Kev cuam tshuam ntawm Kev Ua Siab Ntsig Ntawm Lub Plawv Ntws Ntuj Ecosystem hauv Wildling nas
Oct 30, 2023
Abstract: Mammalian holobiont harbors complex thiab interdependent mutualistic plab kab mob hauv zej zog. Kev hloov pauv hauv kev sib xyaw ntawm cov kab mob no tau paub tias yog lub hauv paus tseem ceeb hauv kev noj qab haus huv, kev tiv thaiv, thiab kab mob. Ntawm ntau lwm tus, kev noj zaub mov zoo yog cov tsav tsheb cuam tshuam rau kev cuam tshuam ntawm cov kab mob-tus tswv tsev kev sib raug zoo. Hauv cov ntsiab lus no, peb yav dhau los tau pom tias kev noj zaub mov muaj ntsev ntau (HSD) ua rau muaj qhov mob dysbiotic ntawm murine plab microbiota, uas yog qhov txo qis lossis txo qis ntawm cov kab mob uas paub zoo txog kev noj qab haus huv hauv plab. Txawm li cas los xij, vim muaj ib puag ncig tswj thiab ua kom huv si, cov pa kuaj cov nas (CLM) muaj cov plab hnyuv microbiota tsawg dua piv rau cov nas qus, ua rau cov txiaj ntsig tsis zoo rau cov kev tshawb fawb plab microbiome, vim tias qhov txo qis hauv plab microbiota ntau yam tuaj yeem ua tsis tau piav qhia txog kev sib cuam tshuam. networks ntawm microbiome. Ntawm no, peb tau soj ntsuam HSD cov nyhuv ntawm lub plab microbiota hauv CLM hauv kev sib piv rau cov nas qus, uas ua rau muaj kev cuam tshuam ntawm lub plab ecosystem ntau ze ze rau qhov xwm txheej ntawm tib neeg. Cov nas tau raug kho nrog kev tswj cov zaub mov lossis HSD thiab lub plab microbiota tau siv cov txheej txheem amplicon-based tsom rau 16S ribosomal gene. Raws li kev tshawb pom yav dhau los, peb cov txiaj ntsig tau nthuav tawm tias HSD ua rau muaj kev cuam tshuam loj ntawm kev sib txawv ntawm alpha thiab kev hloov pauv ntawm plab microbiota muaj pes tsawg leeg hauv CLM, uas yog qhov txo qis ntawm cov kab mob uas muaj txiaj ntsig zoo los ntawm Firmicutes phylum xws li genera Lactobacillus, Roseburia, Tuzzerella, Anaerovorax thiab. nce hauv Akkermansia thiab Parasutterella. Txawm li cas los xij, HSD-kho cov nas qus tsis tau qhia tib yam kev hloov pauv ntawm alpha ntau haiv neeg thiab poob ntawm cov kab mob Firmicutes li CLM, thiab feem ntau, cov tsiaj qus pom tsuas yog hloov me me hauv plab microbiota muaj pes tsawg leeg ntawm HSD. Nyob rau hauv txoj kab nrog qhov no, 16S-raws li kev soj ntsuam ua haujlwm tau pom zoo tsuas yog hloov pauv ntawm plab microbiota ecological functions hauv CLM piv rau cov nas qus thaum HSD. Peb qhov kev tshawb pom qhia tias cov microbiota nplua nuj thiab cov tsiaj qus tau los ntawm cov plab hnyuv microbiota ntau dua rau kev noj zaub mov zoo xws li HSD, piv rau plab microbiota ntawm CLM, uas yuav muaj qhov cuam tshuam tseem ceeb rau yav tom ntej kev tshawb fawb microbiome.
cistanche tubulosa- txhim kho lub cev tiv thaiv kab mob
Ntsiab lus: microbiome; noj ntsev ntau; kev tiv thaiv; hav zoov
1. Taw qhia
Lub plab ntawm cov tsiaj nyeg yog colonized los ntawm cov kab mob sib txawv thiab ntau haiv neeg, uas ua ke nrog tus tswv tsev tsim kom muaj kev sib raug zoo symbiotic [1,2]. Cov kab mob hauv zej zog no siv ntau lub luag haujlwm tseem ceeb rau tus tswv tsev, suav nrog metabolic, immunomodulating, thiab trophic functions [3-7] thiab lub plab microbiota muaj pes tsawg leeg tuaj yeem hloov pauv thaum lub neej, raws li cov kev xav tau tshwj xeeb thiab lub cev ntawm tus tswv [1,8, 9]. Ntau cov txiaj ntsig zoo ntawm cov kab mob hauv plab txhawb kev noj qab haus huv yog kho los ntawm anaerobic fermentation-derived metabolites [10-13] thiab cov kab mob dysbiotic tuaj yeem cuam tshuam rau tus tswv noj qab haus huv [2,11,14,15]. Kev txhawj xeeb loj zuj zus rau kev ua neej nyob rau kev noj qab haus huv tau ua rau muaj kev txaus siab rau kev tshawb fawb hauv plab microbiota kev koom tes thiab nws qhov cuam tshuam txog kev txhais lus [16,17]. Tseeb, lub plab microbiota yog zoo li tus extrinsic (xws li, txoj kev ua neej, kev noj haus, thiab kev kho mob) thiab intrinsic (xws li, host genetics, kev tiv thaiv kab mob thiab metabolic) yam [8,18–20]. Nws feem ntau lees paub tias cov ntsiab lus extrinsic tuaj yeem cuam tshuam qhov cuam tshuam, nrog rau kev noj zaub mov yog ib qho tseem ceeb hauv kev cuam tshuam rau lub plab microbiota muaj pes tsawg leeg thiab kev ua haujlwm [1,2,21]. Cov khoom noj khoom haus sab hnub poob, xws li kev noj ntsev ntau, paub tias ua rau mob homeostasis los ntawm kev cuam tshuam lub cev tsis muaj zog thiab hloov lub plab microbiota thiab kab mob [18,22–37]. Hauv murine gut microbiota, kev noj zaub mov muaj ntsev ntau (HSD) cuam tshuam nrog kev txo qis ntawm cov kab mob txhawb kev noj qab haus huv uas tsis muaj npe hu ua cov neeg tsim cov kab mob luv luv fatty acids (SCFA) xws li Lactobacillus spp., Bifidobacterium, Blautia thiab Faecalibaculum [28, 29,38–41], nrog rau kev nce hauv kev nplua