Ib hom tshiab ntawm Gram-negative, aerobic, ntsev-tiv taus, nquag, qws-zoo li tus, thiab cov kab mob predatory ASxL5T tau cais tawm ntawm lub pas dej nyuj nyuj hauv Nottinghamshire, England, thiab siv Campylobacter ua nws cov tsiaj. Tom qab ntawd, lwm hom Campylobacter thiab cov tswv cuab ntawm tsev neeg Enterobacteriaceae tau tshawb pom raws li cov tsiaj txhu. Tom qab subculture yam tsis muaj cov hlwb, qhov kev loj hlob tsis muaj zog aseptic tau ua tiav ntawm Lub Plawv Lub Plawv Infusion Agar. Qhov kev loj hlob zoo tshaj plaws yog 37 ° C thiab pH yog 7. Kev xa hluav taws xob hluav taws xob microscopy qhia qee qhov txawv txav morphological yam ntxwv ntsig txog qhov muaj cov tsiaj txhu. Phylogenetic tsom xam siv 16S rRNA gene sequence tau qhia tias kev cais tawm muaj feem cuam tshuam nrog ib tus tswv cuab ntawm Marine Spirulina tsev neeg, tab sis tsis tuaj yeem raug cais kom meej tias yog ib tus tswv cuab ntawm cov neeg paub. Tag nrho-genome sequencing ntawm ASxL5T tau lees paub qhov kev sib raug zoo nrog cov tswv cuab ntawm marine spirochetes. Kev tshawb nrhiav cov ntaub ntawv tau qhia tias ob peb ASxL5Ts qhia 16S rRNA gene sequences nrog ob peb cov kab mob tsis zoo los ntawm dej hiav txwv, thaj av thiab dej hauv av. Peb pom zoo tias hom ASxL5T sawv cev rau hom tshiab hauv cov genus tshiab. Peb pom zoo lub npe Venatorbacter cucullus gen. Nov, sp. Thaum lub Kaum Ib Hlis, ASxL5T tau siv los ua hom kab mob.
Cov kab mob Predatory yog cov kab mob uas muaj peev xwm tua tau thiab tua lwm cov kab mob uas muaj sia nyob kom tau txais cov ntaub ntawv biosynthetic thiab lub zog. Qhov no yog qhov txawv ntawm kev rov qab los ntawm cov as-ham los ntawm cov kab mob tuag, thiab nws kuj yog qhov sib txawv ntawm cov kab mob parasitic, uas cov kab mob tsim kev sib raug zoo nrog lawv tus tswv tsev yam tsis tua lawv. Cov kab mob Predatory tau hloov pauv lub neej sib txawv kom tau txais txiaj ntsig zoo ntawm cov khoom noj muaj ntau hauv cov niches uas lawv pom (xws li cov chaw nyob hauv hiav txwv). Lawv yog ib pawg neeg muaj ntau haiv neeg, uas tsuas yog txuas nrog lawv lub neej ua kom tsis muaj menyuam 1. Piv txwv ntawm cov kab mob predatory tau pom nyob rau hauv ntau yam phyla, xws li: Proteobacteria, Bacteroides, thiab Chlorella.3. Txawm li cas los xij, cov kab mob uas tau kawm zoo tshaj plaws yog Bdellovibrio thiab Bdellovibrio-thiab-zoo li kab mob (BALOs4). Cov kab mob Predatory yog qhov muaj txiaj ntsig zoo ntawm cov tshuaj lom neeg lom neeg tshiab thiab cov tshuaj tua kab mob 5.
Cov kab mob Predatory tau ntseeg tias yuav txhim kho microbial ntau haiv neeg thiab muaj kev cuam tshuam zoo rau kev noj qab haus huv, kev tsim khoom thiab kev ruaj ntseg6. Txawm hais tias muaj cov cwj pwm zoo no, muaj qee qhov kev tshawb fawb txog cov kab mob predatory tshiab vim qhov nyuaj ntawm cov kab mob culturing thiab yuav tsum tau ua tib zoo saib xyuas kev sib cuam tshuam ntawm tes kom nkag siab txog lawv txoj kev ua neej nyuaj. Cov ntaub ntawv no tsis yooj yim kom tau los ntawm kev txheeb xyuas lub computer.
Nyob rau hauv ib lub sijhawm ntawm kev tiv thaiv kab mob tiv thaiv kab mob, cov tswv yim tshiab rau kev tsom cov kab mob kab mob tau raug kawm, xws li kev siv cov kab mob bacteriophages thiab cov kab mob predatory7,8. Cov kab mob ASxL5T tau raug cais tawm hauv 2019 siv cov tshuab phage cais tawm los ntawm cov quav nyuj uas tau sau los ntawm Lub Chaw Dairy ntawm University of Nottingham, Nottinghamshire. Lub hom phiaj ntawm kev tshawb nrhiav yog cais cov kab mob uas muaj peev xwm ua cov tshuaj tiv thaiv kab mob lom. Campylobacter hyointestinalis yog ib hom kab mob zoonotic, uas muaj feem cuam tshuam nrog tib neeg txoj hnyuv kab mob10. Nws yog ubiquitous nyob rau hauv cov ntshav thiab siv raws li lub hom phiaj host.
Cov kab mob ASxL5T tau raug cais tawm ntawm cov nqaij nyug jelly vim nws tau pom tias cov plaques nws tsim rau ntawm cov nyom ntawm C. hyointestinalis zoo ib yam li cov khoom tsim los ntawm cov kab mob bacteriophages. Qhov no yog qhov kev tshawb nrhiav tsis tau xav txog, vim hais tias ib feem ntawm cov txheej txheem phage cais tawm cuam tshuam nrog kev lim dej los ntawm 0.2 µm lim, uas yog tsim los tshem tawm cov kab mob. Kev tshuaj xyuas microscopic ntawm cov khoom muab rho tawm los ntawm cov quav hniav qhia tau hais tias cov kab mob gram-negative curved rod-zoo li tus kab mob tsis tuaj yeem khaws cov polyhydroxybutyrate (PHB). Cov kab lis kev cai aseptic ywj pheej ntawm cov tsiaj nyeg yog paub txog cov khoom nruab nrab nplua nuj (xws li lub paj hlwb infusion agar (BHI) thiab ntshav agar (BA)), thiab nws txoj kev loj hlob tsis muaj zog. Nws yog tau tom qab subculture nrog hnyav inoculum txhim kho. Nws loj hlob zoo sib xws nyob rau hauv microaerobic (7% v / v oxygen) thiab atmospheric oxygen, tab sis tsis nyob rau hauv ib qho chaw anaerobic. Tom qab 72 teev, txoj kab uas hla ntawm lub colony yog me me, ncav cuag 2 hli, thiab nws yog beige, translucent, puag ncig, convex thiab ci iab. Kev ntsuam xyuas biochemical Standard yog hampered vim ASxL5T tsis tuaj yeem muaj kev ntseeg siab hauv cov kua xov xwm, uas qhia tias nws yuav vam khom lub neej voj voog ntawm kev tsim biofilm. Txawm li cas los xij, lub phaj ncua kev kawm tau pom tias ASxL5T yog aerobic, zoo rau oxidase thiab catalase, thiab tuaj yeem zam 5% NaCl. ASxL5T yog resistant rau 10 µg streptomycin, tab sis rhiab rau tag nrho lwm cov tshuaj tua kab mob. Cov kab mob ASxL5T tau kuaj xyuas los ntawm TEM (Daim duab 1). Thaum loj hlob tsis muaj prey hlwb ntawm BA, ASxL5T hlwb yog me me Campylobacter, nrog qhov nruab nrab ntev ntawm 1.63 μm (± 0.4), qhov dav ntawm 0.37 μm (± 0.08), thiab ib qho ntev (txog 5 μm) ncej. Kev sib deev flagella. Kwv yees li ntawm 1.6% ntawm cov hlwb tshwm sim kom muaj qhov dav ntawm tsawg dua 0.2 μm, uas yuav tso cai rau kev nkag mus los ntawm cov cuab yeej lim. Ib qho kev nthuav dav txawv txawv tau pom nyob rau sab saum toj ntawm qee lub hlwb, zoo ib yam li kev ncaj ncees (Latin cucullus) (saib cov xub hauv 1D, E, G). Qhov no zoo li yog tsim los ntawm cov txheej txheem sab nrauv, uas tej zaum yuav yog vim qhov txo qis ntawm qhov loj ntawm lub hnab ntawv periplasmic, thaum lub hnab ntawv sab nrauv tseem tsis zoo, uas qhia tau tias "xoo" zoo li. Culturing ASxL5T nyob rau hauv qhov tsis muaj cov as-ham (hauv PBS) ntev ntawm 4 ° C ua rau feem ntau (tab sis tsis yog tag nrho) cov hlwb uas qhia txog coccal morphology (Daim duab 1C). Thaum ASxL5T loj hlob nrog Campylobacter jejuni raws li cov neeg raug tsim txom rau 48 teev, qhov nruab nrab ntawm tes yog qhov ntev thiab nqaim dua li cov hlwb loj hlob yam tsis muaj tus tswv (Table 1 thiab Daim duab 1E). Nyob rau hauv sib piv, thaum ASxL5T loj hlob nrog E. coli raws li prey rau 48 teev, qhov nruab nrab cell loj yog ntev thiab dav dua li thaum nws loj hlob tsis muaj prey (Table 1), thiab lub cell ntev yog variable, feem ntau qhia filamentous (Daim duab 1F). Thaum incubated nrog Campylobacter jejuni los yog E. coli raws li prey rau 48 teev, ASxL5T hlwb pom tsis muaj flagella txhua. Table 1 piav qhia txog cov kev soj ntsuam ntawm cov kev hloov pauv ntawm cov xov tooj ntawm tes raws li qhov muaj, tsis muaj, thiab raug ntaus hom ASxL5T.
