emetụta onye otu ezinụlọ Marine Spirulina, mana enweghị ike ikewa ya nke ọma dị ka onye otu ụdị ama ama ọ bụla. Usoro nke ASxL5T nke genome dum kwadoro mmekọrịta ya na ndị otu spirochetes mmiri. Nnyocha nchekwa data gosiri na ọtụtụ ASxL5T na-ekekọrịta usoro mkpụrụ ndụ ihe nketa 16S rRNA nwere ọtụtụ nje bacteria na-enweghị omenala sitere na oke osimiri, elu ala na mmiri ala. Anyị na-atụ aro na ụdị ASxL5T na-anọchite anya ụdị ọhụrụ na ụdị ọhụrụ. Anyị na-akwado aha Venatorbacter cucullus gen. Nọvemba, sp. N'ime Nọvemba, ASxL5T ejiri mee ụdị ụdị ahụ.
Nje bacteria na-eri anụ bụ nje bacteria na-egosipụta ike ịchụ nta na igbu nje bacteria ndị ọzọ dị ndụ iji nweta ihe ndị na-emepụta ihe na ume. Nke a dị iche na mgbake n'ozuzu nke nri sitere na microorganisms nwụrụ anwụ, ọ dịkwa iche na mmekọrịta parasitic, nke nje bacteria na-enwe mmekọrịta chiri anya na ndị ọbịa ha n'egbughị ha. Nje bacteria na-eri anụ ewepụtala usoro ndụ dị iche iche iji nweta ohere nri dị ukwuu na niche ebe a na-ahụ ha (dị ka ebe obibi mmiri). Ha bụ otu dị iche iche nke taxonomically, nke ejikọtara naanị site na okirikiri ndụ nke ịmụ nwa pụrụ iche1. Achọtala ihe atụ nke nje bacteria na-eri anụ n'ọtụtụ phyla dị iche iche, gụnyere: Proteobacteria, Bacteroides, na Chlorella.3. Otú ọ dị, nje bacteria ndị na-eri anụ ndị a na-amụ nke ọma bụ Bdellovibrio na Bdellovibrio-na-dị ka ntule (BALOs4). Nje bacteria na-eri anụ bụ isi iyi na-ekwe nkwa nke ogige ọhụrụ na-arụ ọrụ ndụ yana ihe nje nje5.
Ekwenyere na nje bacteria na-eri anụ na-eme ka ụdị microbial dị iche iche dịkwuo mma ma nwee mmetụta dị mma na ahụike gburugburu ebe obibi, mmepụta na nkwụsi ike6. N'agbanyeghị àgwà ndị a dị mma, ọ dị ole na ole ọmụmụ banyere nje bacteria ọhụrụ na-eri anụ n'ihi ihe isi ike nke ịzụlite nje bacteria na mkpa ọ dị iji nlezianya na-ekiri mmekọrịta sel iji ghọta usoro ndụ ha dị mgbagwoju anya. Ozi a adịghị mfe ịnweta site na nyocha kọmputa.
N'ime oge nke nguzogide mgbochi nje na-abawanye, a na-amụ atụmatụ ọhụrụ maka ikwado nje nje bacteria, dị ka ojiji nke bacteriophages na nje bacteria7,8. Ekewapụrụ nje bacteria ASxL5T na 2019 site na iji teknụzụ kewapụ phage sitere na nsị ehi anakọtara na ebe mmiri ara ehi nke Mahadum Nottingham, Nottinghamshire. Ebumnuche nke nyocha a bụ ikewapụ ihe ndị dị ndụ nwere ike dị ka ndị na-ahụ maka njikwa ndu. Campylobacter hyointestinalis bụ ọrịa zoonotic, nke na-ejikọta ya na ọrịa eriri afọ mmadụ10. Ọ bụ ebe niile na serum ma jiri ya mee ihe dị ka onye ọbịa lekwasịrị anya.
Nje bacteria ASxL5T dịpụrụ adịpụ na jelii beef n'ihi na a hụrụ na plaques ndị ọ kpụrụ na ahịhịa C. hyointestinal yiri nke nje bacteria na-emepụta. Nke a bụ nchọpụta a na-atụghị anya ya, n'ihi na akụkụ nke usoro ikewapụ phage gụnyere nzacha site na nzacha 0.2 µm, nke e mere iji wepụ mkpụrụ ndụ nje. Nnyocha a na-ahụ anya nke ihe ndị e si na mbadamba nkume ahụ pụta gosiri na obere nje bacteria na-agbagọ agbagọ nke yiri gram adịghị ekpokọta polyhydroxybutyrate (PHB). A na-enweta omenala aseptic nke na-adabereghị na mkpụrụ ndụ anụ oriri na usoro siri ike siri ike (dịka ụbụrụ obi infusion agar (BHI) na agar ọbara (BA)), na uto ya adịghị ike. Ọ na-enwetara mgbe subculture na arọ inoculum mma. Ọ na-eto nke ọma n'okpuru microaerobic (7% v / v oxygen) na ọnọdụ ikuku oxygen, ma ọ bụghị na ikuku anaerobic. Mgbe awa 72 gachara, dayameta nke ógbè ahụ dị obere, na-eru 2 mm, ma ọ bụ beige, translucent, gburugburu, convex na na-egbuke egbuke. A na-egbochi nnwale biochemical maka ASxL5T enweghị ike ịzụlite nke ọma na mgbasa ozi mmiri mmiri, nke na-egosi na ọ nwere ike ịdabere na usoro ndụ mgbagwoju anya nke nhazi biofilm. Agbanyeghị, nkwusioru efere ahụ gosipụtara na ASxL5T bụ aerobic, dị mma maka oxidase na catalase, ma nwee ike ịnabata 5% NaCl. ASxL5T na-eguzogide ọgwụ 10 μg streptomycin, mana ọ na-enwe mmetụta maka ọgwụ nje ndị ọzọ niile a nwalere. A na-enyocha mkpụrụ ndụ nje bacteria ASxL5T site na TEM (Foto 1). Mgbe a na-eto eto na-enweghị mkpụrụ ndụ anụ oriri na BA, mkpụrụ ndụ ASxL5T bụ obere Campylobacter, na nkezi ogologo nke 1.63 μm (± 0.4), obosara nke 0.37 μm (± 0.08), na otu ogologo ogologo (ruo 5 μm). Flaella mmekọahụ. Ihe dị ka 1.6% nke mkpụrụ ndụ na-egosi na ha nwere obosara nke na-erughị 0.2 μm, nke ga-enye ohere ịgafe ngwaọrụ nzacha. Ahụrụ ndọtị nhazi pụrụ iche n'elu sel ụfọdụ, dị ka ihe ngosi (Latin cucullus) (lee akụ ndị dị na 1D, E, G). Nke a dị ka ihe mejupụtara akpụkpọ ahụ dị n'èzí, nke nwere ike ịbụ n'ihi mbelata ngwa ngwa nke envelopu periplasmic, ebe akpụkpọ ahụ dị n'èzí na-anọgide na-adịchaghị mma, na-egosi ọdịdị "adịghị mma". Ịmepụta ASxL5T na enweghị nri (na PBS) ogologo oge na 4 ° C mere ka ọtụtụ (ma ọ bụghị ihe niile) sel na-egosi morphology coccal (Figure 1C). Mgbe ASxL5T tolitere na Campylobacter jejuni dị ka anụ oriri ruo awa 48, nkezi nha cell dị ogologo ma dị warara karịa mkpụrụ ndụ toro na-enweghị onye ọbịa (Table 1 na Figure 1E). N'ụzọ dị iche, mgbe ASxL5T na-eto eto na E. coli dị ka anụ oriri maka awa 48, nkezi sel dị ogologo ma gbasaa karịa mgbe ọ na-eto eto na-enweghị anụ oriri (Table 1), na ogologo cell na-agbanwe agbanwe, na-egosipụtakarị filamentous (Figure 1F). Mgbe etinyere ya na Campylobacter jejuni ma ọ bụ E. coli dị ka anụ oriri maka awa 48, mkpụrụ ndụ ASxL5T egosighi flagella ọ bụla. Tebụl 1 na-achịkọta nleba anya mgbanwe na nha cell dabere na ọnụnọ, enweghị, na ụdị anụ oriri nke ASxL5T.