nuj ntawm Akkermansia, lwm tus neeg tsim khoom siv SCFA uas tau pom tias cuam tshuam rau kev tiv thaiv tus tswv tsev thiab kab mob hauv cov qauv sib txawv [42,43]. Murine tsiaj qauv feem ntau siv los kawm txog kev noj zaub mov zoo li cas tuaj yeem ua rau lub plab microbiota, tiv thaiv kab mob, thiab kab mob [29,44–46]. Txawm hais tias kev siv cov pa kuaj nas (CLM) tseem yog ib qho kev xaiv siv tau rau ntau cov kev tshawb fawb, nws qee zaum ua tsis tau zoo txhais cov plab hnyuv microbiota-tsim daim ntawv thov [47–49]. Piv txwv li, kev tshawb fawb txog kev tiv thaiv kab mob thiab cov kab mob metabolomics hauv cov qauv murine ntawm cov kab mob plab hnyuv (IBD) thiab rog rog tau pom tias ua rau tsis zoo kwv yees cov txiaj ntsig ntawm kev tshawb fawb plab hnyuv microbiota [50]. Qhov no tuaj yeem yog vim muaj ntau qhov sib txawv hauv cov qauv no, xws li lub cev sib txawv ntawm lub plab, caj ces, thiab lub cev [16,50]. Txawm li cas los xij, lwm qhov teeb meem ntawm kev siv CLM rau kev kawm microbiota-immune kev sib cuam tshuam yog kev sib xyaw ntawm cov kab mob hauv plab hauv CLM, uas yog tsom rau kev txo qis ntawm qhov nyuaj thiab kev ua haujlwm ntawm CLM plab microbiota piv rau cov nas qus [51]. Qhov kev xav tau ntawm cov chaw ua kom huv thiab tswj tau ntsib qhov txo qis ntawm cov kab mob thiab cov kab mob cab, uas ntseeg tau tias yog li ua rau muaj kev tiv thaiv "kev kawm" tsawg dua hauv CLM piv rau cov nas qus [51–53]. Txhawm rau daws qhov teeb meem no, tus qauv murine wildling tau tsim los ntawm C57BL / 6 nas-derived embryo hloov mus rau cov nas qus kom tau txais cov tsiaj qus los ntawm lub plab microbiota, kom kov yeej qhov teeb meem kev txhais lus ntawm kev tshawb fawb txog kev tiv thaiv kab mob hauv plab microbiota [54]. Cov kev tshawb fawb tsis ntev los no hais txog tus qauv nas no tau pom cov txiaj ntsig zoo tshaj plaws hauv kev kwv yees tus nqi txhais lus ntawm kev sim tshuaj tiv thaiv kab mob piv rau CLM [54,55]. Ntxiv mus, wildling gut microbiota tau tiv taus ntau dua thiab muaj zog rau kev kho tshuaj tua kab mob thiab kev noj zaub mov muaj roj ntau dua piv rau CLM, piv rau cov xwm txheej nyuaj hauv tib neeg [54,55]. Txawm li cas los xij, txawm hais tias tsim los ntawm HSD ntawm lub plab microbiota, lub cev tiv thaiv kab mob, thiab ntau yam kab mob qauv hauv CLM, qhov cuam tshuam ntawm kev noj ntsev ntau rau ntawm ntuj, tsiaj qus-los ntawm plab microbiota tsis paub. Hauv txoj kev tshawb no, peb tau soj ntsuam cov txiaj ntsig ntawm HSD ntawm cov kab mob plab sib txawv ntawm cov kab mob hauv nruab nrog cev thiab kev kwv yees ua haujlwm ntawm CLM piv rau cov nas qus.
2. Cov ntaub ntawv thiab cov txheej txheem
2.1. Tsiaj thiab Noj Zaub Mov
Cov nas qus-hom C57BL/6 (7-8 lub lis piam poj niam, n=20) tau yuav los ntawm Charles River thiab nyob hauv cov chaw tsiaj ntawm University of Hasselt nyob rau hauv cov xwm txheej txheem. Cov nas qus (C57BL/6 keeb kwm caj ces, txiv neej n=12 thiab poj niam n=11) [54] tau nyob hauv thaj chaw tsiaj ntawm UHasselt nyob rau hauv cov xwm txheej txheem. Cov kev tshawb fawb tsiaj tau pom zoo los ntawm Pawg Saib Xyuas Kev Ncaj Ncees ntawm Tsiaj Txhaj Tshuaj (ECAE) ntawm Hasselt University (ID201618A4V1, ID202235). Cov nas tau nyob hauv tsev (4 nas this / tawb) hauv chav tswj qhov kub thiab txias (21–23 ◦C) nrog lub voj voog ntawm 12:12 teev tsaus ntuj. Cov zaub mov huv hauv qab no tau yuav los ntawm Ssniff (Soest, Lub Tebchaws Yelemees): 0.5% NaCl / tswj kev noj zaub mov (E15430-04), thiab 4% NaCl / HSD (E15431-34). Rau HSD, tsiaj tau pub nrog 1% NaCl hauv cov dej haus ntxiv rau E15431-34, raws li tau piav qhia hauv [28]. CLM nas tau sib npaug ntawm pawg tswj hwm (n=10) thiab HSD (n=10). Rau cov nas qus, txiv neej thiab poj niam cov tib neeg kuj tau sib npaug hauv kev tswj hwm thiab HSD pawg noj zaub mov (6 txiv neej rau kev tswj, 6 txiv neej rau HSD, 5 poj niam rau kev tswj, thiab 6 poj niam rau HSD).
cistanche cog-nce kev tiv thaiv kab mob
2.2. DNA Extraction
Microbial DNA extraction tau ua raws li tau piav qhia hauv [28], los ntawm kev siv cov txheej txheem hloov kho ntawm QIAmp Fast DNA Stool Mini Kit (Qiagen, Hilden, Lub teb chaws Yelemees). Hauv luv luv, fecal pellets tau ntxiv rau ib qho 2-mL Eppendorf uas muaj 0.5 hli iav hlaws thiab 1.5 mL ntawm lysis buffer (ASL) (Qiagen, Hilden, Lub teb chaws Yelemees). Bead-beating yog siv los ua txhua yam homogenization ntawm cov pellets. Kev rho tawm tag nrho tau ua raws li cov chaw tsim khoom raws tu qauv nrog kev hloov kho me me (ntev ntawm cov proteinase K incubation lub sij hawm mus rau 2 h ntawm 70 ◦C). DNA concentrations raug soj ntsuam siv NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, DE, USA) thiab khaws cia ntawm −20 ◦C ua ntej 16S rRNA gene amplification.