TEM zaub ntawm ASx5LT: (A) ASx5LT qhia ntev nplawm; (B) raug ASx5LT roj teeb; (C) cocci ASx5LT hlwb tom qab ntev incubation tsis muaj as-ham; (D) ib pab pawg ntawm ASx5LT hlwb qhia qhov txawv txav (E) ASx5LT cell pab pawg incubated nrog Campylobacter prey pom cov cell ntev ntev piv nrog cov tsis muaj kev loj hlob prey (D) kuj pom cov qauv apical; (F) Loj Filamentous flagella, ASx5LT hlwb, tom qab incubation nrog E. coli prey; (G) Ib leeg ASx5LT cell tom qab incubation nrog E. coli, uas qhia ib tug txawv txawv sab saum toj qauv. Lub bar sawv cev 1 μm.
Kev txiav txim siab 16S rRNA gene sequence (accession naj npawb MT636545.1) ua rau cov ntaub ntawv tshawb fawb los tsim cov kab ke zoo ib yam li cov hauv chav kawm Gammaproteobacteria, thiab ze tshaj rau cov kab mob marine hauv marine spirillum tsev neeg (Daim duab 2), thiab yog cov tswv cuab ntawm Thalasussolituus. Tus txheeb ze ze tshaj plaws rau Marine Bacillus. 16S rRNA gene sequence yog txawv los ntawm cov kab mob predatory uas yog cov Bdelvibrionaceae (Deltaproteobacteria) tsev neeg. Cov kev sib piv ntawm cov sib piv ntawm B. bacteriovorus HD100T (hom strain, DSM 50701) thiab B. bacteriovorus DM11A yog 48.4% thiab 47.7%, thiab rau B. exovorus JSS nws yog 46.7%. Cov kab mob ASxL5T muaj 3 daim ntawv luam ntawm 16S rRNA noob, ob qho tib si zoo ib yam, thiab qhov thib peb yog 3 lub hauv paus sib nrug. Ob tug lwm yam kab mob predatory cais tawm (ASx5S thiab ASx5O; 16S rRNA gene accession naj npawb yog MT636546.1 thiab MT636547.1, raws li) nrog cov yam ntxwv zoo sib xws morphological thiab phenotypic los ntawm tib qhov chaw tsis zoo ib yam, tab sis lawv txawv ntawm ASxtured Bacterial thiab unculate. database sequences yog clustered ua ke nrog rau lwm yam genera hauv Oceanospirillaceae (Daim duab 2). Tag nrho cov genome ib ntus ntawm ASxL5T tau txiav txim siab thiab khaws cia hauv NCBI database, thiab tus lej nkag yog CP046056. Lub genome ntawm ASxL5T muaj ib ncig chromosome ntawm 2,831,152 bp nrog G + C piv ntawm 56.1%. Cov kab ke genome muaj 2653 CDS (tag nrho), ntawm 2567 tau kwv yees rau encode proteins, ntawm 1596 tuaj yeem muab ua haujlwm ua haujlwm (60.2%). Lub genome muaj 67 RNA-coding noob, suav nrog 9 rRNAs (3 txhua rau 5S, 16S, thiab 23S) thiab 57 tRNAs. Cov yam ntxwv genomic ntawm ASxL5T tau muab piv nrog cov genomes ntawm cov kab mob ntawm cov txheeb ze ze tshaj plaws uas tau txheeb xyuas los ntawm 16S rRNA gene sequence (Table 2). Siv cov amino acid tus kheej (AAI) los sib piv tag nrho cov muaj Thalassolituus genomes rau ASxL5T. Qhov ze tshaj plaws muaj (tsis tiav) genome ib ntus txiav txim siab los ntawm AAI yog Thalassolituus sp. C2-1 (ntxiv NZ_VNIL01000001). Cov kab mob no tau raug cais tawm los ntawm qhov sib sib zog nqus dej hiav txwv ntawm Mariana Trench, tab sis tam sim no tsis muaj cov ntaub ntawv phenotypic txog cov kab mob no rau kev sib piv. Piv nrog ASxL5T's 2.82 Mb, kab mob genome loj dua ntawm 4.36 Mb. Qhov nruab nrab genome loj ntawm marine spirochetes yog hais txog 4.16 Mb (± 1.1; n = 92 ua tiav siv genomes tshawb xyuas los ntawm https://www.ncbi.nlm.nih.gov/assembly), yog li cov genome ntawm ASxL5T yog nyob rau hauv txoj kab nrog kev txiav txim Piv rau lwm cov tswv cuab, nws yog me me heev. Siv GToTree 1.5.54 los tsim ib qho genome-raws li kwv yees qhov siab tshaj plaws phylogenetic tsob ntoo (Daim duab 3A), siv cov kev sib txuas thiab txuas cov amino acid sequences ntawm 172 ib hom ntawv luam tshwj xeeb rau Gammaproteobacteria 11,12,13,14,15,16, 17,18 ib. Kev tsom xam pom tias nws muaj feem cuam tshuam nrog Thalassolituus, Kab Mob Kab Mob, thiab Marine Bacterium. Txawm li cas los xij, cov ntaub ntawv no qhia tau tias ASxL5T txawv ntawm nws cov txheeb ze hauv Marine Spirulina thiab nws cov ntaub ntawv genome sequence muaj.
Cov ntoo phylogenetic siv 16S rRNA gene sequence qhia txog txoj haujlwm ntawm ASxL5T, ASxO5, thiab ASxS5 hom (nrog rau lub plab) txheeb ze rau cov kab mob tsis muaj kab mob thiab cov kab mob marine hauv Marine Spirulinaceae. Genbank tus lej nkag mus ua raws li lub npe strain hauv kab lus. Siv ClustalW txhawm rau ua kom sib luag, thiab siv txoj hauv kev zoo tshaj plaws thiab Tamura-Nei tus qauv los txiav txim siab txog kev sib raug zoo ntawm phylogenetic, thiab ua 1000 coj rov ua dua tshiab hauv MEGA X program. Tus lej ntawm cov ceg qhia tau hais tias tus nqi luam tawm yog ntau dua 50%. Escherichia coli U/541T tau siv los ua ib pawg tawm.