Ngosipụta TEM nke ASx5LT: (A) ASx5LT na-egosi ụtarị ogologo; (B) batrị ASx5LT nkịtị; (C) cocci ASx5LT mkpụrụ ndụ mgbe ogologo incubation na-enweghị nri; (D) otu mkpụrụ ndụ ASx5LT na-egosi ihe na-adịghị mma (E) ASx5LT cell group incubated with Campylobacter prey gosiri ụbara ogologo cell ma e jiri ya tụnyere ndị na-enweghị anụ oriri (D) gosikwara apical Ọdịdị; (F) nnukwu Flamentous flagella, sel ASx5LT, mgbe etinyere ya na E. coli prey; (G) Otu cell ASx5LT ka emechara ya na E. coli, na-egosi nhazi elu pụrụ iche. Ogwe ahụ na-anọchi anya 1 μm.
Ịchọpụta usoro mkpụrụ ndụ ihe nketa 16S rRNA (ọnụọgụ nnweta MT636545.1) na-enyere nyocha data aka ịmepụta usoro yiri nke ahụ na klas Gammaproteobacteria, ma dị nso na nje bacteria na mmiri na ezinụlọ spirillum mmiri (Onyonyo 2), ma bụrụ ndị òtù Thalassolituus genus. Onye kacha nso na Marine Bacillus. Usoro mkpụrụ ndụ ihe nketa 16S rRNA dị iche na nje bacteria na-eri anụ nke ezinụlọ Bdelvibrionaceae (Deltaproteobacteria). Ntụnyere ụzọ abụọ nke B. bacteriovorus HD100T (ụdị ụdị, DSM 50701) na B. bacteriovorus DM11A bụ 48.4% na 47.7%, na maka B. exovorus JSS ọ bụ 46.7%. Nje bacteria ASxL5T nwere 3 nke mkpụrụ ndụ ihe nketa 16S rRNA, abụọ n'ime ha yiri ibe ha, nke atọ bụ ntọala atọ dị iche iche. Ihe abụọ ọzọ dịpụrụ adịpụ nje na-eri anụ (ASx5S na ASx5O; 16S rRNA ọnụọgụ ọnụọgụgụ bụ MT636546.1 na MT636547.1, n'otu n'otu) nwere ọdịdị ọdịdị ọdịdị na ọdịdị phenotypic site n'otu ebe ahụ abụghị otu, mana ha dị iche na ASxL5T na nje bacteria na-emeghị omenala. A na-achịkọta usoro nchekwa data yana usoro ndị ọzọ Oceanospirillaceae (ọgụgụ 2). Ekpebiri ma chekwaa usoro genome niile nke ASxL5T na nchekwa data NCBI, yana nọmba ntinye bụ CP046056. Mkpụrụ ndụ ihe nketa nke ASxL5T nwere chromosome okirikiri nke 2,831,152 bp nwere oke G + C nke 56.1%. Usoro genome nwere 2653 CDS (mkpokọta), nke 2567 na-ebu amụma na ọ ga-edobe protein, nke 1596 nwere ike kenye dị ka ọrụ ntinye (60.2%). Mkpụrụ ndụ ihe nketa nwere mkpụrụ ndụ ihe nketa 67 RNA, gụnyere 9 rRNA (3 nke ọ bụla maka 5S, 16S na 23S) na 57 tRNA. Atụnyere njirimara mkpụrụ ndụ ihe nketa nke ASxL5T na mkpụrụ ndụ ihe nketa dịnụ nke ụdị ikwu kacha nso nke achọpụtara na usoro 16S rRNA (Table 2). Jiri njirimara amino acid (AAI) tulee genome Thalassolituus niile dị na ASxL5T. Usoro genome kacha nso dị (ezughị ezu) nke AAI kpebiri bụ Thalassolituus sp. C2-1 (tinye NZ_VNIL01000001). Adịiche a dịpụrụ adịpụ na sedimenti miri emi nke Mariana Trench, mana ugbu a enweghị ozi phenotypic gbasara ụdị a maka ntụnyere. E jiri ya tụnyere ASxL5T's 2.82 Mb, genome nke akụkụ ahụ buru ibu na 4.36 Mb. Nkezi genome nke spirochetes mmiri dị ihe dịka 4.16 Mb (± 1.1; n = 92 genomes ntụaka zuru ezu nyochara site na https://www.ncbi.nlm.nih.gov/assembly), yabụ genome nke ASxL5T dabara na ịtụ atụnyere ndị otu ndị ọzọ, ọ dị obere. Jiri GToTree 1.5.54 wepụta genome nke echere na o nwere ike ịbụ osisi phylogenetic (Onyonyo 3A), na-eji usoro amino acid agbakwunyere na jikọtara ya nke mkpụrụ ndụ ihe nketa 172 akọwapụtara nke ọma na Gammaproteobacteria 11,12,13,14,15,16, 17,18. Nnyocha ahụ gosiri na ọ nwere njikọ chiri anya na Thalassolituus, Bacterial Plane, na Bacterium mmiri. Agbanyeghị, data ndị a na-egosi na ASxL5T dị iche na ndị ikwu ya na Marine Spirulina yana data usoro genome ya dị.
