soinsi2021 Gut microbiota of adults wih diferent metabolic phenotypes | Elsevier Enhanced Reader Nution 90202) 111293 Contents lists available at ScienceDirect Nutrition journal homepage: www.nutritionjral.com LSEVIER Applied nutritional investigation Gut microbiota of adults with different metabolic phenotypes iy Priscila dos Santos Gomes Olivares MSc.°, Ana Beatriz Furlanetto Pacheco Ph.D.‘ Luciana Nicolau Aranha Ph.D.®, Bianca da Silva Oliveira M.Sc.*, Allan Amorim Santos Ph.D.°, Paula Cristina Moreira dos Santos ', José Firmino Nogueira Neto Ph.D. °, Glorimar Rosa Ph.D.*”, Glaucia Maria Moraes Oliveira Ph.D.” Pesaro rogram in eine Carly) Federal Unley of fo dejan, dean. 8 "Depron nd ne js dC Nuriton Feder! nies of Ro deni, eo, “ares Cages i stare py (KC). Federal Unies ie data ode Ba "abort f rpg and Tosca arose Crt Ch Fo ops bu, Fl Uniersiy fio Jani doc La * ins teeta. Rc Medel ees Ro de ae Se nest. dela, re "ese dase Niamey af Ro doe. de ae. a ARTICLE INFO ABSTRACT Arie sory Received 102020 Received in eve oo 15 Apel 2021 eco 18 Ape 2021, ‘objective The aim of this study was o investigate the gut microbiota of individuals with diferent metabolic ‘phenotypes and to compare their characteristics. “Methods: This was a ross-sectional study with 109 adults, classified according to metabolic status, by body mass index, and homeostasis model assessment. Anthropometric and biochemical characteristics and blood pressure were evaluated. Level of physical activity was evaluated by means of a questionnaire, and dietary = Consumption was evalated using a3 fod recor. Fees sample were collect rom each partipant, Cat micrebits and gut microbiota profile was analyzed by sequencing ofthe 165 rRNA gene The Kruskal-Wallis and \?tests obey ‘wer sed acending to the seaeoFmeasirerent othe variables, on the tats program SPSS version 25, Overweight ‘ith significance level of 5% Metaboli healt phenotype hoe Results: Significantly lower values of saturated fa intake and fasting glycemia were observed in metabolically healthy individuals who were overweight and obese compared with their unhealthy counterparts Diversity nd richness of terial communities were lower in individual who were obese and inthe unhealthy phe- notype. The genus Eubacerium rectale and genera ofthe family Pevotéllacee were more abundant in the metabolically healthy obese and metabolically healthy overweight groups than inthe metabolically healthy rnoemal weight one, whereas Bifidobacterium was more abundant in the metabolically healthy normal weight rou. The genera Coprococcs and Ruminococcus were more abundant inthe metabalially healthy over- ‘weight group than inthe metabolically unhealthy overweight one. Conclusions: We observed distinct characteristics in the gut microbiota of diferent metabolic phenotypes. ‘The intestines of individuals with unhealthy phenotypes hosted inflammation-associated microbiota with lower butyrate production potential and reduced bacterial diversity ©2021 Elsevier Inc.All rghts reserved. Introduction Overweight and obesity have increased in all countries, epre- ‘Guo and GX were espns for designing the sty a wring the an Scrip PISGO was responsible for ecru ad asst pallens, borate) ata Yiis DNA extraction, analysis and interpretation of eeu and wing ofthe ‘manuscript. ABEP was esponse for inerpreting he result of he gut meebiota and wing the manescrpt.AAS was responsible for interpreting the resus of the fut micobira UNAasited inthe wring of the manuscript BdSO and MAS Sse in patent cae, abrator anajss and ONA eration, jFMN Was respons. Dietor the laboratory nays A autor ead ae spproved the final mane. ‘heauthorshave no cof of iterest fo deca, "omespnding author: Tel:~35 2139386225, Feat ees: lacamoracsolveraepmai.com (GMM. Ofer) xp dotorg)ton046i.u-2021111293 (29 90070 201 Elsevier nc Alig served, senting a worldwide public health problem [1] In Brazil, accord ing to data from the Surveillance of Risk and Protective Factors, for Chronic Diseases by Telephone Survey, carried out by the Min- istry of Health in 2018 to monitor the frequeney an distribution Of risk and protective factors for non-transmissible chronic dis- eases in the capital cities of Brazilian states and the Federal Dis- trict, 55.7% ofthe adult population was overweight and 19.8% was obese [2 htips:ireaderelsevier.comieadertspiv'S0899900721001558 7token=63034DC8F477B57EBCBFFSEAIE 1A38B120377S7DAASFEAIATECFBBF. 