2.3. 16S rRNA Gene Amplification thiab Sequencing
16S rRNA gene sequence tau nthuav dav los ntawm kev siv primer tshwj xeeb rau thaj tsam V4 (F515/R806), raws li tau piav qhia yav dhau los [56]. Luv luv, 25 ng ntawm DNA tau siv rau ib qho tshuaj tiv thaiv PCR (30 µL) (KAPA HiFi HotStart ReadyMix, Roche, Basel, CH, USA) ntawm qhov pib denaturation rau 30 s ntawm 98 ◦C, ua raws li 25 cycles (10 s ntawm 98 ◦ C, 20 s ntawm 55 ◦C, thiab 20 s ntawm 72 ◦C). Cov tshuaj tiv thaiv tau ua nyob rau hauv triplicate, sib sau ua ke ib qho piv txwv, thiab ua kom huv los ntawm kev sib nqus hlaws-raws li kev huv huv (Agencourt AMPure XP, Beckman Coulter, Brea, CA, USA). Kev npaj tsev qiv ntawv tau ua los ntawm PCR tsis pub dhau lub voj voog kom tau txais cov tsev qiv ntawv indexed siv Nextera thev naus laus zis (Nextera XT Index Kit, Illumina, San Diego, CA, USA), ua raws li qhov thib ob AMPure XP magnetic hlaws ntxuav cov kauj ruam. Indexed cov qauv tom qab ntawd normalized rau tib qhov concentration ntawm 4nM, sib sau ua ke, thiab ua ke ntawm Illumina MiSeq platform PE300 nrog 2 × 300 bp khub-kawg raws tu qauv raws li tuam txhab raws tu qauv (Illumina, Inc., San Diego, CA, USA).
cistanche tubulosa- txhim kho lub cev tiv thaiv kab mob
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2.4. Kev Ua Haujlwm thiab Kev Tshawb Fawb Txog Kev Tshawb Fawb ntawm 16S rRNA Gene Sequencing Data
Raw sequences tau ua tiav siv QIIME 2 [57] pipeline. Tom qab qhov ntev thiab kev lim dej zoo (qhov tsis zoo li qub), cov ntawv nyeem tau lim thiab xa mus rau cov chaw ua haujlwm taxonomic (OTUs) siv DADA2 [58]. Taxonomic assignment tau ua los ntawm VSEARCH algorithm (https://github.com/torognes/vsearch; nkag rau 9 Kaum Ib Hlis 2022) thiab Silva database v128 (https://www.arb-silva.de /; nkag rau 9 Kaum Ib Hlis 2{{40}}}22). Lub rooj ASV tom qab ntawd normalized los ntawm rarefaction ntawm 6.147 qhov tob kom txhua tus qauv mus txog lub toj siab kawg ntawm qhov tsis tshua muaj qhov nkhaus. Alpha-diversity tau soj ntsuam siv ob qhov kev ntsuas sib txawv: OTUs nplua nuj (Saib), Chao1, Shannon, Simpson, Inverse Simpson (InvSimpson) ecological indexes. Rau beta-diversity, Bray-Curtis dissimilarity, Jaccard zoo sib xws, thiab Weighted and Unweighted UniFrac metrics [59] tau xam thiab npaj los ntawm Txoj Cai Tswjfwm Kev Ntsuam Xyuas (PCoA) kom pom qhov deb ntawm cov qauv. Txhawm rau normalize OTU suav cov lus, qhov tsis tshua muaj tshwm sim tau ua tiav ntawm qhov tob ntawm 6305 ntu ntawm ib qho piv txwv 100 zaug. Cov zis tau los ntawm OTU taxonomy assignment, raws li ib tug taxonomy lub rooj, yog siv los tsoo lub normalized OTU rooj rau hauv lub rooj rau taxonomy theem L2 (Phylum), L5 (Tsev Neeg), thiab L6 (Genus). Kev txheeb xyuas txheeb cais tau ua los ntawm kev siv R (https://www.R-project.org/; nkag rau 25 Kaum Ib Hlis 2022; Version 4.2.0). R pob "vegan" (Version 2.6-4) [60] tau siv los tsim cov beta-diversity metrics los sib piv qhov sib txawv ntawm pawg los ntawm PCoA lossis los ntawm kev tsom xam cov ntsiab lus tseem ceeb (PCA). Cov pob khoom thiab cov ntaub ntawv sib cais tau raug sim los ntawm kev ntsuas kev ntsuas nrog pseudo-F piv (function "Adonis" hauv "vegan"). Kev sib cais ntawm kev sib txawv ntawm beta ntawm cov pab pawg tau sim los ntawm Permutational Multivariate Analysis ntawm Variance Siv Distance Matrices (PERMANOVA, muaj nuj nqi "Adonis" hauv "vegan"), thaum qhov sib txawv ntawm cov pab pawg dispersion raug kuaj los ntawm Multivariate homogeneity ntawm pawg dispersions test (PERMDISP , ua haujlwm "betadisper" hauv "vegan"). Cov se uas tsis muaj nyob hauv tsawg kawg 4 cov qauv raug tshem tawm los ntawm kev tshuaj xyuas. Qhov sib txawv ntawm cov kev sib txawv ntawm cov kev sib txawv ntawm taxa thawj zaug tau soj ntsuam nrog qhov kev sim ua ntej Kruskal-Wallis ntawm 4 pawg thiab tom qab ntawd tau soj ntsuam ntxiv nrog Wilcoxon qhov kev sim ntawm cov kev sib piv hauv qab no: CLM Control vs. CLM HSD, wildling Control vs. wildling HSD, CLM Control vs. wildling Control, CLM HSD vs. wildling HSD. Rau kev ntsuam xyuas ntawm taxonomic sib txawv ntawm wildling thiab CLM, Linear Discriminant Analysis Effect Size (LEfSe: https://huttenhower.sph.harvard.edu/galaxy/; nkag