Osisi phylogenetic nke na-eji usoro mkpụrụ ndụ 16S rRNA na-eme ka ọnọdụ nke ASxL5T, ASxO5, na ASxS5 (ya na guts) n'ihe metụtara nje bacteria na-adịghị akọ na mmiri na mmiri Spirulinaceae. Nọmba nbanye Genbank na-esote aha nsogbu dị na mbo. Jiri ClustalW hazie usoro, wee jiri usoro nwere ike yana ụdị Tamura-Nei iji nweta mmekọrịta phylogenetic, wee mee mmegharị 1000 eduzi na mmemme MEGA X. Ọnụọgụ dị na alaka ụlọ ọrụ na-egosi na uru nnomi eduzi karịrị 50%. A na-eji Escherichia coli U/541T mee ihe dị ka mpụta.
(A) Osisi phylogenetic nke dabeere na genome, na-egosi mmekọrịta dị n'etiti mmiri Spirospiraceae bacterium ASxL5T na ndị ikwu ya, E. coli U 5/41T dị ka otu ìgwè. (B) E jiri ya tụnyere T. oleivorans MIL-1T, a na-ebu amụma na ngalaba na-arụ ọrụ nke mkpụrụ ndụ ihe nketa dabere na ụyọkọ orthologous (COG) nke protein ASx5LT. Ọnụ ọgụgụ dị n'aka ekpe na-egosi ọnụọgụ mkpụrụ ndụ ihe nketa na ngalaba COG ọ bụla na-arụ ọrụ na genome ọ bụla. Eserese dị n'aka nri na-egosi pasentị genome dị n'ime otu COG na-arụ ọrụ ọ bụla. (C) E jiri ya tụnyere T. oleiverans MIL-1T, nyocha nke KEGG zuru ezu (Kyoto Encyclopedia of Genes and Genomes) ụzọ modular nke ASxL5T.
Iji nchekwa data KEGG nyochaa mkpụrụ ndụ ihe nketa dị na ASxL5T genome kpughere ụdị ụzọ metabolic nke aerobic gamma Proteus. ASxL5T nwere ngụkọta nke mkpụrụ ndụ ihe nketa 75 e kenyere proteins moto nke nje, gụnyere mkpụrụ ndụ ihe nketa na-etinye aka na chemotaxis, mgbakọ flagella, na ụdị IV fimbriae usoro. Na ngalaba nke ikpeazụ, 9 n'ime mkpụrụ ndụ ihe nketa 10 na-ahụ maka mmegharị mmegharị nke ụdị dị iche iche. Mkpụrụ ndụ ihe nketa nke ASxL5T nwere ụzọ biosynthetic tetrahydropyrimidine zuru oke nke na-ekere òkè na nzaghachi nchebe na nrụgide osmotic20, dị ka a tụrụ anya maka halophiles. genome nwekwara ọtụtụ ụzọ zuru oke maka cofactors na vitamin, gụnyere ụzọ njikọ riboflavin. Ọ bụ ezie na mkpụrụ ndụ alkane 1-monooxygenase (alkB2) dị na ASxL5T, ụzọ ojiji hydrocarbon ezughi oke. N'usoro genome nke ASxL5T, homologues nke mkpụrụ ndụ ihe nketa chọpụtara na ọ bụ isi kpatara mmebi nke hydrocarbons na T. oleiverans MIL-1T21, dị ka TOL_2658 (alkB) na TOL_2772 (mmanya dehydrogenase) adịghị adị. Ọgụgụ 3B na-egosi ntụnyere nkesa mkpụrụ ndụ ihe nketa na ngalaba COG n'etiti ASxL5T na mmanụ oliv MIL-1T. N'ozuzu, genome ASxL5T pere mpe nwere mkpụrụ ndụ ihe nketa pere mpe site na ngalaba COG ọ bụla ma e jiri ya tụnyere genome nwere njikọ buru ibu. Mgbe egosipụtara ọnụ ọgụgụ nke mkpụrụ ndụ ihe nketa na ngalaba ọrụ ọ bụla dị ka pasent nke genome, a na-ahụta ọdịiche dị na pasent nke mkpụrụ ndụ ihe nketa na ntụgharị, usoro ribosomal na ụdị biogenesis, yana mmepụta ume na ọrụ ntụgharị, nke mejupụtara ASxL5T buru ibu. genome A na-atụnyere pasentị ahụ n'otu otu dị na T. oleiverans MIL-1T genome. N'ụzọ dị iche, ma e jiri ya tụnyere ASxL5T genome, T. oleivorans MIL-1T nwere pasentị dị elu nke mkpụrụ ndụ ihe nketa na mmegharị, nhazigharị na nrụzi, na ụdị ederede. N'ụzọ na-akpali mmasị, nnukwu ọdịiche dị na ọdịnaya nke ụdị ọrụ ọ bụla nke genome abụọ ahụ bụ ọnụọgụ nke mkpụrụ ndụ ihe nketa amabeghị dị na ASxL5T (Njirimara 3B). Emere nyocha nkwalite nke modul KEGG, ebe modul KEGG ọ bụla na-anọchite anya otu nkeji arụ ọrụ ejiri aka kọwaa maka nkọwa na nkọwa ndu nke data usoro genome. Ntụnyere nkesa mkpụrụ ndụ ihe nketa na ụzọ KOG module zuru oke nke ASxL5T na olive MIL-1T ka egosiri na eserese 3C. Nyocha a na-egosi na ọ bụ ezie na ASxL5T nwere ụzọ sọlfọ zuru ezu na nitrogen metabolic ụzọ, T. oleiverans MIL-1T adịghị. N'ụzọ dị iche, T. oleiverans MIL-1T nwere usoro cysteine ​​​​na methionine zuru oke, mana ọ ezughị ezu na ASxL5T. Ya mere, ASxL5T nwere modul njirimara maka assimilation sulfate (akọwapụtara dị ka usoro nke mkpụrụ ndụ ihe nketa nwere ike iji dị ka akara phenotypic, dị ka ikike metabolic ma ọ bụ pathogenicity; https://www.genome.jp/kegg/module.html) Na T. Oleiverans MIL-1T. Tụnyere ọdịnaya mkpụrụ ndụ ihe nketa nke ASxL5T na ndepụta nke mkpụrụ ndụ ihe nketa na-egosi ụdị ndụ anụ ọhịa enweghị njedebe. Ọ bụ ezie na waaL gene na-etinye ligase jikọtara ya na O antigen polysaccharide na isi dị na ASxL5T genome (ma ọ na-emekarị na ọtụtụ nje bacteria Gram-negative), tryptophan 2,3-dioxygenase (TDO) Genes nwere ike ịgụnye 60 amino. mpaghara acid a na-ahụkarị na nje bacteria na-eri anụ na-adịghị. Enweghị mkpụrụ ndụ ihe nketa ndị ọzọ na-eri anụ na ASxL5T genome, gụnyere ndị na-etinye enzymes na-etinye aka na isoprenoid biosynthesis na ụzọ mevalonate. Rụba ama na ọnweghị mkpụrụ ndụ ihe nketa nke transcriptional gntR n'ime otu anụ anụ enyochala, mana enwere ike mata mkpụrụ ndụ ihe nketa gntR atọ na ASxL5T.