18soinsi2021 Gut microbiota of adults wih diferent metabolic phenotypes | Elsevier Enhanced Reader 2 P46 Otvere triton 802021) 11293 Obesity is considered an important isk factor for the develop- ‘ment of diverse metabolic complications [3|. It has, nonetheless, ‘been reported in the literature that metabolic changes related 10 ‘obesity vary among individuals (4), Accordingly. the term metaboli- «ally healthy has been proposed to characterize a phenotype that is ‘not accompanied by metabolic dysfunction, Attempts to categorize individuals according to metabolic health and body mass index (BM) have given rise tothe following diferent phenotypes: meta- botically healthy normal weight (MFINW), metabolically unhealthy normal weight, metabolically healthy overweight (MHOW), meta- Dolically unhealthy overweight (MUHOW), metabolically healthy ‘bese (MEO), and metabolically unhealthy obese (MUHO) [4 Lite is understood about the mechanisms that might explain the {avorable metabolic profile of some individuals who are overweight. is known that genetic and lifestyle factors contribute to the develop- ‘ment of obesity; the combination of these factors, however, is not ‘capable of fully explaining the pathogenesis of obesity, and studies from recent years have suggested that gut microbiota is an important factor that may influence the host phenotype, due tits rol in regulat- ing inflammatory responses, energy homeostasis, and metabolism of slucose and lipid [57] contributing tothe development of non-aico- hii fatty ivr diseases, in a high-fat det model |S). ‘We have hypothesized that metabolically healthy and ‘unhealthy individuals may be distinguished based on characteris- ties of gut microbiota; tis differential distribution between meta- bolic phenotypes has been litte studied, however. Knowledge about the composition of gut microbiota of individuals with dtfer- ent metabolic phenotypes may provide a physiologic explanation as to why some individuals are more susceptible to weight gain land metabolic disorders than others. From this perspective, the ‘objective ofthis study was to investigate whether there are differ- fences in gut microbiota of adults with diferent metabolic pheno- types and to compare their characteristics. Paricipansand methods study poplson The study sample comprised 10 volumes sled frm the Cn Nut ‘on esearch and Extension Cente ofthe Josue de Casto Nuitin Intute and ‘he Clementino Fraga Univesty Hostal (HUCF, acronym in Portugese) ot {he Federal Unversity of ie de Janeiro (UFR 2ronym in Portuguese) This stay ‘exe appre rom the Resear Eis Comite ofthe HUTT UFR Al a ‘Spams signed fe and normed consent forms. "ne stay iced adits eeweeh 20 and 59 yo a ofboth sexs, with BA 185 ki The owing vere enced: ind wing dear sippements pre- tii anar pata rng the st 2 to: women wo were rena or re feeding: menopausal Wome nds agnosed wth dabetes melts oF ing typelyennc mediate nd wt pat ie, anny ite tial ess or eal icy, nda sg antic ig the ps2 nates, pid omerng drugs o cried susan: india with recat ep ‘odes ies ding the ps2 rand estan Individuals were dvde no fe groups, cording to BN] and Hameo sis model assessment (HOMA) nde: vidas with HOMA <275 were conse fred metabolically healthy, and these with HOMA =278 were considered ‘mecatolcally unalhy (10), Coups were ienied as MEN, MHOW, MEO, ‘MUHOW. a MUO. Anctrpomeric evaluation. oly composi ant od pressure measurement “Anthropometric ealation was cari ot sing the