tau rau 25 Kaum Ib Hlis 2022) tau siv los paub qhov txawv ntawm cov yam ntxwv tseem ceeb ntawm qib genus [ 61] ib. LEfSe cov txiaj ntsig tau tshwm sim raws li daim duab qhia, nrog Kev Ntsuas Kab Ncauj Tsis Txaus Siab (LDA) cov qhab nia siab dua 1.0. Thaum twg tsim nyog, p-tus nqi ntawm ntau qhov sib piv tau hloov kho los ntawm Benjamini-Hochberg txoj kev. Qhov kev tshawb pom tsis tseeb (FDR) Tsawg dua lossis sib npaug ntawm 0.05 tau suav tias yog qhov tseem ceeb ntawm cov ntaub ntawv: * p Tsawg dua lossis sib npaug rau 0.05; ** p Tsawg dua lossis sib npaug rau 0.01; *** p Tsawg dua lossis sib npaug rau 0.001. Kev ua haujlwm sib txawv ntawm microbiomes ntawm cov ntsiab lus NaCl sib txawv hauv cov khoom noj (0.5% thiab 4% NaCl cov ntsiab lus zaub mov) tau soj ntsuam los ntawm PICRUSt2, bioinformatics software pob los kwv yees metagenome cov ntsiab lus ua haujlwm los ntawm 16s rDNA gene sequencing data (https://huttenhower.sph. harvard.edu/picrust/; nkag rau 29 Kaum Ib Hlis 2022; PICRUST2 2.4.1) [62]. PICRUST2 pipeline tau siv rau cov neeg sawv cev ua ntu zus thiab lawv cov lus nplua nuj los ntawm DADA2 los ntawm kev siv cov qauv ntsuas (https://github.com/ picrust/picrust2/wiki/Full-pipeline-script; nkag mus rau 29 Kaum Ib Hlis 2022). Los ntawm tag nrho cov yeeb nkab tso zis, kev kwv yees metagenomic rau KEGG Orthology thiab MetaCyc txoj hauv kev tau tsim ua cov rooj, nrog kev kwv yees ua haujlwm raws li kab thiab cov qauv ua ke, thiab siv los sib piv cov plab hnyuv microbiota ua haujlwm hauv wildling thiab CLM raws li HSD tsoom fwv. Microbial zej zog kwv yees ua haujlwm uas ua rau muaj txiaj ntsig zoo tshaj plaws rau kev hloov pauv ntawm cov tsiaj qus thiab CLM los ntawm thawj (PC1), thib ob (PC2), thiab tus thawj tswj hwm thib peb (PC3) raug xaiv los tshuaj xyuas ntxiv raws li HSD noj hauv ob tus qauv. Lub matrix nrog rau qhov kev kwv yees muaj nuj nqi ntau yog ces normalized, hloov nyob rau hauv Centered Log Ratio (CLR) qhov tseem ceeb, thiab lub log2mean piv xam (HSD/Control) rau ob wildling thiab CLM. Thaum kawg, cov log2mean piv tau muab piv ntawm cov pab pawg los ntawm Wilcoxon-test thiab plotted raws li ib tug cuneiform zajlus. Qhov sib txawv ntawm cov pab pawg tau suav nrog hauv R software siv Wilcoxon-test thiab Kruskal-Wallis test functions thiab p tus nqi kho los ntawm Holm lossis Benjamini-Hochberg txoj kev.
3. Cov txiaj ntsig
3.1. HSD cuam tshuam rau ntau haiv neeg thiab kev sib xyaw ntawm CLM thiab Wildling Gut Microbiota
Txhawm rau tshawb xyuas qhov cuam tshuam ntawm HSD ntawm lub plab hnyuv microbial ecosystem hauv cov nas, peb tau pub HSD lossis tswj kev noj zaub mov rau cov nas qus thiab CLM. Cov nas tau khaws cia rau hauv kev noj zaub mov noj rau ob lub lis piam thiab fecal plab microbiota muaj pes tsawg leeg tau tshawb xyuas los ntawm 16S RNA gene sequencing los ntawm fecal pellets sau rau hnub 14 (Daim duab 1A). Raws li tsab ntawv tshaj tawm dhau los, tsis muaj qhov sib txawv muaj zog tau pom nyob rau hauv cov ntsiab lus ntawm lub cev hnyav ntawm kev tswj hwm thiab HSD pawg CLM thiab cov nas qus [29]. Txhawm rau ntsuas qhov sib txawv ntawm lub plab microbiota ntawm ob tus qauv CLM thiab cov nas qus ntawm qhov pib, peb kwv yees alpha ntau haiv neeg (Saib lossis nplua nuj, Chao1, Shannon, Simpson, thiab Inverse Simpson indexes), beta ntau haiv neeg (Bray-Curtis dissimilarity), thiab lub ntsiab. taxonomic txawv. Raws li cov kev tshawb fawb yav dhau los [54], wildling gut microbiota yog tus cwj pwm los ntawm kev nplua nuj microbial ntau dua (Daim duab 1B, tag nrho cov alpha diversity indexes), nrog rau qhov sib txawv thiab ntau heterogeneous microbial muaj pes tsawg leeg tshaj CLM (Daim duab 1C, PERMANOVA p =0 }.001 & PERMDISP p=0.0009, wildling vs. CLM; thiab daim duab S1). Nyob rau hauv cov nqe lus ntawm microbial kos npe, CLM thiab wildling nas gut microbiota yog tus cwj pwm los ntawm cov kab mob sib txawv (Daim duab S1). Nyob rau hauv txoj kab nrog Rosshart et al. [54], cov kab mob taxa los ntawm cov nas qus muaj rau Intestinomonas, Desulfovibrio, Tuzzerella, Oscillobacter, Orodibacter, thiab cov kab mob genus Helicobacter, uas qhia txog cov tsiaj qus uas tsis yog hauv tsev ntawm cov qauv no (Daim duab S1).