A na-achịkọta njirimara phenotypic nke ASxL5T na Tebụl 3 wee jiri ya tụnyere njirimara phenotypic nke genera 23, 24, 25, 26, na 27 metụtara na akwụkwọ. Ndị dịpụrụ adịpụ site na T. marinus, T. olevorans, B. sanyensis, na Oceanobacter kriegii na-arụ ọrụ, nnu na-anabata, oxidase-mma mkpanaka yiri ahụ, ma ọ fọrọ nke nta ka ọ bụrụ na ọ dịghị ihe ọzọ phenotypic e ji mara ASxL5T. Nkezi pH nke oké osimiri bụ 8.1 (https://ocean.si.edu/ocean-life/invertebrates/ocean-acidification#section_77), nke na-egosipụta na T. marinus, T. olevorans, B. sanyensis na O. kriegii. ASxL5T dabara maka oke pH buru ibu (4-9) ụdị ụdị ndị na-abụghị mmiri. Njirimara phenotypic nke Thalassolituus sp. C2-1. Amaghi ama. Okpomọkụ dị elu nke ASxL5T na-adịkarị karịa nke ụdị mmiri mmiri (4-42 Celsius C), n'agbanyeghị na ụfọdụ ma ọ bụghị naanị T. marinus dịpụrụ adịpụ na-anabata okpomọkụ. Enweghị ike itolite ASxL5T na mgbasa ozi broth gbochiri njiri mara phenotypic ọzọ. Jiri API 20E nwalee ihe ndị a chụpụrụ na efere BA, ONPG, arginine dihydrolase, lysine decarboxylase, ornithine decarboxylase, citrate utilization, urease, tryptophan deaminase, gelatin hydrolysis Enzyme, nsonaazụ ule niile adịghị mma, mana enweghị indole, acetoin na H2S. e mepụtara. Carbohydrates na-ekoghị eko gụnyere: glucose, mannose, inositol, sorbitol, rhamnose, sucrose, melibiose, amygdalin na arabinose. E jiri ya tụnyere ụdị ntụaka ndị metụtara ya e bipụtara, a na-egosi profaịlụ fatty acid cellular nke ụdị ASxL5T na Tebụl 4. Isi cellular fatty acids bụ C16:1ω6c na/ma ọ bụ C16:1ω7c, C16:0 na C18:1ω9. Hydroxy fatty acids C12:0 3-OH na C10:0 3-OH dịkwa. Oke C16:0 dị na ASxL5T dị elu karịa uru akọpụtara nke mkpụrụ ndụ ihe metụtara ya. N'ụzọ dị iche, ma e jiri ya tụnyere T. marinus IMCC1826TT a kọrọ, ọnụ ọgụgụ nke C18: 1ω7c na / ma ọ bụ C18: 1ω6c na ASxL5T na-ebelata. oleivorans MIL-1T na O. kriegii DSM 6294T, mana achọpụtaghị na B. sanyensis KCTC 32220T. N'ịtụle profaịlụ fatty acid nke ASxL5T na ASxLS gosipụtara ọdịiche dị nro na ọnụọgụ acid fatty acid dị n'etiti ụdị abụọ a, nke dabara na usoro DNA nke otu ụdị ahụ. Enweghị ihe ọ bụla poly-3-hydroxybutyrate (PHB) achọpụtara site na iji nwale ojii Sudan.
A mụọla ọrụ preation nke nje bacteria ASxL5T iji chọpụta oke anụ oriri. Nje bacteria a nwere ike ịmepụta plaques na ụdị Campylobacter, gụnyere: 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 na C. upsaliensis NCTC 11541T. Jiri omenaala ndị edepụtara na ngalaba nyocha nso nso nke usoro iji nwalee ụdị nje bacteria Gram-negative na gram-positive. Nsonaazụ gosiri na ASxL5T nwekwara ike iji Escherichia coli NCTC 86 na Citrobacter freundii NCTC 9750T. Plaques kpụrụ na Klebsiella oxytoca 11466. E gosipụtara mmekọrịta TEM na E. coli NCTC 86 na Figure 4A-D, yana mmekọrịta ya na Campylobacter jejuni PT14 na Campylobacter suis S12 gosipụtara na Figure 4E-H n'etiti. Usoro mwakpo ahụ yiri ka ọ dị iche n'etiti ụdị anụ a nwalere, yana otu mkpụrụ ndụ E. coli ma ọ bụ karịa etinyere na sel ASxL5T ọ bụla wee debe ya n'akụkụ cell gbatịrị agbatị tupu mgbasa. N'ụzọ dị iche, ASxL5T na-egosi na ọ na-ejikọta na Campylobacter site na otu ebe kọntaktị, na-emekarị na kọntaktị na apex nke cell anụ anụ na nso apex nke cell Campylobacter (Nyocha 4H).
TEM na-egosi mmekọrịta dị n'etiti ASx5LT na anụ anụ: (AD) na E. coli anụ anụ; (EH) na C. jejuni anụ oriri. (A) Otu mkpụrụ ndụ ASx5LT ejikọrọ na otu cell E. coli (EC); (B) A filamentous ASx5LT jikọtara na otu cell EC; (C) Mkpụrụ ndụ ASx5LT filamentous jikọtara na ọtụtụ mkpụrụ ndụ EC; (D) Mgbakwunye sel ASx5LT nta na otu cell E. coli (EC); (E) otu sel ASx5LT ejikọrọ na sel Campylobacter jejuni (CJ); (F) ASx5LT na-awakpo mkpụrụ ndụ C. hyointestinelis (CH); (G) abụọ Otu ASx5LT wakporo cell CJ; (H) Nlele dị nso nke ebe mgbakwunye ASx5LT, n'akụkụ ọnụ ọnụ nke cell CJ (ogwe 0.2 μm). Ogwe ahụ na-anọchi anya 1 μm na (A-G).