flowing parameters toy weit (a). eh (tm. waist ccunference (mn) [1] eck ceumerenee (on) [21 BM (kx and watt hgh ratio, BMI as Cleated allowing ‘he Weld Health Organization guieins [9] ody compesiten was evahtd by teapelar Moumpedsnce (dynamics 450, todmames corporation, Washing toa) and bled pesure was measured sing the auscultatary method [13 ood samples were olcted afer 312s ast, and serum was obtained by comifgaton (15 mi, 400) Serum concentrations of lus, waclalycrls (765, high-density ipoprtcn (HDL an toa chokesterol were determine by fhe enaynaie method wan automated iohenncal analy (best Dupe tea SA Vista Alege, Lagoa Sata = Braz), Concentations af lw dns pe proteln were caleulted using the riedevald frm 14. Serum incl was ‘htained by chemiluminescence, and insulin resistance was estimated using the OMA nde 15) Anadis of etry cansumpton and evaluation of pyc acy vel. Dietary intake wasanalyed ung 13-4 food record (2 typical days and 1 ay ica, and the data were entered into the program Foo! Processor. version 72 (Caaesere, Sle, MA USA). Level f psa act was evalted Bf the Short versio he neato Physical Act Question [16 ‘Sumple reparation, DKA sequencing nd dat analysts Fecal samples were cole by participants at home ozen in sealed plastic ‘gs, transported fo the boat, and inwedately sted at 20 Baer DDNA was extracted fom 200mg of fecal sample using BNA stool Ks (Norge, ‘Ona, canada) flowing manufacturer's instructions, quantified by Nanadrop (Thermo Fisher Sci Witham, MA, USA) an stored at 80"C. Equal masses ff DNA from tre individuals in the same group were poo. Five pools were ince in each experimental proup. The VI" variable ren ofthe 165 7DNA ene wa amplified ing specie primers. The DNA amplification, seeing Sin data ana proces were ced ot ing Gene Bote (0 4e Jani. Bran) a deserted i the supplementary mates The sequences ‘were deposited inthe National Center for thotechnology Information database ter ctess number PRINAGS 293 Scenes th = $7 sary were atte! to the se operational o- same ut (OTU To compare communes the abun of TU was somal ting seence rate cvrespod tthe sunple wh the ewe Soqunces We ‘asiered ely OTUs thar conte = Wt the tal sequences a epee este Sunes inthe microbial communities vesity was estimated by celine “versity indies Gos overage Shanon a phgenede dest) and ness {ebsered specs a chs) and vey wasted and represented yma ‘neh mules case) denn Sasa ana Results were express median and inceraarile interval or percentage Te Shapiro” Wik normaly test was apple. For comparison betnten group, "he rua Walls est with post hac Soneron test andthe 2 test were st Sccording to the sae of measurement ofthe arable Analyses were conducted ting the statistical package SPS version 25 (IBM, Armonk, NY. USA). Weaxon tests wee applied to determine siniieantdifrenes 1 vals of ees Indices betwen groups Anais of molecule vance (AMOVA) was perfoaned to determine wheter the difeence nthe truce ofthe mobil community between groups was sgn. The ¢ test was used o determine slgnicant di ferences nthe relative abundance of specific taxonomic groups Between NHOW Sind MUMOW apd benscen MHD snd MUO, Rests with P O15 were cons ‘red satisically signa Results Data from 109 individuals were evaluated; the majority were ‘women (858), and the median age was 39 y. Sociodemographic characteristics and dietary intake ofthe study groups are shown in Table 1, and their anthropometric, clinical, and biochemical data are shown in Table 2. Significant differences were observed in Socioeconomic status; alcohol and saturated fat intake; and TG, ‘glucose, and insulin levels, For analysis of the composition of gut microbiota by metage- nomics, each group was represented by five samples (except MHNW, with four samples) and each sample comprised a pool of thtee individuals For comparison, the samples were normalized to aan equal number of sequences (30. 