(A) Ib tsob ntoo phylogenetic raws li cov genome, qhia txog kev sib raug zoo ntawm cov kab mob marine Spirospiraceae ASxL5T thiab nws cov txheeb ze ze, E. coli U 5/41T ua ib pawg tawm. (B) Muab piv nrog T. oleivorans MIL-1T, cov kev ua haujlwm ntawm pawg faib cov noob yog kwv yees raws li pawg orthologous (COG) pawg ntawm ASx5LT protein. Daim duab ntawm sab laug qhia txog cov noob hauv txhua pawg COG ua haujlwm hauv txhua genome. Daim duab ntawm sab xis qhia qhov feem pua ​​​​ntawm cov genomes muaj nyob rau hauv txhua pawg COG ua haujlwm. (C) Muab piv nrog T. oleiverans MIL-1T, kev tsom xam ntawm kev ua tiav KEGG (Kyoto Encyclopedia of Genes and Genomes) modular pathway of ASxL5T.
Siv cov ntaub ntawv KEGG los tshuaj xyuas cov noob caj noob ces tam sim no hauv ASxL5T genome tau nthuav tawm cov txheej txheem metabolic ntawm aerobic gamma Proteus. ASxL5T muaj tag nrho ntawm 75 genes muab rau cov kab mob lub cev muaj zog, suav nrog cov noob koom nrog chemotaxis, flagella sib dhos, thiab hom IV fimbriae system. Hauv qeb kawg, 9 ntawm 10 cov noob yog lub luag haujlwm rau kev sib tw ntawm ntau yam ntawm lwm yam kabmob. Lub genome ntawm ASxL5T muaj tag nrho tetrahydropyrimidine biosynthetic txoj hauv kev uas koom nrog kev tiv thaiv cov lus teb rau osmotic stress20, raws li xav tau rau halophiles. Lub genome tseem muaj ntau txoj hauv kev rau cov cofactors thiab cov vitamins, suav nrog riboflavin synthesis pathways. Txawm hais tias alkane 1-monooxygenase (alkB2) noob muaj nyob rau hauv ASxL5T, txoj kev siv hydrocarbon tsis tiav. Nyob rau hauv genome ib ntus ntawm ASxL5T, homologues ntawm cov noob txheeb xyuas raws li lub luag haujlwm tseem ceeb rau kev degradation ntawm hydrocarbons hauv T. oleiverans MIL-1T21, xws li TOL_2658 (alkB) thiab TOL_2772 (alcohol dehydrogenase) yog qhov tsis pom tseeb. Daim duab 3B qhia txog kev sib piv ntawm cov noob faib hauv COG qeb ntawm ASxL5T thiab txiv roj roj MIL-1T. Zuag qhia tag nrho, qhov me me ASxL5T genome muaj feem tsawg dua cov noob los ntawm txhua pawg COG piv rau cov genome loj dua. Thaum tus naj npawb ntawm cov noob hauv txhua pawg ua haujlwm tau qhia raws li feem pua ​​​​ntawm cov genome, qhov sib txawv tau sau tseg nyob rau hauv feem pua ​​​​ntawm cov noob hauv kev txhais lus, ribosomal qauv thiab biogenesis pawg, thiab cov khoom siv hluav taws xob thiab kev hloov pauv muaj nuj nqi, uas yog qhov loj dua ASxL5T. genome Qhov feem pua ​​​​yog piv nrog tib pab pawg tam sim no hauv T. oleiverans MIL-1T genome. Hauv qhov sib piv, piv nrog ASxL5T genome, T. oleivorans MIL-1T muaj feem pua ​​​​ntawm cov genes ntau dua hauv kev rov ua dua tshiab, rov ua dua thiab kho dua tshiab, thiab cov kev hloov pauv. Interestingly, qhov sib txawv loj tshaj plaws nyob rau hauv cov ntsiab lus ntawm txhua qhov kev ua haujlwm ntawm ob lub genomes yog tus naj npawb ntawm cov noob tsis paub muaj nyob hauv ASxL5T (Daim duab 3B). Ib qho kev ntsuam xyuas ntxiv ntawm KEGG modules tau ua tiav, qhov twg txhua KEGG module sawv cev rau cov txheej txheem ntawm kev ua haujlwm ntawm tus kheej rau cov lus piav qhia thiab kev txhais lus lom neeg ntawm cov ntaub ntawv genome sequence. Kev sib piv ntawm cov noob tis nyob rau hauv tag nrho KOG module txoj kev ntawm ASxL5T thiab txiv ntseej MIL-1T yog qhia nyob rau hauv daim duab 3C. Qhov kev tshuaj ntsuam no qhia tau hais tias txawm hais tias ASxL5T muaj tag nrho cov sulfur thiab nitrogen metabolic txoj kev, T. oleiverans MIL-1T tsis. Hauv qhov sib piv, T. oleiverans MIL-1T muaj tag nrho cysteine ​​​​thiab methionine metabolic txoj hauv kev, tab sis nws tsis tiav hauv ASxL5T. Yog li ntawd, ASxL5T muaj tus yam ntxwv module rau sulfate assimilation (txhais tau tias yog ib txheej ntawm cov noob uas yuav siv tau raws li phenotypic cim, xws li metabolic muaj peev xwm los yog pathogenicity; https://www.genome.jp/kegg/module.html) Hauv T oleivrans MIL-1T. Muab piv cov noob cov ntsiab lus ntawm ASxL5T nrog rau cov npe ntawm cov noob uas qhia txog kev ua neej nyob tsis muaj tseeb. Txawm hais tias waaL noob encoding lub ligase txuam nrog O antigen polysaccharide rau cov tub ntxhais yog tam sim no nyob rau hauv ASxL5T genome (tab sis nws muaj ntau yam kab mob Gram-negative), tryptophan 2,3-dioxygenase (TDO) Genes yuav suav nrog 60 amino. thaj chaw acid feem ntau pom nyob rau hauv cov kab mob predatory uas tsis muaj nyob. Tsis muaj lwm yam kab mob uas tshwm sim hauv ASxL5T genome, suav nrog cov encoding enzymes koom nrog hauv isoprenoid biosynthesis hauv txoj kev mevalonate. Nco ntsoov tias tsis muaj transcriptional regulatory gene gntR nyob rau hauv pawg predator soj ntsuam, tab sis peb gntR-zoo li noob tuaj yeem txheeb xyuas hauv ASxL5T.