Nje bacteria na-eri anụ ewepụtala iji nweta ohere dị ukwuu nke anụ oriri. N'ụzọ doro anya, ha dị ebe niile n'ọtụtụ ebe dị iche iche. N'ihi nha dị warara nke ndị mmadụ, ọ ga-ekwe omume ikewapụ nje bacteria ASxL5T na slurry site na iji usoro nkewa nke phage. Mkpa genomic nke ASxL5T na ndị òtù ezinụlọ oceanospirillaceae nke nje bacteria na-eju anya, ọ bụ ezie na organism na-anabata nnu ma nwee ike itolite na ọkara nwere 5% nnu. Nyocha banyere ịdị mma mmiri nke slurry gosiri na ọdịnaya sodium chloride erughị 0.1%. Ya mere, apịtị dị anya na gburugburu mmiri - ma mpaghara ala ma na kemịkal. Ọnụnọ nke atọ ndị metụtara ma dị iche iche site n'otu ebe na-enye ihe akaebe na ndị anụ ọhịa ndị a na-eme nke ọma na gburugburu ebe a na-abụghị mmiri. Na mgbakwunye, nyocha microbiome (faịlụ data dị na https://www.ebi.ac.uk/ena/browser/view/PRJEB38990) gosiri na otu usoro mkpụrụ ndụ 16S rRNA dị na taxa 50 kachasị ukwuu (OTU) ) N'ime oge nlele ole na ole nke apịtị. Ahụrụ ọtụtụ nje bacteria na-enweghị omenala na nchekwa data Genbank, nke nwere usoro mkpụrụ ndụ 16S rRNA yiri nje bacteria ASxL5T. Usoro ndị a, yana usoro nke ASxL5T, ASxS5, na ASxO5, yiri ka ọ na-anọchi anya clades dị iche iche kewapụrụ na Thalassolituus na Oceanobacter (Nyocha 2). Ụdị nje bacteria atọ na-enweghị omenala (GQ921362, GQ921357 na GQ921396) dịpụrụ adịpụ na mmiri fissure na omimi nke 1.3 kilomita na ebe a na-egwupụta ọla edo South Africa na 2009, na abụọ ndị ọzọ (DQ256320 na DQ337006) sitere na mmiri dị na South Africa. n'afọ 2005). Usoro mkpụrụ ndụ ihe nketa 16S rRNA nwere njikọ chiri anya na ASxL5T bụ akụkụ nke usoro mkpụrụ ndụ 16S rRNA enwetara site na ọdịbendị ịba ụba nke sedimenti aja enwetara n'akụkụ osimiri nke ugwu France na 2006 (nọmba nnweta AM29240828). Usoro mkpụrụ ndụ ihe nketa 16S rRNA ọzọ nwere njikọ chiri anya sitere na nje bacteria HQ183822.1 a na-emebeghị eme ka enwetara site na tankị nnakọta nke ewepụtara na ebe mkpofu obodo na China. N'ụzọ doro anya, nje bacteria ASxL5T abụghị onye nnọchi anya nke ukwuu na ọdụ data taxonomic, mana usoro ndị a sitere na nje bacteria na-enweghị omenala nwere ike ịnọchite anya ihe dị ndụ yiri ASxL5T, nke a na-ekesa n'ụwa niile, na-emekarị na gburugburu ebe dị egwu. Site na nyocha genome phylogenetic dum, onye kacha nso na ASxL5T bụ Thalassolituus sp. C2-1, T. marinus, T. oleivorans. Na O. kriegii 23, 24, 25, 26, 27. Thalassolituus bụ onye so na mmiri na-amanye hydrocarbon fragmentation bacteria (OHCB), nke bụ ebe nile na mmiri na terrestrial gburugburu, na-emekarị na-aghọ ndị na-achị mgbe hydrocarbon mmetọ events30,31. Nje bacteria na mmiri abụghị ndị otu OHCB, kama ọ dịpụrụ adịpụ na gburugburu mmiri.
Data phenotypic na-egosi na ASxL5T bụ ụdị ọhụrụ na onye so n'otu ụdị nke a na-amabughị na ezinụlọ spirospiraceae mmiri. Ugbu a enweghị ọkọlọtọ doro anya iji kewaa ụdị dịpụrụ adịpụ ọhụrụ ka ọ bụrụ ụdị ọhụrụ. Emeela mgbalị iji chọpụta ókèala genera zuru ụwa ọnụ, dịka ọmụmaatụ, dabere na pasent nke genome nke protein conservative (POCP), a na-atụ aro na ọnụ ahịa mbelata bụ 50% yiri nke strain33. Ndị ọzọ na-atụ aro iji ụkpụrụ AAI, nke nwere uru karịa POCP n'ihi na enwere ike nweta ha site na genomes34 ezughị ezu. Onye edemede ahụ kwenyere na ọ bụrụ na uru AAI na-erughị 74% ma e jiri ya tụnyere ụdị ụdị nke ụdị ihe nlereanya ahụ, eriri ahụ bụ ihe nnọchianya nke ụdị dị iche iche. Ụdị ụdị dị na mmiri spirillaceae bụ mmiri spirillum, na ụdị ụdị bụ O. linum ATCC 11336T. Uru AAI n'etiti ASxL5T na O. linum ATCC 11336T bụ 54.34%, na uru AAI n'etiti ASxL5T na T. oleivorans MIL-1T (ụdị ụdị ụdị) bụ 67.61%, na-egosi na ASxL5T na-anọchite anya ụdị ọhụrụ dị iche na Thalassolituus. Iji usoro mkpụrụ ndụ ihe nketa 16S rRNA dị ka ọkọlọtọ nhazi, oke mmachi genus tụrụ aro bụ 94.5%35. Enwere ike itinye ASxL5T na ụdị Thalassolituus, na-egosi njirimara usoro 95.03% 16S rRNA na T. oleivorans MIL-1T na 96.17%. marinus IMCC1826T. Otú ọ dị, a ga-etinyekwa ya na ụdị Bacteroides nke nwere 94.64% 16S rRNA geneidentity na B. sanyensis NV9, na-egosi na iji otu mkpụrụ ndụ ihe dị ka 16S rRNA gene nwere ike iduga na nhazi na ọrụ aka ike. Ụzọ ọzọ a tụrụ aro na-eji ANI na Genome Alignment Score (AF) iji nyochaa nchịkọta nke data sitere na ụdị niile na ụdị na-abụghị ụdị nke genera dị adị. Onye edemede ahụ na-atụ aro ka ijikọta ókèala genus na ntụgharị ntụgharị nke ọdịdị nke a na-eme atụmatụ kpọmkwem maka taxa a na-enyocha. Agbanyeghị, ọ bụrụ na ezughị usoro genome zuru oke sitere na Thalassolituus iche, ọ gaghị ekwe omume ịchọpụta ma ASxL5T ọ bụ nke Thalassolituus genus site na usoro a. N'ihi obere ohere nke usoro genome zuru oke maka nyocha, a ga-eji nlezianya kọwaa osisi genome phylogenetic dum. Nke abuo, usoro ntụnyere genome dum enweghị ike ịkọwapụta nnukwu ọdịiche na nha genome atụnyere. Ha tụlere myirịta nke mkpụrụ ndụ ihe nketa echekwabara n'etiti mkpụrụ ndụ ihe nketa metụtara ya, mana ha eburughị n'uche ọnụọgụ mkpụrụ ndụ ihe nketa na-anọghị na genome dị ntakịrị nke ASxL5T. N'ụzọ doro anya, ASxL5T na ndị otu gụnyere Thalassolituus, Oceanobacter, na Bacterioplanes nwere otu nna ochie, mana evolushọn ewerela ụzọ dị iche iche, na-eduga ná mbelata nke genome, nke nwere ike ime mgbanwe na ndụ anụ ọhịa. Nke a dị iche na T. oleivorans MIL-1T, nke bụ 28% ibu ma malite n'okpuru nrụgide gburugburu ebe obibi dị iche iche iji jiri hydrocarbons23,30. Enwere ike iji ntụnyere na-adọrọ mmasị na parasaiti intracellular na symbionts dị ka Rickettsia, Chlamydia, na Buchnera. Ogo genome ha dị ihe dịka 1 Mb. Ikike iji metabolites sel ndị ọbịa na-eduga na mfu mkpụrụ ndụ ihe nketa, yabụ enwere mmebi nke mkpụrụ ndụ mkpụrụ ndụ ihe dị ịrịba ama. Mgbanwe evolushọn sitere na ntule nri kemịkalụ mmiri gaa n'ụdị ndụ anụ ọhịa nwere ike bute mbelata otu ahụ na nha genome. Nyocha COG na KEGG na-eme ka ọnụọgụ mkpụrụ ndụ ihe nketa na-arụ ọrụ dị iche iche na ọdịiche dị n'ụwa niile na ụzọ genomic n'etiti ASxL5T na T. oleivorans MIL-1T, nke na-abụghị n'ihi na enwere ike ịnweta ihe mkpụrụ ndụ ihe nketa mobile. Ọdịiche dị na nha G + C nke genome dum nke ASxL5T bụ 56.1%, na nke T. oleivorans MIL-1T bụ 46.6%, nke na-egosikwa na e kewapụrụ ya.