079 readings). The resulting number of sequences was sufficient to characterize the bacterial ‘communities, as illustrated by the rarefaction curves that approached saturation and by Good's coverage values >99% (Sup- plementary Fig. 1 and Supplementary Table 1) Each sample con- tained, on average, 608 OTUs. When comparing a-diversty indices according to all parame- ters evaluated, the MUHO group showed less diversityichness hitips:ireaderelsevier-comiteadertspiv'S0899900721001558 7token=63034DC8F477857EBCBFFSEAIE 1A38B12037737DAASFEAIATECFBBF. 218soinsi2021 Gut microbiota of adults wih diferent metabolic phenotypes | Elsevier Enhanced Reader PAS Olivares eta Nurion 902021) 111293, 3 able Siodemograpic characteristics and dietary ike variates Toal(N= 109) MNW{n=18) __MHOW(a=17)__MHO(n=29) __MUNOW(n=15) __MUNO(a=30)__Falue nee) 396250) «SATIS 385) GSS) AGRA) ——«SAT-ATS)—«ATGGLIS 5125) “NS Sox 30) Men 15010) 110) 1139) 1030) 220) 230) Ns Women 56) 349(16) 332(15) 387005) 387) 787123) Noite race (0) 6407) 65(12) 570) 585(17) ae) 70(21) xs Sine. 50) 54038) 585(10) our) aia) 45110) 533(10) is Levelofscooting=12y.3(n) $3(00) sua) 10017) 335025) 780) 30123) ks Soxilaleool use (0) 41085) 4408) 420) se(i8) 3330) 230) ‘03s, Tebaco se 2(6) 30) 36(1) na) 1033) 83) 330) xs Sedetaceon (0) «2(on) 7a (7) 2118) ai) 667 (10) 787 (23) Xs Hypertension (a) i820) 5601) 136) 210) 13330) 30) ns Dyson, (9) 1) a ° ° 0 672) xs Perea incre betwee ‘ow Dapd2imes MW. 2(H)——_54(70) a5(12) 785 (13) 7352) 200) 5565 (20) (aloes (ka) 1752(0589_ 1850) 1652(1566_ 1847) 175 (1585-185) 16501585 1793) 1820(1620. 1891) 1782(1695. 1891) “NS Proteins TEV 165-12) 16UI5- 18) N66 19) GUIS 18) T6UIS-17) 1615-17) ONS Carbobyarates TEV S556) S865. 8)Sa(S3-58) © Sa(S3 Se) SSK Sc) SAIS 58) ONS Fas xTEV 2805.2) 275 28) 2810525) 2825.29) 8 BHD) Sate ft TEV 0 1 97-10) S(av10) nti) ior toto" “a0. Gholestera (ng) 215(170-258) 240(185-275) 104(173-279) 196(138-222)216(170-320) 218(174256) “NS Dietary fiber 3) 16(13-20) 1903-28) 163-19) 61220) 14K03-17) SATB) ONS sugars 1006 16) 9613) 31615) 8615) 146-18) 90-15) 8S ‘Maw, metabolealy Realy sormal weight: MH. metabolally Realy obese: MEOW, mcabo ally eahy overweight; MUO, mtaboalyunealthy obs [MUHOW setaailly wae overwelgt MW, mim wage! NS niga TY, tl energy vale Values exressd s median and interouarte ange or equeney( "Revs Wales *Minum wage 298.0 aan reals (US847n), Superript syns epost Pv between gous Pas between MAO and ther groups ‘agar between MEN and othe groups able2 ‘Anthropometric nical and bochemical ata variates ew (= 18) OW (= 17) MHO(a=29) MUHOW{a= 15) MURO(S=30), Weigh) 5805 (5225-6627) 705(656-8245) _BA55(8027-9768) __755(7035-801) __1012(on5 12297) BNC) 2138(2057-2340) ——2785(2047-2888) 3287(3105- 3624) 2837(2765-2885) _3638(3887 4642) Waistercunterence(em) _"74{6575 7825) 85(82.96) s9(003-1025) —853(838.8875) 10610712175) Neckcieumference(em) _ 31.75(31-3273) 34002-3825) 35(345-3775) 35G425.3780) —382(3638-263) Wat to bagi oay(o4i-o47) 033(050-058) 50(057-054), astiast 059) ‘65 (059-074) Fase (2) 213(1747-2425) 312(246 384) 3R5(348 4135) 509(259 3435) 4135(373.4433) Lean mass) 78217492235) 30616 754) G1A(SRA5 652) 616545 TH) 5835(5483 48) Syste BP 1175(100- 120) e010 120) 120(110-130), 120(1175-125), 120(120-130), (com a) DBiatleae 50(70.50) s0(725 80) 50(70.50) 50(80-80) ‘0(60.80) (om Hg) Total coleerol(mgldl) —1835(15675.21525) 196159. 239) woyr71-21ss) 215 2055-236) LDLc{msidl) 1075 (85513575) 107(835 154) 119(80-1315), iaa(izs 175) 125(1055- 1495) HDLC sit) 2(3875 70) 531475 5) 59(30_6t) s0(as.61) '35(4875. 6225) Tricity (msl) 5713075 975) ‘0,70. 