Cov yam ntxwv phenotypic ntawm ASxL5T tau sau tseg hauv Table 3 thiab muab piv nrog cov yam ntxwv phenotypic ntawm lwm yam 23, 24, 25, 26, thiab 27 qhia hauv cov ntaub ntawv. Kev cais tawm ntawm T. marinus, T. olevorans, B. sanyensis, thiab Oceanobacter kriegii yog cov nquag, ntsev-tiv taus, oxidase-zoo pas nrig-zoo li lub cev, tab sis yuav luag tsis muaj lwm yam phenotypic yam ntxwv nrog ASxL5T. Qhov nruab nrab pH ntawm dej hiav txwv yog 8.1 (https://ocean.si.edu/ocean-life/invertebrates/ocean-acidification#section_77), uas yog reflected nyob rau hauv T. marinus, T. olevorans, B. sanyensis thiab O. kev krie. ASxL5T yog haum rau qhov loj pH ntau (4-9) uas tsis yog-marine hom. Phenotypic yam ntxwv ntawm Thalassolituus sp. C2-1. Tsis paub. Qhov kev loj hlob ntawm qhov kub ntawm ASxL5T feem ntau yog dav dua li ntawm marine hom (4-42 ° C), txawm hais tias qee qhov tab sis tsis yog tag nrho cov T. marinus cais tawm yog cov cua sov. Qhov tsis muaj peev xwm loj hlob ASxL5T hauv cov xov xwm broth tiv thaiv ntxiv phenotypic characterization. Siv API 20E los ntsuam xyuas cov ntaub ntawv pov tseg los ntawm BA phaj, ONPG, arginine dihydrolase, lysine decarboxylase, ornithine decarboxylase, citrate utilization, urease, tryptophan deaminase, gelatin hydrolysis Enzyme, cov txiaj ntsig tau zoo tag nrho, tab sis tsis muaj indole, H2Sacetoin. raug tsim tawm. Unfermented carbohydrates muaj xws li: qabzib, mannose, inositol, sorbitol, rhamnose, sucrose, melibiose, amygdalin thiab arabinose. Piv nrog rau cov ntaub ntawv hais txog cov kab mob uas tau tshaj tawm, cov cellular fatty acid profile ntawm ASxL5T hom yog qhia nyob rau hauv Table 4. Lub ntsiab cellular fatty acids yog C16:1ω6c thiab/los yog C16:1ω7c, C16:0 thiab C18:1ω9. Hydroxy fatty acids C12: 0 3-OH thiab C10: 0 3-OH kuj muaj. Qhov piv ntawm C16: 0 hauv ASxL5T yog siab dua li qhov qhia txog tus nqi ntawm cov genera. Hauv qhov sib piv, piv nrog T. marinus IMCC1826TT, qhov piv ntawm C18: 1ω7c thiab / lossis C18: 1ω6c hauv ASxL5T yog txo. oleivorans MIL-1T thiab O. kriegii DSM 6294T, tab sis tsis kuaj pom hauv B. sanyensis KCTC 32220T. Sib piv cov fatty acid profiles ntawm ASxL5T thiab ASxLS qhia qhov sib txawv me me ntawm tus kheej fatty acids ntawm ob hom, uas zoo ib yam nrog cov genomic DNA sib lawv liag ntawm tib hom. Tsis muaj poly-3-hydroxybutyrate (PHB) particles tau kuaj pom siv Sudan dub xeem.
Kev ua txhaum ntawm cov kab mob ASxL5T tau kawm los txiav txim siab qhov ntau ntawm cov tsiaj txhu. Cov kab mob no tuaj yeem tsim cov plaques ntawm Campylobacter hom, suav nrog: 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 and C. upsaliensis NCTC 11541T. Siv cov kab lis kev cai uas tau teev nyob rau hauv qhov kev txiav txim siab ntawm tus tswv tsev ntawm txoj hauv kev los sim ntau yam kab mob Gram-negative thiab Gram-positive. Cov txiaj ntsig tau pom tias ASxL5T tuaj yeem siv tau hauv Escherichia coli NCTC 86 thiab Citrobacter freundii NCTC 9750T. Plaques tsim ntawm Klebsiella oxytoca 11466. TEM kev cuam tshuam nrog E. coli NCTC 86 yog qhia hauv daim duab 4A-D, thiab kev cuam tshuam nrog Campylobacter jejuni PT14 thiab Campylobacter suis S12 yog qhia hauv daim duab 4E-H nruab nrab. Cov txheej txheem kev tawm tsam zoo li sib txawv ntawm cov hom tsiaj uas raug kuaj, nrog ib lossis ntau dua E. coli hlwb txuas rau txhua ASxL5T cell thiab muab tso rau tom qab ntawm lub xovtooj txuas ntxiv ua ntej adsorption. Hauv qhov sib piv, ASxL5T zoo li txuas rau Campylobacter los ntawm ib qho kev sib cuag, feem ntau yog nyob rau hauv kev sib cuag nrog lub apex ntawm cov tsiaj txhu ntawm tes thiab nyob ze ntawm lub apex ntawm Campylobacter cell (Daim duab 4H).
TEM qhia txog kev sib cuam tshuam ntawm ASx5LT thiab cov tsiaj txhu: (AD) thiab E. coli prey; (EH) thiab C. jejuni prey. (A) Ib tug raug ASx5LT cell txuas nrog ib leeg E. coli (EC) cell; (B) Ib qho filamentous ASx5LT txuas rau ib leeg EC cell; (C) Ib qho filamentous ASx5LT cell txuas nrog ntau lub EC hlwb; (D) Txuas me me ASx5LT hlwb ntawm ib leeg E. coli (EC) cell; (E) ib leeg ASx5LT cell txuas nrog Campylobacter jejuni (CJ) cell; (F) ASx5LT tawm tsam C. hyointestinalis (CH) hlwb; (G) ob leeg Ib ASx5LT ntawm tes tawm tsam CJ cell; (H) Lub zeem muag pom ntawm ASx5LT txuas taw tes, ze ntawm lub apex ntawm CJ cell (bar 0.2 μm). Qhov bar sawv cev rau 1 μm hauv (A-G).
Cov kab mob Predatory tau hloov zuj zus mus ua kom zoo dua ntawm ntau qhov chaw ntawm prey. Obviously, lawv muaj dav nyob rau hauv ntau qhov chaw sib txawv. Vim qhov nqaim ntawm cov neeg koom nrog, nws muaj peev xwm cais cov kab mob ASxL5T los ntawm slurry siv txoj kev sib cais phage. Qhov cuam tshuam genomic ntawm ASxL5T rau cov tswv cuab ntawm oceanospirillaceae tsev neeg ntawm cov kab mob marine yog qhov xav tsis thoob, txawm hais tias cov kab mob no muaj ntsev-tiv taus thiab tuaj yeem loj hlob ntawm nruab nrab uas muaj 5% ntsev. Kev ntsuas dej zoo ntawm cov slurry pom tias cov ntsiab lus sodium chloride tsawg dua 0.1%. Yog li ntawd, cov av nkos nyob deb ntawm thaj chaw marine - ob qho tib si thaj chaw thiab tshuaj lom neeg. Lub xub ntiag ntawm peb qhov cuam tshuam tab sis txawv cais los ntawm tib lub hauv paus muab pov thawj tias cov tsiaj nyeg no vam meej hauv qhov chaw tsis muaj dej hiav txwv. Tsis tas li ntawd, kev tshuaj xyuas microbiome (cov ntaub ntawv cov ntaub ntawv muaj los ntawm https://www.ebi.ac.uk/ena/browser/view/PRJEB38990) tau qhia tias tib lub 16S rRNA gene sequence yog nyob rau sab saum toj 50 feem ntau tshaj kev ua haujlwm taxa (OTU ) Hauv ob peb qhov piv txwv ntawm cov av nkos. Ob peb cov kab mob tsis zoo tau pom nyob rau hauv Genbank database, uas muaj 16S rRNA gene sequences zoo li ASxL5T kab mob. Cov kab ke no, ua ke nrog cov kab ke ntawm ASxL5T, ASxS5, thiab ASxO5, zoo li sawv cev sib txawv clades sib cais los ntawm Thalassolituus thiab Oceanobacter (Daim duab 2). Peb hom kab mob uas tsis muaj kab mob (GQ921362, GQ921357 thiab GQ921396) raug cais tawm ntawm cov dej tawg ntawm qhov tob ntawm 1.3 kilometers hauv South African kub mine hauv xyoo 2009, thiab lwm yam ob (DQ256320 thiab DQ337006) yog nyob rau hauv South Africa. hauv 2005). 16S rRNA gene sequence feem ntau ze ze rau ASxL5T yog ib feem ntawm 16S rRNA gene sequence tau los ntawm cov kab lis kev cai enrichment ntawm cov xuab zeb sediments tau los ntawm ntug hiav txwv dej ntawm sab qaum teb Fabkis xyoo 2006 (accession naj npawb AM29240828). Lwm qhov zoo sib xws ntawm 16S rRNA gene sequence los ntawm cov kab mob tsis zoo HQ183822.1 tau txais los ntawm lub tank sau leached los ntawm lub nroog pov tseg hauv Suav teb. Pom tseeb, ASxL5T cov kab mob tsis yog cov neeg sawv cev hauv cov ntaub ntawv sau se, tab sis cov kab mob no los ntawm cov kab mob tsis zoo yuav sawv cev rau cov kab mob zoo ib yam li ASxL5T, uas tau faib thoob plaws ntiaj teb, feem ntau hauv cov cheeb tsam nyuaj. Los ntawm tag nrho cov genome phylogenetic tsom xam, tus txheeb ze ze tshaj plaws rau ASxL5T yog Thalassolituus sp. C2-1, T. marinus, T. oleivorans. Thiab O. kriegii 23, 24, 25, 26, 27. Thalassolituus yog ib tug tswv cuab ntawm marine obligate hydrocarbon fragmentation kab mob (OHCB), uas yog dav nyob rau hauv marine thiab terrestrial ib puag ncig, thiab feem ntau ua tus tseem ceeb tom qab hydrocarbon pa phem tshwm sim30,31. Cov kab mob Marine tsis yog cov tswv cuab ntawm pab pawg OHCB, tab sis raug cais tawm ntawm ib puag ncig marine.