Nyocha nke ọdịnaya nzuzo nke ASxL5T genome na-enye nghọta na-arụ ọrụ n'ime njirimara phenotypic. Ọnụnọ nke mkpụrụ ndụ ihe nketa nke ụdị IV fimbriae (Tfp) bụ ihe na-adọrọ mmasị karịsịa n'ihi na ha na-akwalite mmegharị nke cell, nke a na-akpọ gliding ma ọ bụ mgbagwoju anya, na-enweghị flagella n'elu. Dị ka akụkọ si kwuo, Tfp nwere ọrụ ndị ọzọ, gụnyere predation, pathogenesis, nhazi biofilm, DNA eke eke, nchịkọta cell akpaka na mmepe38. Mkpụrụ ndụ ASxL5T nwere mkpụrụ ndụ ihe nketa iri na asatọ nke na-edobe diguanylate cyclase (enzyme nke na-eme ka ntụgharị nke 2 guanosine triphosphate banye guanosine 2 phosphate na cyclic diGMP) na mkpụrụ ndụ ihe nketa 6 na-etinye diguanylate cyclase phosphate diguanylate kwekọrọ. Mkpụrụ ndụ maka esterase (na-eme ka mmebi nke cyclic di-GMP gaa guanosine monophosphate) bụ ihe na-adọrọ mmasị n'ihi na cycl-di-GMP bụ ozi nke abụọ dị mkpa na-etinye aka na mmepe biofilm na nkewa, mmegharị, ntinye cell na virulence 39, 40 na usoro ahụ. Okwesiri iburu n'uche na na Bdellovibrio bacteriovorus, egosila cyclic okpukpu abụọ GMP iji chịkwaa mgbanwe n'etiti ndụ efu na ụdị ndụ anụ ọhịa41.
Ọtụtụ nchọpụta banyere nje bacteria na-eri anụ lekwasịrị anya na Bdellovibrio, Bdellovibrio-dị ka ntule, na ụdị Myxococcus. Ihe atụ ndị a na ndị ọzọ a ma ama nke nje bacteria na-eri anụ na-etolite otu dị iche iche. N'agbanyeghị ụdị dị iche iche a, a chọpụtala otu ezinụlọ protein e ji mara nke na-egosipụta phenotypes nke nje bacteria iri na otu a ma ama3,22. Agbanyeghị, ọ bụ naanị mkpụrụ ndụ ihe nketa na-etinye O antigen ligase (waaL) ka achọpụtara, nke a na-ahụkarị na nje bacteria Gram-negative. Ụdị nyocha a adịghị enye aka n'ịkọpụta ASxL5T dị ka onye na-eri anụ, eleghị anya n'ihi na ọ na-eji atụmatụ ọgụ ọhụrụ. Ịnweta ụdị mkpụrụ ndụ nje na-eri anụ dị iche iche ga-enyere aka ịmepụta nyocha mkpebi dị mma nke na-eburu n'uche ihe akaebe nke ọrụ na esemokwu gburugburu ebe obibi n'etiti ndị otu. Ihe atụ nke nje bacteria na-eri anụ adịghị etinye na nyocha a gụnyere ndị òtù Cupriavidus necator42 na Bradymonabacteria43, n'ihi na ka ndị nchọpụta na-enyocha obodo dị iche iche microbial, a na-ewepụtakwu ụtụ isi anụ.
Akụkụ kachasị ama nke nje bacteria ASxL5T nke TEM weghaara bụ ọdịdị ọdịdị ya pụrụ iche na nke na-agbanwe agbanwe, nke nwere ike ịkwalite mmekọrịta ya na nje bacteria na-eri anụ. Ụdị mmekọrịta a hụrụ dị iche na nje bacteria ndị ọzọ na-eri anụ, a chọpụtabeghị ma ọ bụ kọọ ya na mbụ. A na-egosi usoro ndụ anụ ọhịa ASxL5T nke a tụrụ aro na eserese 5. Enwere ihe atụ ole na ole n'ime akwụkwọ ndị nwere ụdị apical yiri nke a ka anyị na-akọ ebe a, mana ihe atụ ndị a gụnyere Terasakiispira papahanaumokuakeensis, nje nje spirillum nke mmiri na-ebuwanye ibu 44, na Alphaproteobacteria, Terasakiella pus. , nke bụbu nke genus Oceanospirillum, na-egosipụta ihe a na-akpọ "ihe nkiri polar" 45. A na-ahụkarị ụdị Cocci na omenala ochie, karịsịa maka nje bacteria nwere ụdị agbagọ, dị ka Vibrio, Campylobacter, na Helicobacter 46, 47, 48, nke nwere ike ịnọchite anya ala dara ada. A chọrọ ọrụ ọzọ iji dokwuo anya usoro ndụ nje bacteria ASxL5T. Iji chọpụta ka o si ejide ma na-eri anụ, yana ma mkpụrụ ndụ ihe nketa ya na-edobe ogige ndị na-arụ ọrụ ndụ nke enwere ike iji mee ihe maka ahụike ma ọ bụ teknụzụ biotechnological.