122) 0s(655°1555) 106(735-1185) 1185 (855162), Fasting teem rl) 73(675 905) 78(735-86) 79(75-87) 4075539) 99(5125-95) Fasting m) 5125-125) 743-95) 8515-1125) 16(15-2): 195 1675-275): Howat 146(08- 223) 151(089 193) 174(103 235) 3400324416) 4161345 675) [a ody mas ox Pond presse: HDL, ih-densty Hpopotein cholexrok HOM, homcostass model assessment Of nstn resistance DLC low. dens "popreen cholesterol MIN, metabo eathy nmal weight MNO, metabolealy healthy obese; MHOW, metsbolaly Rely overacieht; MUHO, metabo ‘ueath bese; MUOM. metaoballyuneltyoxerreahe Vals expressed x median nd intreartle ange: Ks Wal text with ost hoc was ar out to anslze eve a nificance Speserpt sys represent Pale between BOS than the MHNW and MHOW groups. Moreover, considering, observed species, MUHO had lower values than all the other groups. Richness was significantly lower in the MUHOW group (Chaot, P < 0.05) than in the MHOW group and significantly lower (Chaot, P< 005) in the MHO group than in the MHOW group (Fig. 1, Supplementary Tables 1 and 2) ‘The similarity ofthe microbial communities of the five groups was evaluated by nMDS based on distribution of bacterial OTUs (3% dissimilarity: Fig. 2). The difference between groups (MHNW-MUHO-MHO-MUHOW.MHOW) was supported by AMOVA (F = 2.19, P= 0.03), as well a dferences between the fol- lowing pairs: MHNW and MHOW (F= 2.04, P= 0.046); MUHO and hitips:ireaderelsevier.comieadertspi'S0899900721001558 7token=63034DC8F477857EBCBFFSEAIE 1A38B120377S7DAASFEAIATECFBBF. 38s01nsi2021 Gut microbiota of adults wih diferent metabolic phenotypes | Elsevier Enhanced Reader 4 Pa Otvares tl Mutriton 902021) 111253 Fig. 1. boxplts representing difleencesin-dversty beween group sed on the umber of abseved specs, Shatin Chan, ad phylogenies nies tats ‘ei tests for comparison ofl rb are presented suppemenay material, *P O05; P= 01 P~ O01, MANW. meatal est nal weight MAO, met nial eat bese: MHOW metabo healthy overweight MUMO, metab unhealthy bes: MUNOW, metabolic nhelyaverweg BHI ig. 2. ak plt in wo dimension showing the dstrbtion of eat samples fete fie group according tothe compesiton ofthe teil coments Pos were bed on Bay-Cuts mare dste with 97 slaty met MEO, eabocally healthy abe: MHOW, metabolic healthy overveht: MUHO, ‘metabolically saheahy otese: MUBOW, meatal atealhy overweight MOS, poamescmulimensoal sealing MUHOW (F » 3.54, P = 0.048); and MENW and MUHO (F = 3.97, 028; Supplementary Table 3). ‘The composition of all bacterial communities was dominated by Firmicutes and Bacteroidetes, which contributed to 78% to 91% of the community. The F/B ratio was ower in the MUHO group. How- ever, there was no difference inthis relationship between the other metabolic phenotypes (Fig. 3A, Supplementary Table 4). The relative abundance of Actinobacteria accounted for 4% to 11% and that of Proteobacteria ranged from 3% to 8% (Fig. A). The most abundant orders were Clostridiales (43-38%) and Bacteroidales (14-43%; ), whereas the most represented genera were Prevo- fella9, Bacteroides, Eubacterium rectale, and Blautia. Compared ‘with other groups, Dialister was more abundant in the MUHOW ‘group. and Bifidobacterium was more abundant in the MENW sroup (Fig. 3). ‘The proportions of shared and exclusive OTUs attributed to ‘each group are represented in Venn diagrams (Supplementary Fi 2}. Considering all groups. 34% of all designated OTUs were shared by all, and between 5% and 9% were exclusive to each group. With respect to MHOW and MUHOW, 60% of OTUs were shared, ‘whereas 23% and 17% were exclusive to MHOW and MUHOW, respectively. Regarding MHO and MUHO, 60% of OTUs were shared, whereas 16% were exclusive to MUHO, and 24% were ‘exclusive to MHO, Unique OTUs corresponded to taxonomic ‘groups with lower felative contribution to the composition of the ‘community (1%). In comparing groups with healthy phenotypes (MHNW, MHOW, and MHO), no differences were observed in the relative abundance of Firmicutes and the corresponding taxonomic levels, class Clostridia, order Clostridiales, and families Lachnospiraceae and Ruminacaccaceae (supplementary Fig. 3). The phylum Bacteroi- ddetes and related taxonomic levels, class Bacteroidi, order Bacter- ‘idales, and family Prevotelaceae were more abundant in MHO and MHOW than in MHNW. More detailed analysis revealed the main OTUs contributing to distinctions between MHNW and MHOW, as well as between MHNW and MHO, estimated by SIM- PER analysis (Supplementary Fig. 4), The genera Prevotela 9, Prevo- tellaceae UCGO03, uecinvibrio, and Eubacterium rectale were more hitips:ireaderelsevier.comieadertspiv'S0898900721001558 7token=63034DC8F477B57EBCBFFSEAIE 1A38B12037737DAASFEAIATECFBBF. 