THE MICROBIOME IN HIV
Cindy Monaco, MD, PhDUniversity of Rochester School of
Medicine and Dentistry
No Disclosures
Outline
• What is the microbiome?
• Why does it matter?
• What is the relationship between the microbiome and HIV?
What is human?
• We are comprised of more than just human cells
http://scalar.usc.edu/works/lope/diagram-of-human-body
What is human?
• We are an ecosystem of bacteria, viruses, fungi, and human cells
• Human cells are outnumbered at least 3:1, not including viruses
• Genes from our microbes impact our health
Mpkb.org
What do these microbes do?
• Commensal microbes (microbes that live within us) can protect against diseases:
– Parasitic worm infection prevents allergies
– Skin/vaginal yeast infections after antibiotics
– C diff colitis after antibiotic use
– Bacterial vaginosis (BV) associated with increased risk of STIs including HSV, HIV, etc.
What do these other organisms do?
• The majority of organisms that colonize us are not able to be cultured
• How can we study those?
• Advent of NGS (next generation sequencing) aka deep sequencing, second generation sequencing– allows us to discover the unculturable microbes
at various body sites
• Dawn of “microbiome” research
Why has it taken so long to study the microbiome?
Price per base of DNA sequencing• 1869: DNA discovered• 1953: DNA revealed to be
genetic blueprint• 1970s: RNA sequencing
developed• 1977: first full DNA
sequence (bacteriophage phiX174) by Sanger sequencing (chain termination)
• 2001: human genome published (Sanger sequencing) after > 10 year effort
• 2005: first next generation sequencing (NGS) machine introduced
Rob Carlson, 3/2016, www.synthesis.cc
Sanger sequencing NGS sequencing
Microbiome components - Definitions
• Bacteria: – Microscopic (i.e. can see using microscope)
– single celled organisms
– DNA genome
– Come in various shapes
– Free-living (i.e. do not require another host cell machinery to live)
– Some require oxygen, others do not
– Differentiated by type of cell wall into gram positive and gram negative
– Can be treated with antibiotics
Microbiome components - Definitions
• Viruses– Submicroscopic (i.e. cannot see by microscope)
– Infectious agent
– Replicates only inside living cells of other organisms
– DNA or RNA genomes
– Eukaryotic viruses: viruses that infect animals or plants• Few viruses have effective treatments (HIV, HCV, Flu, CMV)
• Usually rely on vaccination (HPV, Flu, chickenpox/zoster, rotavirus, yellow fever, measles/mumps/rubella)
– Bacteriophage: viruses that infect bacteria• can be either lysogenic (replicate harmlessly in the bacteria)
• Or lytic (break open and destroy bacteria after replication)
• Can carry antibiotic resistance or toxin genes to other bacteria
Microbiome components - Definitions
• Fungus: multicellular organisms like yeast or molds
• Other prokaryotes: i.e amoeba, helminth, tapeworms, etc
What is the microbiome?
• Microbiome: all bacteria, archaea, viruses, fungi, and other microbial Eukarya
• Bacteriome: bacterial components of the microbiome
• Virome: viral microbiome, including eukaryotic viruses, bacteriophage, endogenous retroviruses– Estimated as little as 1%
sequenced
Virgin HW. Cell. 157, 2014; Clemente JC et al, Cell. 148, 2012; Cox MJ et al, Hum Mol Genet, 22, 2013
What is the microbiome?
Blum HE. Adv in Med Sci. (2017), 62
• Defined by body site
• Unique between individuals
• Diversity greater between sites than between individuals
Clemente JC et al, Cell. 148, 2012; Muszr M et al, Arch Immunol Ther. Exp (2015), 63.
Bacterial microbiome composition by body site
The bacteriome varies by age
Ottman, N et al. Aug 2012 Frontiers in Cellular and Infection Microbiology
Virome
• Much less known about the viral components of the microbiome
– lack of adequate culture methods
– highly divergent sequences between viral species
– prevents development of broad PCR-based assays
• Culture-independent, high-throughput sequencing methods
– unbiased look at the composition of the microbiome
– discovering potentially novel, unculturable viruses
Virgin HW. Cell. 157, 2014;
What is a typical gut bacteriome?