Daim duab 1. HSD cuam tshuam rau cov kab mob muaj pes tsawg leeg ntawm CLM (n=10/pab pawg) thiab cov nas qus (n=11 rau wildling Ctrl thiab n=12 rau wildling HSD). (A) Kev sim tsim. C57BL/6 CLM lossis cov nas qus tau pub rau ntawm 0.5% NaCl (tswj, Ctrl) lossis siab ntsev 4% NaCl (HSD) thiab cov kab mob hauv plab hauv zej zog lub plab uas muaj 16S rRNA gene amplicon sequencing. (B) Indexs rau alpha ntau haiv neeg ntawm fecal plab microbiota ntawm CLM thiab wildling; Los ntawm sab laug mus rau sab xis, cov kev ntsuas hauv qab no tau qhia: Pom (Tawm nplua nuj), Chao1, Shannon, Simpson, Simpson (Inverse Simpson). Qhov sib txawv ntawm cov pab pawg raug ntsuas los ntawm Wilcoxon-test. (C) Tus Thawj Saib Xyuas Kev Tswj Xyuas Kev Ua Haujlwm ntawm kev sib txawv ntawm beta los ntawm Bray-Curtis dissimilarity metric ntawm CLM vs. wildling (sab saum toj), CLM tswj vs. CLM HSD (hauv qab sab laug), thiab wildling control vs. wildling HSD (hauv qab sab xis); Kev sib cais thiab homogeneity ntawm pab pawg tau suav los ntawm PERMANOVA thiab PERMDISP cov kev xeem raws li. * p Tsawg dua lossis sib npaug rau 0.05; ** p Tsawg dua lossis sib npaug rau 0.01; **** p Tsawg dua lossis sib npaug rau 0.0001. Daim duab 1. HSD cuam tshuam rau cov kab mob muaj pes tsawg leeg ntawm CLM (n=10) / pawg) thiab cov nas qus (n=11 rau wildling Ctrl thiab n=12 rau wildling HSD). (A) Kev sim tsim. C57BL/6 CLM los yog wildling nas tau noj ntawm 0.5% NaCl (tswj, Ctrl) los yog siab ntsev 4% NaCl (HSD) thiab plab kab mob hauv zej zog lub plab uas muaj 16S rRNA gene amplicon sequencing. (B) Indexs rau alpha ntau haiv neeg ntawm fecal plab microbiota ntawm CLM thiab wildling; Los ntawm sab laug mus rau sab xis, cov kev ntsuas hauv qab no tau qhia: Pom (Tawm nplua nuj), Chao1, Shannon, Simpson, Simpson (Inverse Simpson). Qhov sib txawv ntawm cov pab pawg raug soj ntsuam los ntawm Wilcoxon test. (C) Tus Thawj Saib Xyuas Kev Tswj Xyuas Kev Ua Haujlwm ntawm kev sib txawv ntawm beta los ntawm Bray-Curtis dissimilarity metric ntawm CLM vs. wildling (sab saum toj), CLM tswj vs. CLM HSD (hauv qab sab laug), thiab wildling control vs. wildling HSD (hauv qab sab xis); Kev sib cais thiab homogeneity ntawm pab pawg tau suav los ntawm PERMANOVA thiab PERMDISP cov kev xeem raws li. * p Tsawg dua lossis sib npaug li 0.05; ** p Tsawg dua lossis sib npaug rau 0.01; **** p Tsawg dua lossis sib npaug rau 0.0001.
HSD ua rau muaj kev txo qis hauv cov kab mob sib txawv (Daim duab 1B, tag nrho cov alpha diversity indexes) nrog rau kev hloov pauv microbial tseem ceeb hauv cov muaj pes tsawg leeg ntawm CLM (Daim duab 1C, PERMANOVA p=0.001, PERMDISP p=0} .1, CLM Ctrl vs. CLM HSD). Nyob rau hauv sib piv, lub plab microbiota ntawm wildling nas yog tus cwj pwm los ntawm ntau ntau haiv neeg raws li HSD (Daim duab 1B, Saib & Chao1 indexes), divergently los ntawm CLM, thiab lawv kuj tau characterized los ntawm tsawg pronounced microbial muaj pes tsawg leeg hloov raws li HSD piv rau CLM (Daim duab 1C, PERMANOVA p=0.001, PERMDISP p=0.5, wildling Ctrl vs. wildling HSD).