418s01nsi2021 Gut microbiota of adults wih diferent metabolic phenotypes | Elsevier Enhanced Reader 4 Ores 5202111283 5 ER «con | mt caus mm os St I EE ess “+ Cine | oo ee A ae ear ome SS MA ie g.3. OTV composition ofthe Five stay groups atthe (A) pum, {011 of relate abundance are represented. Fr genus, ely the ores. an (Cems eves. For phylum and order. only the major taxonomic groups that conte mln taxonomic group are represented WHO, metal heathy asses MO, metabolically bly overweghr: MUH, metabo untealty bes: MUNOW, metaeleallyunheslhyoveraight- UL operations fexonamc wi abundant in MHO in comparison with MHNW. The genera Prevo- tellacene UCGO03, Faecalibacterium, and Eubacterium recrale were ‘more abundant in MHOW in comparison with MHNW. The genera Bifidobacterium and Butyrivibrio were more abundant in MHNW ‘than in MHOW and MHO. ‘We also identified taxonomic groups that showed significant differences in relative contribution (¢ test, P< 0.05) of MHOW and [MUHOW, in several taxonomic categories (Fig. 4, Supplementary Table 5). The following taxa showed greater abundance in [MUHOW than in MHOW: class Bacilli, order Lactobacillals, genera Paraprevotella and Ruminococcaceae UCG-010. The most abundant taxa in MHOW included the genera Coprococcus and Ruminococcus. Regarding the obese groups, the taxa that differed between MHO and MUHO were minorities, contributing to <1% of relative abun- ‘dance, likely representing transitory members ofthe communities In general, individuals with metabolically healthy phenotypes hhad lower anthropometric measurements, in comparison with their unhealthy pairs. Pest hoc analysis revealed significant dffer- cences in fasting. glucose and serum insulin between MHO and MUHO, These results are similar to those observed by other researchers [10,17] Significantly higher serum concentrations of HDL cholesterol (HDL-C) have been reported in MHO, in comparison with MUHO (18) Similarly, although the difference was not statistically signift- ‘ant, we found that the MHO group showed higher concentrations of HDL-C than the MUHO group. This may be related to greater alcohol intake [19] and higher level of physical activity in this ‘group (20). Slagter et al, also found that individuals who were [MHO were more frequently moderate alcohol users [21 in con- trast with the findings of Goday et al, who found that individuals ‘who were MHO were less likely to consume alcohol (22) Other factors, such as physical activity and healthy dietary hab- its, have been associated with a healthier phenotype in previous studies [21,22]. n the present study, we observed statistical signifi- cance only in saturated fat intake, which was lower in the healthy phenotype. Similarly, some studies did not reveal significant dffer- fences in level of piysical activity and dietary intake between ‘metabolically healthy and unhealthy adults who were overweight, or obese [23,24], which stimulates discussion regarding other pos- sible factors involved, When analyzing the gut microbiota of the participants, our results revealed reduced bacterial richness and diversity with increased body weight and lower diversity in groups with unhealthy phenotypes compared with the healthy ones, which vwas consistent with other studies [7.25]. This decrease in diversity ‘may modify the functional profile ofthe microbiota. In analyzing the two dominant phyla, no difference was observed in the relative abundance of Firmicutes between groups. In contrast, the phylum Bacteroidetes was more abundant in indi- Viduals wo were overweight or obese compared with those who were