Nkọwa nke Venatorbacter gen. Novemba Venatorbacter (Ven.a.tor, ba'c.ter, L. bụ nke ndị na-eri nri sitere na L. n. venator,' hunter' na Gr. n. bacter,' mkpanaka '. Venatorbacter,' mkpanaka ịchụ nta' Sel bụ aerobic, nnu-na-anabata, curved Gram ntụpọ, ọrụ oxidase adịghị amụba Okpomọkụ nke 4 ruo 42 Celsius C na-etolite n'ụdị pH nke 4-9 bụ ihe a na-adịghị ahụkebe na pH nke acid acid. C18: 1ω9 ; C12: 0 3-OH na C10: 0 3-OH dị ka abụba hydroxy Acids. Ha anaghị eto na mgbasa ozi broth DNA G + C bụ 56.1 mol .
Nkọwa nke Venatorbacter cucullus sp. November Venatorbacter cucullus (cu'cull.us.; L. n. cucullus pụtara fairing).
Na mgbakwunye, njirimara nkọwa nke ụdị ndụ a bụ na mgbe a na-etolite na BA ma ọ bụ BHI, mkpụrụ ndụ dị 1.63 µm ogologo na 0.37 µm n'obosara. Mpaghara ndị dị na BHI agar dị obere, na-eru 2 mm na dayameta mgbe awa 72 gachara. Ha bụ beige, translucent, gburugburu, convex na onwunwu. Ndị otu ụdị a nwere ike iji Escherichia coli na Klebsiella. Campylobacter na ọtụtụ nje bacteria gram-adịghị mma na-eje ozi dị ka anụ oriri.
Ụdị ụdị ASxL5T dịpụrụ adịpụ na mmiri ara ehi beef na Nottinghamshire, UK, ma debe ya na National Ụdị Culture Collection (UK): ntinye nọmba NCTC 14397 na Netherlands Bacterial Culture Collection (NCCB) ntinye nọmba NCCB 100775. Usoro genome zuru ezu nke ASxL5T etinyela ya na Genbank dịka mgbakwunye CP046056 si dị.
Ekewapụrụ nje bacteria ASxL5T na mmiri ara ehi site na iji teknụzụ kewapụrụ phage9,49. A na-agbaze slurry 1: 9 (w / v) na SM buffer (50 mM Tris-HCl [pH 7.5], 0.1 M NaCl, 8 mM MgSO4.7H2O na 0.01% gelatin; Sigma Aldrich, Gillingham, UK), wee tinye ya. na 4 Celsius C maka awa 24, na-atụgharị nwayọọ nwayọọ ka ndị na-eri anụ banye n'ime nchekwa. A na-etinye nkwusioru ahụ na 3000g maka nkeji 3. A na-anakọta supernatant na centrifud na 13,000g maka oge nke abụọ maka nkeji ise. A na-agafe ihe dị elu site na nzacha akpụkpọ anụ 0.45 µm (Minisart; Sartorius, Gottingen, Germany) na 0.2 µm membrane filter (Minisart) iji wepụ mkpụrụ ndụ nje ọ bụla fọdụrụnụ. ASxL5T nwere ike ịgafe nzacha ndị a. A na-edozi ahịhịa agar dị nro nke Campylobacter enterosus S12 (Nchịkọta NCBI CP040464) site na otu slurry ahụ site na iji usoro ọkọlọtọ. A na-ekesa slurry a kpochara na nke ọ bụla n'ime efere cell ndị ọbịa na 10 µl ụmụ irighiri mmiri dị okpukpu atọ ma hapụ ya ka ọ kpọọ nkụ. A na-etinye efere ahụ na tank microaerophilic na 37 ° C maka awa 48 n'okpuru ọnọdụ microaerobic (5% O2, 5% H2, 10% CO2, na 80% N2). E weputara akara ngosi a na-ahụ anya n'ime ihe nchekwa SM wee bufee ya na ahịhịa ọhụrụ nke C. hyointestinalis S12 ka ọ gbasaa n'ihu n'ime ihe ndị ahụ lysed. Ozugbo e kpebisiri ike na nje bacteria bụ ihe na-akpata lytic plaque ọ bụghị phage, gbalịa itolite organism n'adabereghị na onye ọbịa ma kọwaa ya ọzọ. Emere omenala ikuku ikuku na 37 Celsius C na 5% v / v defibrinated ọbara ịnyịnya (TCS Biosciences Lt, Buckingham, UK, mgbakwunye). Dị ka ntụziaka nke National Clinical Standards Committee si kwuo, a na-eji usoro mgbasa ozi diski eme ihe maka nyocha nke nje bacteria. Azụlitere BHI agar na 37 Celsius C site na iji diski nwere ọgwụ nje ndị a (Oxoid) maka ọdịbendị aerobic: amoxicillin na clavulanic acid 30 µg; cefotaxime 30 µg; streptomycin 10 g; ciprofloxacin 5 µg; Ceftazidime 30 µg Nalidixic acid 30 µg; Imipenem 10 µg; Azithromycin 15 µg; chloramphenicol 30 g; Cefoxitin 30 g; Tetracycline 30 g; nitrofurantoin 300 g; Aztreonam 30 µg; Ampicillin 10 g; Cefpodoxime 10 µg; Trimethoprim-Sulfamethoxazole 25 μg. Emebere nnabata nnu site na incubation aerobic na efere BHI agar na 37 Celsius. Agbakwunyere NaCl agbakwunyere na efere agar BHI iji nye oke ịta ahụhụ ruru 10% w/v. A na-ekpebi oke pH site na omenala ikuku na efere BHI agar na 37 Celsius C, ebe agbanweela pH dị n'etiti 4 na 9 nke nwere HCl na-adịghị mma ma ọ bụ NaOH na-adịghị mma, a na-achọpụtakwa uru pH ezubere tupu ịwụsa efere ahụ. Maka nyocha cellular fatty acid, ASxL5T ka emebere na BHI agar maka ụbọchị 3 yana ikuku ikuku na 37 Celsius. Dịka MIDI (Sherlock Microbial Identification System, ụdị 6.10) ọkọlọtọ ọkọlọtọ nke FERA Science Ltd, (York, UK) si kwuo, ewepụtara, kwadebere na nyochara acid fatty cell.