Phenotypic cov ntaub ntawv qhia tias ASxL5T yog ib hom tshiab thiab ib tug tswv cuab ntawm ib tug yav tas los unrecognized genus nyob rau hauv lub marine spirospiraceae tsev neeg. Tam sim no tsis muaj tus qauv qhia meej los faib cov kab mob tshiab uas raug cais tawm mus rau hauv cov genus tshiab. Kev sim tau ua los txiav txim siab thoob ntiaj teb genera ciam teb, piv txwv li, raws li qhov feem pua ​​​​ntawm cov genome ntawm kev saib xyuas cov protein (POCP), nws raug pom zoo tias tus nqi txiav tawm yog 50% zoo ib yam rau cov ntaub ntawv siv strain33. Lwm tus hais kom siv AAI qhov tseem ceeb, uas muaj qhov zoo dua POCP vim tias lawv tuaj yeem tau txais los ntawm cov genomes34 tsis tiav. Tus kws sau ntawv ntseeg tias yog tus nqi AAI tsawg dua 74% piv nrog cov qauv qauv ntawm hom qauv, hom kab mob yog tus sawv cev ntawm ntau hom. Tus qauv genus nyob rau hauv marine spirillaceae yog marine spirillum, thiab cov qauv hom yog O. linum ATCC 11336T. Tus nqi AAI ntawm ASxL5T thiab O. linum ATCC 11336T yog 54.34%, thiab AAI tus nqi nruab nrab ntawm ASxL5T thiab T. oleivorans MIL-1T (genus hom hom) yog 67.61%, qhia tias ASxL5T sawv cev rau cov genus tshiab txawv ntawm Thalassol. Siv 16S rRNA gene sequence raws li cov qauv kev faib tawm, qhov kev pom zoo genus delimitation ciam teb yog 94.5% 35. ASxL5T tej zaum yuav muab tso rau hauv lub genus Thalassolituus, qhia 95.03% 16S rRNA sequence tus kheej nrog T. oleivorans MIL-1T thiab 96.17%. MARINUS IMCC 1826T UAS. Txawm li cas los xij, nws tseem yuav muab tso rau hauv Bacteroides genus uas muaj 94.64% 16S rRNA gene tus kheej nrog B. sanyensis NV9, qhia tias kev siv tib lub noob xws li 16S rRNA noob tuaj yeem ua rau kev faib tawm thiab kev ua haujlwm. Lwm txoj kev qhia siv ANI thiab Genome Alignment Score (AF) los tshuaj xyuas cov pawg ntawm cov ntaub ntawv cov ntsiab lus los ntawm txhua hom thiab tsis yog hom kab mob ntawm cov genera uas twb muaj lawm. Tus sau pom zoo kom sib txuas cov genus ciam teb nrog rau qhov inflection point ntawm kwv yees genus tshwj xeeb rau taxa raug tshuaj xyuas. Txawm li cas los xij, yog tias tsis muaj cov genome ua tiav txaus los ntawm Thalassolituus cais, nws tsis tuaj yeem txiav txim siab seb ASxL5T belongs rau Thalassolituus genus los ntawm txoj kev no. Vim tias muaj tsawg tsawg ntawm cov genome ua tiav rau kev soj ntsuam, tag nrho genome phylogenetic tsob ntoo yuav tsum tau txhais nrog ceev faj. Qhov thib ob, tag nrho cov kev sib piv genome tsis tuaj yeem suav txog qhov sib txawv loj ntawm qhov loj ntawm qhov sib piv genomes. Lawv ntsuas qhov zoo sib xws ntawm kev khaws cia cov tub ntxhais kawm ib leeg ntawm cov noob caj noob ces, tab sis tsis suav nrog cov noob loj uas tsis muaj nyob hauv cov genome me me ntawm ASxL5T. Obviously, ASxL5T thiab pawg xws li Thalassolituus, Oceanobacter, thiab Bacterioplanes muaj ib tug poj koob yawm txwv, tab sis evolution tau coj ib txoj kev sib txawv, ua rau txo qis hauv genome, uas tej zaum yuav hloov mus rau kev ua neej nyob. Qhov no yog qhov sib piv rau T. oleivorans MIL-1T, uas yog 28% loj dua thiab tau hloov zuj zus nyob rau hauv ib puag ncig sib txawv los siv hydrocarbons23,30. Ib qho kev sib piv nthuav tuaj yeem ua tau nrog cov kab mob hauv nruab nrog cev thiab kev sib koom ua ke, xws li Rickettsia, Chlamydia, thiab Buchnera. Lawv genome loj yog hais txog 1 Mb. Lub peev xwm los siv tus tswv tsev cell metabolites ua rau cov noob poob, yog li Underwent tseem ceeb evolutionary genomic degradation. Kev hloov pauv hloov pauv los ntawm cov kab mob hauv hiav txwv chemical mus rau kev ua neej nyob yuav ua rau muaj kev txo qis hauv genome. COG thiab KEGG tsom xam qhia txog cov noob caj noob ces siv rau kev ua haujlwm tshwj xeeb thiab lub ntiaj teb sib txawv ntawm txoj hauv kev genomic ntawm ASxL5T thiab T. oleivorans MIL-1T, uas tsis yog vim muaj ntau ntawm cov noob caj noob ces. Qhov sib txawv ntawm G + C piv ntawm tag nrho cov genome ntawm ASxL5T yog 56.1%, thiab ntawm T. oleivorans MIL-1T yog 46.6%, uas kuj qhia tau tias nws yog cais.
Kev tshuaj xyuas ntawm cov ntsiab lus coding ntawm ASxL5T genome muab kev ua haujlwm pom zoo rau cov yam ntxwv phenotypic. Lub xub ntiag ntawm cov noob encoding hom IV fimbriae (Tfp) yog qhov txaus siab tshwj xeeb vim lawv txhawb kev txav ntawm tes, hu ua kev sib raug zoo gliding lossis convulsions, tsis muaj flagella ntawm qhov chaw. Raws li cov lus ceeb toom, Tfp muaj lwm yam dej num, suav nrog kev tua tsiaj, kab mob, tsim biofilm, ntuj DNA uptake, tsis siv neeg cell aggregation thiab kev loj hlob38. ASxL5T genome muaj 18 genes encoding diguanylate cyclase (ib qho enzyme uas ua rau kev hloov pauv ntawm 2 guanosine triphosphate rau hauv guanosine 2 phosphate thiab cyclic diGMP) thiab 6 noob encoding qhov sib thooj diguanylate cyclase phosphate diguanylate. Cov noob rau esterase (catalyzing degradation ntawm cyclic di-GMP rau guanosine monophosphate) yog qhov nthuav vim hais tias cyclic-di-GMP yog ib qho tseem ceeb thib ob tus tub txib koom nrog hauv biofilm txoj kev loj hlob thiab kev sib cais, txav mus los, kev sib txuas ntawm tes thiab virulence 39, 40 hauv cov txheej txheem. Nws yuav tsum tau muab sau tseg tias nyob rau hauv Bdellovibrio bacteriovorus, cyclic ob GMP tau pom los tswj kev hloov ntawm lub neej dawb thiab predatory txoj kev ua neej41.