• Human gastrointestinal tract is colonized by trillions of microorganisms
• Differs between the developed world and agrarian cultures
• Prevotella-rich and Bacteroides-poor community associated with agrarian cultures
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Prevotella
Bacteroides
De Filippo et al., PNAS, 2010
What can alter the microbiome?
• Diet– Plant versus animal based– Even artificial sweetener
• Medications, especially antibiotics or antivirals, heartburn medications
• Sexual practices – Anal intercourse associated with increased Prevotella in fecal samples – Differences in vaginal microbiome
• Hormones and hormonal contraceptives• Immunosuppression (steroids, AIDS, cancers)• Some environmental factors (pets, family members)• Hygiene practices (skin microbiome with antimicrobial soaps,
vaginal microbiome and douching)
Why Does the microbiome matter?
Effect of intestinal bacterial microbiome in health
• For a long time we have known that diet can affect many diseases in including aging, cardiovascular disease, dementia– May be due to the gut bacterial microbiome
• Intestinal bacterial communities and their metabolites– shape and maintain gut immunity– influence systemic immune response– alter behavior– produce metabolites used by us as energy source. – promote gut epithelial barrier integrity
Dillon SM et al, AIDS, 2017 31:511-521
Alterations in the enteric microbiomeare associated with disease
• Metabolic disorders such as obesity and diabetes mellitus– Fecal transplant from obese mice to lean mice led to
weight increase despite no change in diet.
• Neuropsychiatric disorders such as schizophrenia, autistic disorders, anxiety disorders and major depressive disorders.
• Gut-brain axis– signals originating in the gut can also influence brain
function through interface with the enteric and central nervous systems
– gut microorganisms are capable of producing and delivering neurotransmitters such as serotonin and GABA
Evrensel A and Ceylan ME. Clin Psychopharmacol Neurosci. 2015 Dec; 13(3): 239–244; Perry RJ et al, Nature. 2016 June; 534: 213–217
The gut microbiome and stroke
• Ischemic stroke – highly prevalent disease – limited therapeutic options– Inflammation is key component
• Treated mice with augmentin (a PCN antibiotic) for 2 weeks – decreased diversity of intestinal
bacteriome
• Then induced transient middle cerebral artery occlusion (MCAO). – 60% reduction in stroke volume in
Augmentin-treated mice.
• Similar effects with oral vancomycin• Ciprofloxacin and Flagyl treatment
reduced survival following MCAO• Suggesting that particular microbial
species cause differential effects on brain injury recovery
Benakis C et al, Nature Medicine volume22 , pages516–523 (2016)
Control Augmentin
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Control Augmentin
The gut microbiome and Austism
• Autism spectrum disorders (ASDs), a complex neurobiological disorder– impaired social interactions and communication – leads to restricted, repetitive, and stereotyped patterns of behavior, interests,
and activities
• Causes of poorly understood – Complex interplay of genetic and environmental factors
• Many also experience significant GI symptoms, which correlate with ASD severity– Many ASD kids have gotten more Abx therapy in first 3 years of life
• Several studies showed altered gut bacterial microbiomes in ASD kids compared with neurotypcial children– lower abundances of fermentative bacteria (e.g., Prevotella copri)– lower overall bacterial diversity
• ASD mouse model demonstrated that Bacteroides fragilis treatment could alter gut microbiota and blood metabolite profiles, correct increased gut permeability, and improve ASD symptoms
Kang DW et al, Microbiome20175:10; Fung TC et al, Nature Neuroscience volume20, pages145–155 (2017)
The gut microbiome and Austism
• 2 open-label studies in children with ASD, – 8 weeks of oral vancomycin transient
improvements in both GI symptoms and ASD symptoms
– Fecal Transplant: 2-week antibiotic treatment, a bowel cleanse, and then an extended fecal microbiota transplant (FMT) using a high initial dose followed by daily lower maintenance doses for 7–8 weeks. • 80% reduction of GI symptoms at the end of treatment,
persisted 8 weeks after treatment.