3.2. Lub plab microbial muaj pes tsawg leeg ntawm Wildling nas yog tiv taus HSD dua li CLM
Bacterial compositional sib txawv ntawm wildling thiab CLM tau ntxiv taxonomically characterized. Nyob rau theem phylum, cov phyla ntau tshaj plaws ntawm cov txheeb ze abundance yog: Firmicutes (CLM: 52 ± 12%, wildling: 32 ± 34%), Bacteroidota (CLM: 24 ± 23%, wildling: 57 ± 19%), Actinobacteriota (CLM: 1{{1{16}}}} ± 7%, wildling: 0.7 ± 1.3%) thiab Verrucomicrobiota (CLM: 24 ± 23%, wildling: 0% / tsis kuaj) (Daim duab 2). Lub plab microbial profile tau pom ntau qhov sib txawv ntxiv rau tag nrho cov phyla kuaj pom hauv cov qauv fecal ntawm cov nas qus thiab CLM (Daim duab 2). Tshwj xeeb, cov tub ntxhais microbiota phyla Firmicutes, Bacteroidota, thiab Verrucomicrobiota tau sib txawv ntawm ob tus qauv (Daim duab 2). Tshwj xeeb tshaj yog, nyob rau theem tsev neeg, qhov sib txawv tau pom nyob rau hauv wildling vs. CLM plab microbiota rau feem ntau ntawm cov kab mob yav dhau los tau tshaj tawm tias HSD rhiab heev [28], suav nrog Lactobacillaceae, Clostridiaceae, Peptostreptococaceae, thiab Akkermansiaceae (Daim duab 3). Raws li qhov no, cov qauv zoo sib xws tau lees paub nyob rau theem genus ntawm wildling thiab CLM cov qauv rau cov tswv cuab tseem ceeb ntawm cov tsev neeg tau hais tseg; Ntawm cov no, cov neeg sawv cev tshaj plaws yog Lactobacillus, Roseburia, Tuzzerella, Faecalibaculum thiab Akkermansia (Figures S1 thiab 4). Txhawm rau ua kom pom qhov cuam tshuam ntxiv ntawm HSD ntawm CLM thiab wildling gut microbiota compositions, peb kuj tau txheeb xyuas qhov cuam tshuam ntawm kev noj zaub mov ntawm ntau qib sib txawv. Nyob rau theem phylum, HSD-kho CLM plab microbiota yog tus cwj pwm los ntawm kev txo qis ntawm Firmicutes thiab enrichment ntawm Verrucomicrobiota (Daim duab 2), tab sis tsis muaj ib qho loj phyla cuam tshuam los ntawm HSD hauv cov qauv wildling (Daim duab 2). Nyob rau theem tsev neeg, CLM plab microbiota yog tus cwj pwm los ntawm kev txo qis ntawm cov kab mob lactic acid uas tsim tawm xws li Lactobacillaceae, nrog rau SCFA-producers xws li Peptostreptococaceae thiab Clostridiaceae (Daim duab 3). Tsis tas li ntawd, hauv HSD-fed CLM, peb pom qhov nce hauv Akkermansiaceae, Sutterellaceae, Defluvitaleaceae, thiab Eggerthellaceae (Daim duab 3). Hauv qhov sib piv, HSD cuam tshuam cov kab mob sib txawv hauv cov kab mob hauv lub plab microbiota, ntawm lawv ob lub siab-ntaus Muribaculaceae thiab Prevotellaceae, ob qho tib si tau nce raws HSD (Daim duab 3). Cov kab mob hloov pauv uas feem ntau ua rau HSD-tshuaj cuam tshuam hauv CLM suav nrog kev nce ntawm genera Akkermansia, Parasutterella, thiab Enterorhabdus, nrog rau kev txo qis ntawm Lactobacillus, Roseburia, Tuzzerella, (Eubacterium) oxidoreducens pawg, Muribaculum thiab Anaerovorax (Daim duab 4). Tsuas yog Roseburia, tsis muaj ib qho ntawm cov kab lus hais saum toj no tau cuam tshuam los ntawm HSD hauv wildling gut microbiota, thaum lub genus Anaerovorax tau pom qhov sib txawv ntawm CLM (Daim duab 4).
Daim duab 2. HSD-kev cuam tshuam rau cov kab mob phyla los ntawm plab microbiota ntawm CLM (n=10/pab pawg) thiab cov nas qus (n=11 rau wildling Ctrl thiab n=12 rau wildling HSD). Tag nrho cov muaj pes tsawg leeg nyob rau hauv cov nqe lus ntawm phyla txheeb ze abundance yog qhia los ntawm bar zaj duab xis rau txhua tus neeg (sab saum toj) thiab boxplot rau tshwj xeeb phyla (hauv qab); Kev sib piv cov lej tau ua los ntawm cov pab pawg los ntawm Wilcoxon-test. * p Tsawg dua lossis sib npaug rau 0.05; ** p Tsawg dua lossis sib npaug rau {{10}}.01; *** p Tsawg dua lossis sib npaug rau 0.001; **** p Tsawg dua lossis sib npaug rau 0.0001.
Daim duab 3. Kev cuam tshuam ntawm kev noj zaub mov muaj ntsev ntau rau cov tsev neeg kab mob ntawm CLM (n=10/pab pawg) thiab nas qus (n=11 rau wildling Ctrl thiab n=12 rau wildling HSD). Tag nrho cov muaj pes tsawg leeg ntawm tsev neeg yog sawv cev los ntawm bar zaj duab xis rau txhua tus neeg (saum toj) thiab boxplot rau cov tsev neeg tshwj xeeb (hauv qab); Kev sib piv cov lej tau ua los ntawm cov pab pawg los ntawm Wilcoxon-test. * p Tsawg dua lossis sib npaug rau 0.05; ** p Tsawg dua lossis sib npaug rau {{10}}.01; *** p Tsawg dua lossis sib npaug rau 0.001; **** p Tsawg dua lossis sib npaug rau 0.0001.
Daim duab 4. Kev hloov ntawm cov kab mob hauv CLM (n=10}/pab pawg) thiab cov nas qus (n=11 rau wildling Ctrl thiab n=12 rau wildling HSD). Zuag qhia tag nrho cov txheeb ze abundance pab nyob rau hauv lub genus theem yog plotted raws li ib ncig bar zaj duab xis rau txhua tus neeg (sab saum toj) thiab ib lub boxplot rau tej genera (hauv qab); Kev sib piv cov lej tau ua los ntawm cov pab pawg los ntawm Wilcoxon-test. * p Tsawg dua lossis sib npaug rau 0.05; ** p Tsawg dua lossis sib npaug rau 0.01; *** p Tsawg dua lossis sib npaug rau 0.001; **** p Tsawg dua lossis sib npaug rau 0.0001.