Kev tshawb fawb feem ntau ntawm cov kab mob predatory tau tsom rau Bdellovibrio, Bdellovibrio-zoo li kab mob, thiab Myxococcus hom. Cov no thiab lwm cov piv txwv paub txog cov kab mob predatory tsim muaj ntau pawg neeg. Txawm hais tias muaj kev sib txawv no, ib pawg ntawm cov tsev neeg muaj protein ntau uas muaj kev cuam tshuam cov phenotypes ntawm 11 cov kab mob uas paub txog cov kab mob predatory tau txheeb xyuas 3,22. Txawm li cas los xij, tsuas yog cov noob encoding O antigen ligase (waaL) tau raug txheeb xyuas, uas tshwj xeeb tshaj yog cov kab mob Gram-negative. Daim ntawv tshuaj ntsuam no tsis muaj txiaj ntsig rau kev xaiv ASxL5T ua tus neeg tua tsiaj, tej zaum vim nws siv lub tswv yim tawm tsam tshiab. Qhov muaj cov kab mob sib txawv ntawm cov kab mob sib txawv yuav pab txhim kho kev daws teeb meem zoo dua uas suav nrog cov pov thawj ntawm kev ua haujlwm thiab ib puag ncig sib txawv ntawm cov tswv cuab. Piv txwv ntawm cov kab mob predatory tsis suav nrog hauv qhov kev tshuaj ntsuam no suav nrog cov tswv cuab ntawm Cupriavidus necator42 thiab Bradymonabacteria43, vim tias raws li cov kws tshawb fawb tshawb xyuas cov zej zog sib txawv, ntau cov tsiaj txhu raug tsim.
Qhov tseem ceeb tshaj plaws ntawm ASxL5T cov kab mob ntes tau los ntawm TEM duab yog nws cov cim thiab hloov tau yooj yim morphology, uas tuaj yeem txhawb kev cuam tshuam nrog cov kab mob prey. Hom kev sib cuam tshuam tau pom txawv ntawm lwm cov kab mob predatory thiab tsis tau pom yav dhau los lossis tshaj tawm. Lub tswv yim ASxL5T predatory lub neej voj voog yog qhia nyob rau hauv daim duab 5. Muaj ob peb piv txwv nyob rau hauv cov ntaub ntawv uas zoo sib xws apical lug raws li peb qhia ntawm no, tab sis cov piv txwv no muaj xws li Terasakiispira papahanaumokuakeensis, ib tug marine spirillum kab mob nrog tej lub sij hawm apex o 44, thiab Alphaproteobacteria, , yav tas los teej tug mus rau lub genus Oceanospirillum, nthuav tawm Lub npe hu ua "polar film" 45. Cocci cov ntaub ntawv feem ntau pom hauv cov kab lis kev cai qub, tshwj xeeb tshaj yog rau cov kab mob uas muaj cov ntaub ntawv nkhaus, xws li Vibrio, Campylobacter, thiab Helicobacter 46, 47, 48, uas tuaj yeem sawv cev rau lub xeev degraded. Kev ua haujlwm ntxiv yog xav tau los qhia meej txog lub neej voj voog ntawm ASxL5T kab mob. Txhawm rau txiav txim siab seb nws ntes thiab tua tsiaj li cas, thiab seb nws cov genome encodes biologically active compounds uas yuav siv tau rau kev kho mob los yog biotechnological hom phiaj.
Kev piav qhia ntawm Venatorbacter gen. Lub Kaum Ib Hlis Venatorbacter (Ven.a.tor, ba'c.ter, L. yog tsim los ntawm venators los ntawm L. n. venator, 'hunter' thiab Gr. n. bacter,' rod'. Venatorbacter, 'ib tug yos hav zoov qws' Cells yog aerobic, ntsev-tiv taus, nkhaus Gram stain tsis zoo, qoj qws Catalase thiab oxidase kev ua haujlwm zoo Nyob rau hauv qhov kub ntawm 4 mus rau 42 ° C Ingrown Lub pH ntau ntawm 4-9 yog txawv nyob rau hauv marine snails, feem ntau yog intolerant ntawm acidic pH Lub ntsiab fatty acids yog C16: 1ω6c thiab / los yog C16: 1ω7c , C16:0 thiab C18:1ω9; C12:0 3-OH thiab C10:0 3-OH yog pom raws li hydroxy fatty acids Lawv tsis loj hlob nyob rau hauv broth media Cov ntsiab lus ntawm DNA G + C yog 56.1 mol%. nyob hauv tsev neeg.
Kev piav qhia ntawm Venatorbacter cucullus sp. Kaum ib hlis Venatorbacter cucullus (cu'cull.us.; L. n. cucullus txhais tau hais tias fairing).
Tsis tas li ntawd, cov lus piav qhia ntawm cov genus no yog tias thaum loj hlob ntawm BA lossis BHI, cov hlwb yog 1.63 µm ntev thiab 0.37 µm dav. Cov cheeb tsam ntawm BHI agar yog me me, ncav cuag 2 hli hauv txoj kab uas hla tom qab 72 teev. Lawv yog beige, translucent, puag ncig, convex thiab ci iab. Cov tswv cuab ntawm hom no tuaj yeem siv Escherichia coli thiab Klebsiella. Campylobacter thiab ob peb lwm cov kab mob Gram-negative ua rau raug mob.
Cov kab mob ASxL5T raug cais tawm ntawm cov nqaij nyuj hauv Nottinghamshire, UK, thiab muab tso rau hauv National Type Culture Collection (UK): accession naj npawb NCTC 14397 thiab Netherlands Bacterial Culture Collection (NCCB) accession number NCCB 100775. Qhov ua tiav genome sequence ntawm AexL5T tau tso rau hauv Genbank raws li qhov sib ntxiv ntawm wb 046056.
Cov kab mob ASxL5T raug cais tawm ntawm cov nqaij nyuj siv phage cais tshuab9,49. Cov slurry yog diluted 1:9 (w/v) hauv SM tsis (50 mM Tris-HCl [pH 7.5], 0.1 M NaCl, 8 mM MgSO4.7H2O thiab 0.01% gelatin; Sigma Aldrich, Gillingham, UK), Tom qab ntawd incubate ntawm 4 ° C rau 24 teev, tig maj mam mus rau elute lub predators nyob rau hauv lub buffer. Qhov kev ncua yog centrifuged ntawm 3000g rau 3 feeb. Lub supernatant tau sau thiab centrifuged ntawm 13,000g rau zaum ob rau 5 feeb. Tom qab ntawd tus supernatant tau dhau los ntawm 0.45 µm membrane lim (Minisart; Sartorius, Gottingen, Lub teb chaws Yelemees) thiab 0.2 µm membrane lim (Minisart) kom tshem tawm cov kab mob uas tseem tshuav. ASxL5T tuaj yeem dhau cov lim dej no. Cov nyom nyom ntawm Campylobacter enterosus S12 (NCBI tus lej CP040464) los ntawm tib slurry tau npaj siv cov txheej txheem txheej txheem. Cov slurry lim tau muab faib rau ntawm txhua tus tswv cuab ntawm tes daim hlau hauv 10 µl droplets hauv triplicate thiab tso cai kom qhuav. Lub phaj yog incubated nyob rau hauv lub microaerophilic tank ntawm 37 ° C rau 48 teev nyob rau hauv microaerobic mob (5% O2, 5% H2, 10% CO2, thiab 80% N2). Cov quav hniav uas tau pom tau raug muab rho tawm mus rau hauv SM tsis thiab xa mus rau cov nyom tshiab ntawm C. hyointestinalis S12 kom nthuav tawm cov kab mob lysed ntxiv. Thaum nws tau txiav txim siab tias cov kab mob yog qhov ua rau lytic plaque thiab tsis yog phage, sim ua kom cov kab mob loj hlob ntawm nws tus kheej ntawm tus tswv tsev thiab ntxiv cov yam ntxwv rau nws. Cov kab lis kev cai aerobic tau ua ntawm 37 ° C nrog 5% v / v defibrinated nees ntshav (TCS Biosciences Lt, Buckingham, UK, ntxiv). Raws li cov lus qhia ntawm National Clinical Standards Committee, cov txheej txheem disc diffusion yog siv rau kev kuaj tshuaj tua kab mob. BHI agar yog kab lis kev cai ntawm 37 ° C siv lub disc uas muaj cov tshuaj tua kab mob hauv qab no (Oxoid) rau kab lis kev cai aerobic: amoxicillin thiab clavulanic acid 30 µg; cefotaxime 30 µg; streptomycin 10 µg; ciprofloxacin 5 µg; Ceftazidime 30 µg Nalidixic acid 30 µg; 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. Kev kam rau ntsev yog tsim los ntawm aerobic incubation ntawm BHI agar daim hlau ntawm 37 ° C. Ntxiv NaCl tau ntxiv rau BHI agar daim hlau los muab qhov sib txawv ntawm ntau txog 10% w / v. Qhov pH ntau yog txiav txim siab los ntawm kab lis kev cai aerobic ntawm BHI agar daim hlau ntawm 37 ° C, qhov pH ntau yam tau hloov kho rau ntawm 4 thiab 9 nrog sterile HCl los yog tsis muaj menyuam NaOH, thiab lub hom phiaj pH tus nqi raug txheeb xyuas ua ntej ncuav lub phaj. Rau kev txheeb xyuas cellular fatty acid, ASxL5T tau coj los ntawm BHI agar rau 3 hnub thiab aerobic ntawm 37 ° C. Raws li MIDI (Sherlock Microbial Identification System, version 6.10) tus qauv txheej txheem ntawm FERA Science Ltd, (York, UK), cell fatty acids tau muab rho tawm, npaj thiab tshuaj xyuas.