• Significantly improved behavioral ASD symptoms, persisted 8 weeks after treatment ended.
Kang DW et al, Microbiome20175:10; Fung TC et al, Nature Neuroscience volume20, pages145–155 (2017)
Inflammatory Bowel Disease linked to alterations in enteric bacteriome and virome
Norman JM,…, Monaco CL, et al. Cell, 2015
Gut microbiome may predict response to chemotherapy medications
• Melanoma patients undergoing anti-PD-1 immunotherapy (n=112)
• Significantly higher gut bacterial diversity in patients who responded to therapy (R)
• Significant difference in mortality depending on gut bacterial diversity
Gopalkrishnan V et al, Gut microbiome modulates response to anti-PD-1 immunotherapy in melanoma patients, Science. 2018 Jan 5;359(6371):97-103.
Div
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Diversity
Compositional differences in the gut
microbiome are associated with responses to anti-PD-1 immunotherapy
• Higher relative abundance of Ruminococcaceae bacteria (p<0.01) in responding patients.
• Metagenomic studies revealed functional differences in gut bacteria in responders (R) including enrichment of anabolic pathways.
• Immune profiling suggested enhanced systemic and anti-tumor immunity in responders
• Verified results in gnotobioticmice (germ-free mice) with transplanted fecal microbiomes
• This has profound implications for all chemotherapy
Gopalkrishnan V et al, Gut microbiome modulates response to anti-PD-1 immunotherapy in melanoma patients, Science. 2018 Jan 5;359(6371):97-103.
Cardiovascular disease and the gut microbiome
Blum HE. Adv in Med Sci. 62 (2), 2017.
THE MICROBIOME AND HIV
HIV• Major global public health issue
– > 39 million dead
– ~ 36 million people living with HIV
• Sub-Saharan Africa disproportionally affected– 66% of new HIV infections globally
– 24.7 million people with HIV in 2013
• ART has transformed HIV from a fatal disease to a chronic condition
WHO 2014.; Maartens G et al, Lancet, 2014; Brenchley JM. Mucosal Immunology. 2013; SMART study et al, NEJM, 355 (2006);
Prevalence of HIV/AIDs in Africa (% population in 2011) (World Bank)
Over 15%5-15%2-5%1-2%0.5-1%0.1-0.5%Insufficient data
Uganda
HIV
• ~ 50% of deaths in people on ART in developed world are not due to AIDS– non-AIDS- defining cancers (23·5%),
cardiovascular disease (15·7%), and liver disease (14·1%).
– 50% increased risk of MI after adjustment for vascular risk factors
– ART does not fully correct this risk
• May be a consequence of chronic immune activation– Linked to CVD, renal, bone disease and
increased mortality
– Uncertain mechanism, microbial products likely involved
Maartens G, Celum C, and Lewin SR, HIV infection: epidemiology, pathogenesis, treatment and prevention, Lancet, 2014; Brenchley JM. Mucosal Immunology. 2013, April 1-9; Marchetti G, et al. Clin Micro Rev. 2013, 26(1): 2-18;
Why look at the microbiome and HIV?