3.3. HSD cuam tshuam rau Predictive Microbial Functions hauv CLM tab sis tsis nyob hauv Wildling nas
PICRUST 2 cov zis tsis pom qhov sib txawv tseem ceeb ntawm microbial zej zog kev ua haujlwm ntawm wildling HSD vs. cov nas qus tsis tau kho rau ob qho tib si KEGG Orthology thiab MetaCyc txoj kev annotations, nrog rau tsuas yog kev zam ntawm HSD-induced muaj nuj nqi ntawm recG noob rau ATP-nyob ntawm helicase los ntawm the KEGG Orthology (Figure 5A). Kev cuam tshuam HSD ntawm CLM yog tus cwj pwm los ntawm qhov txo qis ntawm kev kwv yees ua haujlwm rau KEGG Orthology, ntawm lawv cov noob spp (sucrose-6-} phosphatase) thiab pfkA (phosphofructokinase 1), ob qho tib si koom nrog cov hmoov txhuv nplej siab thiab sucrose metabolism, uas yog nyob rau hauv kab. nrog cov kev tshawb pom yav dhau los [28] (Daim duab 5A). Tsis tas li ntawd, lub plab microbiota ntawm HSD-fed CLM yog tus cwj pwm los ntawm kev txo qis kev ua haujlwm ntawm cov noob koom nrog hauv kev thauj mus los (feoB rau hlau thauj, AB 2P AB 2 permease protein, AB 2A AB 2 ATP binding protein), glutamine biosynthesis (glnA) , LacI tsev neeg transcriptional regulator (lacI, galR) thiab transketolase (tktA, tktB) (Daim duab 5A). Rau MetaCyc txoj hauv kev, HSD tau txhawb nqa CLM lub plab microbiota ntawm kev kwv yees ua haujlwm cuam tshuam nrog kev txo nitrate (txoj kev denitrification), galactose degradation (D-galactarate degradation, super txoj kev ntawm D-glucarate thiab D-galactarate degradation), phenyl-propanoate degradation, fatty acid salvage, succinate degradation rau butanoic acid thiab amino acid degradation (aromatic amine degradation, L-leucine degradation) (Daim duab 5B). Tsis tas li ntawd, nyob rau hauv txoj kab nrog kev tshawb pom yav dhau los [28], HSD plab microbiota hauv CLM poob kev ua haujlwm rau ammino acid biosynthesis (super pathway ntawm L-alanine biosynthesis, L-lysine biosynthesis), tov kua qaub fermentation, nrog ntxiv tshiab kos npe poob zoo li N- acetylglucosamine/N-acetyl-mannosamine/N-acetylneuraminate degradation thiab deoxyribonucleosides degradation (pyrimidine thiab purine degradation, inosine5phosphate biosynthesis III) (Daim duab 5B).
Daim duab 5. Cont.
Daim duab 5. Qhov cuam tshuam ntawm HSD ntawm lub plab kwv yees metagenomic ua haujlwm hauv CLM (n=10} / pawg) thiab wildling (n=11 rau wildling Ctrl thiab n=12 rau wildling HSD) plab microbiota. PICRUST2 cov ntaub ntawv tso tawm tau npaj raws li cov phiaj xwm cuneiform rau KEGG Orthology annotation (A) thiab MetaCyc txoj hauv kev (B) qhia raws li log2 txhais tau tias piv ntawm kev kwv yees ua haujlwm suav nrog HSD vs. Ctrl cov qauv. Txhua qhov kev sib piv cov lej tau ua ntawm Ctrl vs. HSD pawg los ntawm Wilcoxon-test.
4. Kev sib tham
Complex thiab ntau haiv neeg wildling gut microbiota paub tias yuav muaj zog dua rau qee cov qauv kab mob [51] thiab kev noj zaub mov zoo, xws li kev noj cov rog ntau [54,55]. Txawm li cas los xij, tsis muaj kev tshawb fawb yav dhau los tau ntsuas qhov cuam tshuam ntawm kev noj cov sodium siab ntawm murine wild-derived plab microbiota. Ntawm no, peb tau tshawb xyuas thawj zaug yuav ua li cas HSD cuam tshuam rau wildling plab microbiota piv rau CLM. Interestingly, peb cov txiaj ntsig tau pom tias piv rau CLM, cov tsiaj qus microbiome tiv taus ntau dua rau HSD kev cuam tshuam ntawm ob qho tib si muaj pes tsawg leeg thiab kev kwv yees ua haujlwm. Nws yog qhov tsim nyog tias kev noj ntsev ntau tuaj yeem ua rau muaj kev pheej hmoo ntawm ntau yam kab mob, xws li kab mob plawv lossis kab mob autoimmune, los ntawm kev hloov cov plab hnyuv microbiome muaj pes tsawg leeg thiab kev tiv thaiv kab mob homeostasis [25,29,31,34,63–65]. Raws li cov ntaub ntawv dhau los, HSD-vim hloov pauv hauv plab microbiota hauv CLM tau pom los ntawm kev hloov pauv loj hauv microbial ntau haiv neeg, muaj pes tsawg leeg, thiab kev kwv yees ua haujlwm [28]. Cov kab mob txhawb kev noj qab haus huv xws li tsev neeg Peptostreptococcaceae thiab genera Lactobacillus, Roseburia, thiab Tuzzerella txo qis ntawm cov txheeb ze ntau hauv CLM, thaum Akkermansia nce ntau hauv HSDfed pawg. Peb kuj tau kuaj pom cov txheeb ze ntau dua ntawm HSD hauv Defluvitaleaceae, Enterorhabdus, thiab Parasutterella. Interestingly, genus Parasutterella yog ib qho tseem ceeb ntawm lub plab microbiota ntawm ob qho tib si CLM thiab tib neeg, qhov twg nws coj li asaccharolytic thiab tsim tawm ntawm succinate [66]. Ob leeg Enterorhabdus los ntawm tsev neeg Eggerthellaceae thiab Parasutterella los ntawm tsev neeg Sutterellaceae paub tias muaj txiaj ntsig zoo rau cov neeg mob IBD [67,68], qhia ntxiv tias HSD yuav cuam tshuam li cas rau kev txhim kho kab mob. Txawm li cas los xij, qhov zoo siab, cov nas qus tsis tau pom qhov chaw zoo sib xws ntawm HSD-vim cov kab mob microbial hloov, xws li CLM. Txawm li cas los xij, kev sib txawv ntawm cov tsiaj qus tau nce ntxiv ntawm HSD rau kev soj ntsuam OTUs thiab Chao1 cov kev ntsuas, thiab tsuas yog ob peb cov se tau koom nrog HSD kev cuam tshuam ntawm cov plab hnyuv microbiota, ntawm lawv qhov nce ntawm Anaerovorax, ua ke nrog kev txo qis ntawm Erysipelatoclostridium, Roseburia, thiab Lachnospiraceae. UCG-004 genus. Roseburia tsuas yog cov kab mob kos npe uas feem ntau sib koom ntawm HSD pawg piv nrog cov kev tswj hwm, txawm hais tias HSD-piv CLM tseem yog tus cwj pwm los ntawm ntau dua ntawm cov kab mob no piv rau HSD-fed wildling nas. Nco ntsoov, butyrate-ua cov kab mob xws li Roseburia tau pom tias muaj cov txheeb ze qis dua hauv cov neeg mob ulcerative colitis [69] thiab qhov kev txo qis no kuj tau pom tias muaj feem cuam tshuam nrog IBD caj ces kev pheej hmoo ntawm tib neeg kev kawm [70]. Qhov no yog nyob rau hauv txoj kab nrog kev tshawb pom yav dhau los, qhov hloov pauv hauv cov kab mob xws li Roseburia lossis Lactobacillus tau pom tias muaj feem cuam tshuam nrog kev pheej hmoo ntawm kev kub siab, tej zaum yuav txhawb nqa los ntawm Western noj mov [71]. Cov kab mob hauv plab kuj tseem cuam tshuam nrog lub plab motility thiab physiology [72].