Rau TEM, ASxL5T yog kab lis kev cai aerobic los ntawm kev sib kis ntawm BA ntawm 37 ° C rau 24 teev, thiab tom qab ntawd sau rau hauv 1 ml ntawm 3% (v / v) glutaraldehyde hauv 0.1 M cacodylate tsis nyob hauv chav sov kho rau 1 teev, ces centrifuge. ntawm 10,000 g rau 3 feeb. Tom qab ntawd maj mam rov muab cov pellet nyob rau hauv 600 μl 0.1 M cacodylate tsis. Hloov cov ASxL5T ncua kev ncua mus rau Formvar/carbon zaj duab xis ntawm 200 mesh tooj liab kab sib chaws. Cov kab mob tau stained nrog 0.5% (w/v) uranyl acetate rau 1 feeb thiab tshuaj xyuas los ntawm TEM siv TEI Tecnai G2 12 Biotwin microscope. Raws li tau hais los saum toj no, muab tib cov tsiaj nyeg thiab tsiaj txhu hauv NZCYM broth (BD Difco™, Fisher Scientific UK Ltd, Loughborough) thiab incubate rau 48 teev nyob rau hauv microaerobic tej yam kev mob ntawm Campylobacter los yog Campylobacter ntawm 37 ° C, kev sib cuam tshuam ntawm predator thiab prey. kuj tau tshuaj xyuas los ntawm TEM. Aerobic tej yam kev mob rau Escherichia coli. Tshawb xyuas cov tsiaj nyeg thiab cov kab mob predatory los txiav txim seb muaj kev hloov pauv hauv cell morphology vim kev ua phem. Txoj kev Sudan dub tau siv rau kho qhov muag microscopy ntawm PHB tsub zuj zuj.
Loj hlob ASxL5T thaum hmo ntuj kab lis kev cai los ntawm smearing kev loj hlob ntawm BHI los yog BA daim hlau nrog ib tug sterile swab. Sau ASxL5T hlwb thiab ncua lawv hauv MRD (CM0733, Oxoid), thiab muab tso rau ntawm 4 ° C rau 7 hnub kom tshaib plab. NCTC siv los yog kuaj cov kab mob hauv cov kab mob hauv cov kab mob hauv BHI los yog No. 2 nutrient broth (CM007, Oxoid), incubated overnight, centrifuged ntawm 13,000g thiab resuspended hauv MRD kom txog thaum OD600 yog 0.4. Kab lis kev cai: Bacillus subtilis NCTC 3610T, Citrobacter freundii NCTC 9750T, Enterobacter aerogenes NCTC 10006T, Enterococcus faecalis NCTC 775T, Escherichia coli NCTC 86, Klebsiella NCTC146ocostca, 10817, Listeria Cov kab mob tshwj xeeb NCTC 4885, Bacillus macerans NCTC 6355T, Providencia stuartsii NCTC 10318, Pseudomonas fluorescens SMDL, Rhodococcus submarine hamburger NCTC 1621NC, Salmonella5TC7 intestinville NCTC 10861, Staphylococcus aureus NCTC 8532T, Streptococcus pneumoniae NCTC 7465T, Yersinia enterocolitica NCTC 10460. Lub Campylobacter party yog microaerobically incubated ntawm BA daim hlau ntawm 37 ° C. Cov neeg kuaj Campylobacter yog: C. coli 12667 NCTC, C. jejuni 12662, C. jejuni PT14, C. jejuni NCTC 11168T, C. helveticus NCTC 12472, C. lari NCTC 111458NC5, C. lari PT14, C... Sau cov hlwb hauv MRD, centrifuge ntawm 13,000g thiab resuspend hauv MRD kom txog thaum OD600 yog 0.4. Ntxiv ib qho aliquot ntawm 0.5 ml suspension rau 5 ml melted NZCYM sab saum toj agar (0.6% agar) thiab ncuav nws mus rau 1.2% NZCYM hauv qab phaj. Tom qab kho thiab ziab, cov serially diluted ASxL5T tau muab faib ua 20 µl droplets ntawm txhua lub rooj tsav xwm nyom hauv triplicate. Cov kab lis kev cai kub thiab huab cua nyob ntawm qhov yuav tsum tau ua ntawm cov kab mob kuaj.
Siv GenElute™ Bacterial Genomic DNA Kit (Sigma Aldridge) los npaj DNA los ntawm cov kab mob sib cais. Cov txheej txheem txheem tau siv rau PCR amplification ntawm 16S rRNA gene thiab cov khoom sib lawv liag txiav txim siv dye termination chemistry (Eurofins Value Read Service, Lub teb chaws Yelemees). Siv qhov kev pab cuam BLAST-N los sib piv cov kab ke no nrog rau lwm cov 16S rRNA gene sequences los txheeb xyuas thiab sau cov tsiaj sib ze. Cov no tau ua raws li siv ClustalW hauv MEGA X program. Cov ntoo phylogenetic tau tsim kho dua tshiab siv MEGA X siv txoj kev muaj peev xwm siab tshaj plaws raws li tus qauv Tamura-Nei, nrog 1000 daim ntawv qhia 54. Siv PureLink™ Genomic DNA Kit (Fisher Scientific, Loughborough, UK) los rho tawm DNA rau tag nrho-genome sequencing. Lub genome ib ntus ntawm ASxL5T tau txiav txim siab siv Illumina MiSeq ua ke, uas muaj 250 bp ob-kawg nyeem uas tsim los ntawm lub tsev qiv ntawv npaj siv cov khoom siv Nextera labeling thiab 2 txog 20 kb ntev nyeem los ntawm PacBio platform. Genomics DNA Sequencing Research Facility ntawm Sembia University. Lub genome tau sib sau ua ke siv CLC Genomics Workbench 12.0.3 (Qiagen, Aarhus, Denmark). ASxL5T kab lis kev cai tau muab tso rau hauv National Type Culture Collection (UK) thiab Netherlands Bacterial Culture Collection (NCCB). Cov genomes ntawm cov kab mob uas muaj feem xyuam siv rau kev sib piv yog: Thalassolituus oleivorans MIL-1T (accession naj npawb HF680312, tiav); Bacterioplanes sanyensis KCTC 32220T (accession tooj BMYY01000001, tsis tiav); Oceanobacter kriegii DSM 6294T (accession naj npawb NZ_AUGV00000000, tsis tiav); Marinamonas zej zog DSM 5604T (ntxiv ASM436330v1, tsis tiav), Oceanospirullum linum ATCC 11336T (ntxiv MTSD02000001, tsis tiav) thiab Thalassolituus sp. C2-1 (ntxiv NZ_VNIL01000001, tsis tiav). Siv JGI Genome Portal36 ntawm https://img.jgi.doe.gov//cgi-bin/mer/main.cgi?section=ANI&page= los txiav txim qhov qhab nias (AF) thiab nruab nrab nucleic acid tus kheej (ANI). Hauv khub. Txoj kev ntawm Rodriguez-R & Konstantinidis55 tau siv los txiav txim siab amino acid tus kheej (AAI). Siv GToTree 1.5.5411,12,13,14,15,16,17,18 los tsim qhov kwv yees qhov siab tshaj plaws ntawm tsob ntoo phylogenetic. Cov tswv yim genome uas sawv cev rau cov ntaub ntawv siv genome yog xaiv los ntawm cov khoom siv siv tau txheeb xyuas tias muaj feem xyuam rau ASxL5T los ntawm 16S rRNA phylogeny. Sau ntawv rau tsob ntoo siv kev sib tham sib tsob ntoo ntawm lub neej hauv online cuab yeej (https://itol.embl.de/). Cov lus piav qhia ua haujlwm thiab kev tshuaj xyuas ntawm ASxL5T genome yog ua los ntawm BlastKOALA KEGG cov cuab yeej online siv KEGG (Kyoto Encyclopedia of Genes and Genomes) module enrichment distribution. Kev faib tawm ntawm COG pawg (orthologous pawg) yog txiav txim siab siv eggNOG-mapper online cuab yeej.