normal weight. The Felatonship between these two phyla, expressed as the F/B ratio, has been associated with several patho logic conditions and often is considered as a possible signature of obesity [25] Inthe present study, we found no significant differ- ence in this relationship. Perhaps the small number of participants included has made statistical power irrelevant for detecting small, variations, ‘Some studies have reported an increased FYB rato in the micro- biota of obese individuals [27] compared with lean individuals, whereas other studies have observed no difference [25], or they observed an opposite association, with increased Bacteroidetes in individuals who were overweight and obese [28] and increased Firmicutes in lean individuals [29]. Moreno-Navarrete et al. on the other hand, observed a lower F/B ratio in obese individuals who were resistant to insulin compared with those who were sensitive to insulin (30) Inconsistencies among studies may result from het- exogeneity of groups, genetic history, geographic location, habitual lifestyle, diet, and the methodology used. Many of these studies did not control factors such as dietary intake, medication use, and physical activity level; furthermore, they included elderly individ uals in their population or had significantly different percentages, of mea ina given group. Iti also likely that compositional changes, at more specific levels, such as genus or species, maybe more rele~ vant than the FB ratio for association witha given health condition [26 Increased abundance of gram-negative bacteria Bacteroidetes), hich express liposaccharides, is consistent with the hypothesis of hitips:ireaderelsevier-comieader!spiv'S0899900721001558 7token=63034DC8F477857EBCBFFSEAIE 1A38B120377S7DAASFEAIATCFBBF. 58soos cette in tre eta herp] har nha Ree moe no —_ vc = 7 Se a ' Coprocoeas F io ce * Meansin groups” °" —‘Biference baween grouns.”” men ems oo —— = ay * Meansingroups"” ** —Biference between groups” vow EB MuHow ~ ‘90% cantidence eva —_— coo | ~ | ot ‘Means in groups : Difference between groups ig tarsal analss( test) determine OTUs with sigan vari in eae abundance in KHON and MUMOWY (P< 0.0) nario taenom categories. ars ‘epresent sverige valves of TU abundance meach grap, On the ight, mean P-value of spec or variation Between groups and 3 confidence tervals wth ower { uppe iit MHOW. metabolic healhy overweight: MUKOW, metabolically unhealthy overweight CTU. operational taxonomic nit ‘metabolic endotoxemia, with low-degtee inflammation, character- istic of obesity (31. thas been suggested that Bacteroidetes are regulated mainly by environmental factors, whereas Firmicutes depend on the sgenetic composition ofthe host [32] Reyes et al. discovered that the abundance of Bacteroidetes was correlated with fiber intake [29], which is coherent with our results. Reduced consumption of fibers has also been indicated as an important contributing factor toss of microbial richness and diversity [33], as has elevated con- sumption of saturated ft 34), ‘The genus Bifidobacterium was more abundant in the MHNW ‘group compared with MHOW and MHO, whieh is consistent with the literature [7,29], Abundance of bifidobacteria has been associ- ated with reduced inflammatory markers, improved homeostasis ‘of glucose and lipid metabolism [35], and higher fiber intake (29) ‘Another intresting finding of ths Study was the observed inerease {nthe genera of the family Prevotellaceae and the genera Eubacterim rectale and Faeealbacerium in the MHOW and MHO groups, in com- parison with MHNW. Although greater abundance ofPrevoteli as fre- ‘quently been associated with health [36] anda dit rich in fibers [37 an association wit obesity has also been reported [38,39] It has been ‘suggested that this genus is more efficient in extracting the energy of ‘complex polysaccharides [20], contributing tothe development of abe- sity inthe high-calorie Westem diet (39) Eubacterum