cistanche cov txiaj ntsig rau txiv neej-ua kom muaj zog tiv thaiv kab mob
Lub genus Anaerovorax yav dhau los tau pom nyob rau hauv cov nas nrog lub plab physiology txawv txav thiab txo motility [73]; Txawm li cas los xij, kev ua kom muaj txiaj ntsig ntawm Anaerovorax hauv HSD rau cov nas qus tuaj yeem ua rau muaj lub luag haujlwm sib txawv ntawm cov taxa no hauv cov ntsiab lus ntawm lub plab homeostasis thiab kev ua haujlwm zoo. Raws li kev tshawb pom yav dhau los, peb tau pom qhov nce hauv cov genus Akkermansia hauv HSD pawg CLM [28], thaum lub plab microbiota ntawm cov nas qus tau depleted ntawm cov genus no, uas kuj zoo ib yam nrog cov kev tshawb fawb ua ntej ntawm cov qauv no [51, 53–55] ib. Txawm hais tias lub genus Akkermansia yog qhov muaj peev xwm probiotic vim nws cov txiaj ntsig zoo ntawm kev txhim kho cov tswv tsev tiv thaiv kab mob thiab cov kab mob metabolic (xws li, rog thiab ntshav qab zib hom 2) [42,74–77], lub luag haujlwm ntawm cov genus no tseem tsis meej vim nws qhov tsis zoo. Kev sib raug zoo nrog cov txiaj ntsig kev kho mob hauv kab mob qog noj ntshav [78], Parkinson's disease [79,80] thiab ntau tus neeg mob sclerosis [81]. Raws li peb cov txiaj ntsig yav dhau los tau nrog MetaCyc txoj hauv kev [28], CLM raws li HSD tau pom tias txo qis kev ua haujlwm ntawm microbial cuam tshuam nrog cov hmoov txhuv nplej siab thiab sucrose metabolism rau KEGG orthology. Txawm li cas los xij, kev hloov pauv me me hauv plab kab mob muaj pes tsawg leeg ntawm HSD-fed wildling nas tsis tuaj yeem ua rau muaj kev hloov pauv tseem ceeb hauv kev kwv yees cov kab mob ua haujlwm, qhia tias cov tsiaj qus-derived plab microbiota thiab metabolic / ecological networks ruaj khov dua thiab yuav hloov tau yooj yim dua. HSD-induced dietary variations piv rau CLM plab ecosystems, uas lav kev tshawb nrhiav ntxiv. Tsim nyog hais kuj yog qhov ua tau ntawm lub plab fungal zej zog ntawm lub plab kab mob network raws li kev noj zaub mov txawv. Cov kev tshawb fawb yav dhau los tau qhia tias yuav ua li cas muaj peev xwm cuam tshuam ntawm cov kab mob thiab cov fungi cuam tshuam rau hauv lub cev tiv thaiv kab mob homeostasis thiab kab mob kev loj hlob [82-85]. Nyob rau hauv cov ntsiab lus no, CLM raug txwv ntxiv los ntawm lawv cov kab mob qis dua piv rau cov nas qus, uas tuaj yeem cuam tshuam rau kev tsim muaj ntau lub plab mycobiota [54]. Cov kev tshawb fawb yav tom ntej yuav tuaj yeem txiav txim siab qhov kev koom tes ntawm cov kab mob hauv plab hauv cov chaw ntawm plab microbiota thiab kev tiv thaiv tus tswv tsev los ntawm kev siv cov qauv wildling. Hauv cov ntsiab lus, peb txoj kev tshawb fawb muab cov ntaub ntawv hais txog kev noj cov tshuaj sodium ntau npaum li cas cuam tshuam rau lub cev, cov tsiaj qus tau los ntawm plab hnyuv microbial ecosystem hauv kev sib piv rau cov kab mob hauv tsev neeg ntawm CLM. Peb txoj kev tshawb fawb pom tau tias HSD tsis cuam tshuam rau cov kab mob taxa thiab plab microbiota nyob rau hauv cov nas qus nyob rau hauv tib txoj kev raws li nws ua rau ib tug domesticated plab microbiota los ntawm CLM. Qhov kev sib txawv no, raws li tau hais yav dhau los rau lwm cov kev noj haus lossis cov xwm txheej xws li kev noj zaub mov muaj rog [54,55], qhia tias kev tshawb fawb yav tom ntej yuav tsum muaj nyob rau hauv cov qauv murine los rov ua dua thiab kwv yees qhov cuam tshuam ntawm kev noj zaub mov rau ntau lub plab hnyuv ecosystems, zoo li hauv tib neeg.
cistanche tubulosa- txhim kho lub cev tiv thaiv kab mob
Cov ntaub ntawv
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