Maka TEM, ASxL5T azụlitere ikuku ikuku site n'ịgbasa n'otu n'otu na BA na 37 ° C maka awa 24, wee weta ya n'ime 1 ml nke 3% (v/v) glutaraldehyde na 0.1 M cacodylate buffer na ụlọ okpomọkụ Fix maka 1 hour, wee centrifuge. na 10,000 g maka nkeji 3. Mgbe ahụ jiri nwayọọ maliteghachi pellet ahụ na 600 μl 0.1 M cacodylate nchekwa. Nyefee nkwusioru ASxL5T etinyere na ihe nkiri Formvar/carbon na grid ọla kọpa 200 mesh. Ejiri 0.5% (w/v) uranyl acetate mee nje bacteria ahụ maka nkeji 1 wee nyochaa ya site na TEM site na iji TEI Tecnai G2 12 Biotwin microscope. Dị ka e kwuru n'elu, jikọta otu ọnụ ọgụgụ anụ anụ na anụ anụ na NZCYM broth (BD Difco ™, Fisher Scientific UK Ltd, Loughborough) na incubate maka awa 48 n'okpuru ọnọdụ microaerobic nke Campylobacter ma ọ bụ Campylobacter na 37 ° C, Mmekọrịta nke anụ anụ na anụ anụ. TEM nyochakwara ya. Ọnọdụ aerobic maka Escherichia coli. Nyochaa onwe gị na-enyocha anụ oriri na nje bacteria na-eri anụ iji chọpụta mgbanwe ọ bụla na cell morphology n'ihi anụ anụ. Ejiri usoro ojii Sudan maka nyocha anya nke mkpokọta PHB.
Too omenala ASxL5T n'abali site na iji swab na-adịghị mma na-eto eto na BHI ma ọ bụ efere BA. Chịkọta mkpụrụ ndụ ASxL5T ma kwụsịtụ na MRD (CM0733, Oxoid), wee debe ha na 4°C maka ụbọchị 7 ka mkpụrụ ndụ gụchaa agụụ. A na-etinye akwụkwọ ntụaka NCTC ma ọ bụ omenala nje bacteria n'ime broth BHI ma ọ bụ No. 2 nutrient broth (CM007, Oxoid), tinye ya n'otu abalị, centrifuged na 13,000g wee maliteghachi na MRD ruo mgbe OD600 bụ 0.4. Omenala: Bacillus subtilis NCTC 3610T, Citrobacter freundii NCTC 9750T, Enterobacter aerogenes NCTC 10006T, Enterococcus faecalis NCTC 775T, Escherichia coli NCTC 86, Klebsiella 4conostu NCTC 10817, Listeria pụrụ iche nje NCTC 4885, Bacillus macerans NCTC 6355T, Providencia stuartsii NCTC 10318, Pseudomonas fluorescens SMDL, Rhodococcus submarine hamburger NCTC 1621T, Salmonella NCTC intestinal. 10861, Staphylococcus aureus NCTC 8532T, Streptococcus pneumoniae NCTC 7465T, Yersinia enterocolitica NCTC 10460. Onye na-elekọta Campylobacter bụ nke a na-etinye microaerobically na efere BA na 37 ° C ma kwụsịtụrụ na broth N.Z. Ndị ọbịa Campylobacter a nwalere bụ: 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, C... Na-anakọta mkpụrụ ndụ na MRD, centrifuge na 13,000g wee maliteghachi na MRD ruo mgbe OD600 bụ 0.4. Tinye aliquot nke 0.5 ml nkwusioru na 5 ml gbazere NZCYM top agar (0.6% agar) ma wunye ya na 1.2% NZCYM efere ala. Mgbe a gwọchara na ihicha, ekesara ASxL5T a gbanyere mkpọrọgwụ ka ọ bụrụ 20µl ụmụ irighiri mmiri na bọọdụ ahịhịa ọ bụla n'ime atọ. Okpomọkụ omenala na ikuku na-adabere n'ihe achọrọ nke nje bacteria.
Jiri GenElute™ Bacterial Genomic DNA Kit (Sigma Aldridge) kwado DNA site na mpụ nje. Eji usoro ọkọlọtọ mee ihe maka mmụba PCR nke 16S rRNA gene na mkpebi n'usoro ngwaahịa site na iji kemịka ịcha agba (Eurofins Value Read Service, Germany). Jiri mmemme BLAST-N tulee usoro ndị a na usoro mkpụrụ ndụ 16S rRNA ndị ọzọ iji chọpụta na ịnakọta ụdị nwere njikọ chiri anya. A kwadoro ihe ndị a site na iji ClustalW na mmemme MEGA X. Emegharịrị osisi phylogenetic site na iji MEGA X site na iji usoro ohere kachasị dabere na ụdị Tamura-Nei, yana mbipụta 1000 eduzi54. Jiri PureLink™ Genomic DNA Kit (Fisher Scientific, Loughborough, UK) wepụ DNA maka usoro genome dum. Ekpebiri usoro genome nke ASxL5T site na iji nchikota Illumina MiSeq, nke nwere 250 bp nwere ngwụcha ngwụcha nke mejupụtara ọbá akwụkwọ a kwadebere site na iji ngwa akara Nextera na 2 ruo 20 kb ogologo na-agụ site na ikpo okwu PacBio. Genomics DNA Sequencing Research Facility na Mahadum Sembia. Ejikọtara genome ahụ site na iji CLC Genomics Workbench 12.0.3 (Qiagen, Aarhus, Denmark). A na-edobe omenala ASxL5T na National Ụdị Culture Collection (UK) na Netherlands Bacterial Culture Collection (NCCB). Mkpụrụ ndụ ihe nketa nke ihe ndị metụtara ya eji atụnyere ya bụ: Thalassolituus oleivorans MIL-1T (ọnụọgụ nnweta HF680312, zuru ezu); Bacterioplanes sanyensis KCTC 32220T (Nnweta ọnụọgụY01000001, ezughi oke); Oceanobacter kriegii DSM 6294T (nọmba nnweta NZ_AUGV00000000, ezughị ezu); Obodo Marinamonas DSM 5604T (agbakwunyere ASM436330v1, ezughị ezu), Oceanospirullum linum ATCC 11336T (agbakwunyere MTSD02000001, ezughị ezu) na Thalassolituus sp. C2-1 (tinye NZ_VNIL01000001, ezughị ezu). Jiri JGI Genome Portal36 na https://img.jgi.doe.gov//cgi-bin/mer/main.cgi?section=ANI&page= iji chọpụta akara nzizi (AF) na nkezi njirimara nucleic acid (ANI). Na ụzọ abụọ. Ejiri usoro Rodriguez-R & Konstantinidis55 chọpụta njirimara amino acid (AAI). Jiri GToTree 1.5.5411,12,13,14,15,16,17,18 wepụta osisi phylogenetic a na-eme atụmatụ kachasị. A na-ahọrọ genome ntinye nke na-anọchite anya genome ntụaka dị site na nrụtụ aka nke achọpụtara dị ka ihe metụtara ASxL5T sitere na 16S rRNA phylogeny. Edere osisi ahụ site na iji ngwa ọrụ ịntanetị na-emekọrịta ihe (https://itol.embl.de/). A na-eme nkọwa ọrụ na nyocha nke genome ASxL5T site na iji ngwa ịntanetị BlastKOALA KEGG site na iji KEGG (Kyoto Encyclopedia of Genes and Genomes) modul nkesa nkwalite. A na-ekpebi nkesa nke ngalaba COG (otu orthologues) site na iji ngwa ịntanetị eggNOG-mapper.
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