rectale and Fae- calbacterium are important producers of butyrate, and, although a large body of evidence has suggested potential health benefits, the results have been contradictory and inconclusive in invuals who are obese [41 Comparing the MHOW and MUHOW groups, we identified taxa ‘with significant differences in relative abundance, Some are partic- ularly interesting because they are linked to human health, such as, the genera Coprococcus and Rurninacoceus, which are producers of butyrate and which were more abundant in MHOW, Butyrate has anti-inflammatory potential; it prevents endotoxemia, improves insulin sensitivity, and increases intestinal barrier function [42 A ‘dysfunctional junction ofthe intestinal barter allows components ‘derived from the gut microbiota, such as liposaccharides, o enter the bloodstream, which may exacerbate inflammation of adipose tissue [43]. Org et al. also reported an association between the ‘genus Coprococcus and lower TC levels [6 Unhealthy phenotypes show greater relative abundance of patho- ‘genic bacteria that may compromise intestinal bartier function. In gen- feral, the order Lactobacilales and the class Bacili, bacteria that produce lactic acid, are considered beneficial, but an increased abun- ‘dance ofthese taxa has also been observed in individuals with coro- nary artery disease (44) diabetes [45), and obesity [725] In previous studies, elevated abundance of Akkermansia was associated with healthy metabolism in individuals who were over- ‘weight [46], while the genera Pseudomonas, Prevotela, Veillonela, land Fusobacterium were mote abundant in obese individuals hitips:ireaderelsevier.comieadertspiv'S0899900721001556 7token=63034DC8F477857EBCBFFSEAIE 1A38B12037737DAASFEAIATECFBBF. 68soinsi2021 Gut microbiota of adults wih diferent metabolic phenotypes | Elsevier Enhanced Reader PAS Olivares eta Nurion 902021) 111293, 7 resistant to insulin (47); nevertheless, our study did not abserve ny difference in these taxa. Manar et al. suggested that certain associations between bacterial species and host factors are the result of specific arrangements inthe microbial community, which are not always determined by individual taxa, and that the same ‘genus may have different implications for health and disease [48], Some taxa, which differed in abundance between our study soups, have been litte studied in humans, and their functional ‘ole is still not clear. In common with other studies, we identified a decrease in butyrate-producing bacterial groups and an increase in ‘opportunistic pathogens in individuals with metabolic disorders. Ie is important to note that it was not possible to consider all confounding factors, such as the hormonal phase of the women included in the study, given that estrogen levels can influence the ‘composition ofthe gut microbiota and cause metabolic and immu- rnologic changes [49,50]. Although differences in the composition ‘of gut microbiota between sexes have been described in the litera- ture, the proportion of men and women was similar in all groups inthis study, and we did not identity gender dimorphism. Study limitations ‘The small sample size was a limitation ofthe present study. Fur~ ‘ther studies with larger numbers of individuals are needed to con- firm our results Study strengths ‘This was a pioneer study in Brazil, comparing the gut micro- biota of metabolically healthy and unhealthy individuals who are ‘bese and overweight. Conelusion ‘We found thatthe intestines of overweight and obese individuals with metabolic disorders hosted inflammation-associated microbiota, ‘ith lower butyrate production potential and reduced bacterial diver- sity, along with an increase in opportunistic pathogens. Supplementary materials Supplementary material associated with this article can be found inthe online version at doi:10.1016jj.ut.2021.111293. References [1 Jr Pav. 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