Pérez, J., Moraleda-Muñoz, A., Marcos-Torres, FJ thiab Muñoz-Dorado, J. Cov kab mob tua kab mob: 75 xyoo thiab nws txuas ntxiv mus! . ib puag ncig. kab mob. 18, 766–779 (2016).
Linares-Otoya, L. thiab lwm yam. Muaj ntau haiv neeg thiab tshuaj tua kab mob muaj peev xwm ntawm cov kab mob predatory ntawm Peruvian ntug dej hiav txwv. March tshuaj. 15 Ib., 308. https://doi.org/10.3390/md15100308 (2017).
Pasternak, Z. et al. Los ntawm lawv cov noob, koj yuav nkag siab lawv: cov yam ntxwv genomic ntawm cov kab mob predatory. ISME J. 7, 756–769 (2013).
Sockett, RE Lub neej ua neej nyob ntawm tus kab mob bacteriophage Bdellovibrio. nruab. Xibhwb microbes. 63, 523–539 (2009).
Korp, J., Vela Gurovic, MS & Nett, M. Tshuaj tua kab mob los ntawm cov kab mob predatory. Beilstein J. Histochemistry 12, 594–607 (2016).
Johnke, J., Fraune, S., Bosch, TCG, Hentschel, U. & Schulenburg, H. Bdellovibrio thiab cov kab mob zoo sib xws yog cov kwv yees ntawm microbiome ntau haiv neeg hauv ntau tus tswv tsev. kab mob. Ecology. 79, 252–257 (2020).
Vila, J., Moreno-Morales, J. thiab Ballesté-Delpierre, C. Tshawb nrhiav cov xwm txheej tam sim no ntawm cov tshuaj tua kab mob tshiab. kho mob. kab mob. Kab mob. https://doi.org/10.1016/j.cmi.2019.09.015 (2019).
Hobley, L. et al. Qhov kev sib tw ob ntawm phage thiab phage tuaj yeem tshem tawm E. coli prey yam tsis muaj ib qho kev ua phem. J. Kab mob. 202, ib 00629-19. https://doi.org/10.1128/JB.00629-19 (2020).
El-Shibiny, A., Connerton, PL & Connerton, YOG Tus suav thiab ntau haiv neeg ntawm Campylobacter thiab bacteriophages cais thaum lub sij hawm pub mis ntawm pub dawb-ntau thiab organic qaib. Kev thov ib puag ncig. kab mob. 71, 1259–1266 (2005).
Wilkinson, DA thiab lwm yam. Hloov kho genomic taxonomy thiab epidemiology ntawm Campylobacter swine. kev kawm. Tus neeg sawv cev 8, 2393. https://doi.org/10.1038/s41598-018-20889-x (2018).
Lee, MD GToTree: Tus neeg siv-phooj ywg ua haujlwm rau cov kab ke genomics. Bioinformatics 35, 4162–4164 (2019).
Edgar, RC MUSCLE: Ib txoj hauv kev sib txuas ntau yam uas txo lub sijhawm thiab qhov chaw nyuaj. BMC biological ntaub ntawv. 5, 113 (2004).
Capella-Gutiérrez, S., Silla-Martínez, JM & Gabaldón, T. TrimAl: Ib qho cuab yeej rau kev sib dhos tsis siv neeg thiab txiav hauv qhov loj-scale phylogenetic tsom xam. Bioinformatics 25, 1972–1973 (2009).
Hyatt, D., LoCascio, PF, Hauser, LJ & Uberbacher, EC gene thiab metagenomic sequence translation pib qhov twv twv. Bioinformatics 28, 2223-2230 (2012).
Shen, W. & Xiong, J. TaxonKit: Hla-platform thiab npaum NCBI kev faib cov cuab yeej. Bio Rxiv. ( Nkag mus rau Lub Rau Hli 1, 2021); https://www.biorxiv.org/content/10.1101/513523v1 (2019).
Nqe, MN, Dehal, PS & Arkin, AP FastTree 2-kwv yees li qhov siab tshaj plaws tsob ntoo nrog kev sib tw loj. PLoS One 5, e9490 (2010).
Tange, O. GNU Parallel. ( Nkag mus rau Lub Rau Hli 1, 2021); https://zenodo.org/record/1146014#.YOHaiJhKiUk (2018).
Kanehisa, M. & Goto, S. KEGG: Kyoto Encyclopedia of Genes and Genomes. Kev tshawb nrhiav nucleic acid. 28, 27-30 (2000).
Czech koom pheej, L. thiab lwm yam Lub luag hauj lwm ntawm extremolytes ectoine thiab hydroxyectoine raws li kev tiv thaiv kev ntxhov siab thiab cov as-ham: noob caj noob ces, system genomics, biochemistry, thiab kev soj ntsuam cov qauv. Gene (Basel). 9., ib 177. https://doi.org/10.3390/genes9040177 (2018).
Gregson, BH, Metodieva, G., Metodiev, MV, Golyshin, PN & McKew, BA Differential protein qhia thaum lub sij hawm loj hlob ntawm obligate marine hydrocarbon-degrading bacterium Thalassolituus oleivorans MIL-1 thaum lub sij hawm kev loj hlob ntawm nruab nrab thiab ntev saw alkanes. pem hauv ntej. kab mob. 9, 3130 (2018).
Pasternak, Z., Ben Sasson, T., Cohen, Y., Segev, E., thiab Jurkevitch, E. Ib txoj kev sib piv genomics tshiab rau kev txhais cov cim qhia tshwj xeeb qhia txog kev xeeb tub hauv cov kab mob predatory. Public Science Library Ib. 10. e0142933. https://doi.org/10.1371/journal.pone.0142933 (2015).
Yakimov, MM, thiab lwm yam Thalassolituus oleivorans noob. Nov, sp. nov., ib hom kab mob marine tshiab uas tshwj xeeb hauv kev siv cov hydrocarbons. kev thoob ntiaj teb. J. System. evolution. kab mob. 54, 141–148 (2004).
Wang, Y., Yu, M., Liu, Y., Yang, X. & Zhang, XH Bacterioplanoides pacificum gen. Nov, sp. Thaum lub Kaum Ib Hlis, nws tau sib cais los ntawm cov dej hiav txwv ncig hauv South Pacific. kev thoob ntiaj teb. J. System. evolution. kab mob. 66, 5010–5015 (2016).