• SIV-infected vs uninfected macaque monkeys – same diet, separately housed – 24 wks post intra-rectal SIV infection
• Pathogenic SIV is associated with an expanded enteric virome– >10-fold increase in viral sequences – New viruses found– Expansion of adenoviruses – Adenovirus-positive enteric
epithelial cells assoc with enteropathy (damage to gut)
• Bacterial microbiome was unchanged
• No difference in microbiome in non-pathogenic infection of African green monkeys
• Alterations in the enteric virome may be important in the pathogenesis of AIDS
Handley S et al., Cell. 2012
The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vesselCD4Tcell
CD4Tcell
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vesselCD4Tcell
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Veazey et al, Science 1998
Pe
rip
her
al L
N
G
ALT
Bacteria
Bacteriophage
CD4Tcell
The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
sCD14:- LPS binding protein- increased levels in HIV- marker of microbial translocation- Linked to increased mortality in HIVSandler et al, JID, 2011
Bacteria
Bacteriophage
HIV gut microbiome studies
Gootenberg DB et al, Current Opinion in Infectious Diseases, 2017
Effect of intestinal microbiome in HIV
• Fecal microbiome in most studies shows increases in potentially “pathogenic” bacterial families with decreases in “beneficial” bacterial
– In western world see increase Prevotella species, Firmicutes and Bacteroides reduced
– In developing world, Prevotella species already high (likely due to higher fiber/plant diet)
Dillon SM et al, AIDS, 2017 31:511-521
Purpose of this study
• What is the effect of HIV, ART, and immune status on the enteric virome and bacteriome?
Monaco CL et al, Cell Host and Microbe, 2016
ISS clinic in Mbarara, Uganda• Founded November 1998
• Initially compassionate care to HIV-positive patients
• UARTO cohort: Uganda AIDS Rural Treatment Outcomes
– 40 HIV negative subjects
– 40 HIV-infected on ART ≥ 5 yrs
– 42 HIV-infected, ART naïve
– HIV neg and HIV untreated enrolled on initial test for HIV
– Matched stool and plasma
Monaco CL et al, Cell Host and Microbe, 2016
CD4 T cell count correlates with sCD14
Monaco CL et al, Cell Host and Microbe, 2016
Adenoviridae and Anelloviridae are expanded in AIDS
Monaco CL et al, Cell Host and Microbe, 2016
Bacterial richness and diversity differ by CD4 T cell count
Richness Diversity
Monaco CL et al, Cell Host and Microbe, 2016
Discriminant Bacterial Families
Monaco CL et al, Cell Host and Microbe, 2016
HIV Negative versus CD4 T cell count < 200
Conclusions• CD4 < 200 was associated with changes in the
enteric DNA virome • Adenoviridae sequences were increased in
patient with advanced HIV disease– Similar to SIV
• CD4 <200 associated with decreased bacterial richness and diversity
• Enterobacteriaceae associated with low CD4 T cell count– Similar to SIV
• Decreased Ruminococcaceae associated with CD4 <200 in multivariate model
Model of gut in HIV
Gut Lumen
Blood vesselCD4Tcell
CD4Tcell
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
• Severe immunodeficiency destabilizes enteric commensals
• Intestinal dysfunction and disease
• Treatment of enteric pathogens, in conjunction with ART, may limit HIV-associated enteropathy
• May minimize systemic immune activation
Shigella
Ruminococcus
Gut epithelium
Model of gut in HIV
Gut Lumen
Blood vesselCD4Tcell
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
• Severe immunodeficiency destabilizes enteric commensals
• Intestinal dysfunction and disease
• Treatment of enteric pathogens, in conjunction with ART, may limit HIV-associated enteropathy
• May minimize systemic immune activation
Shigella
Ruminococcus
Gut epithelium
Model of gut in HIV
Gut Lumen
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
• Severe immunodeficiency destabilizes enteric commensals
• Intestinal dysfunction and disease
• Treatment of enteric pathogens, in conjunction with ART, may limit HIV-associated enteropathy
• May minimize systemic immune activation
Shigella
Ruminococcus
Gut epithelium
Model of gut in HIV
Gut Lumen
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
• Severe immunodeficiency destabilizes enteric commensals
• Intestinal dysfunction and disease
• Treatment of enteric pathogens, in conjunction with ART, may limit HIV-associated enteropathy
• May minimize systemic immune activation
Shigella
Ruminococcus
Gut epithelium
Model of gut in HIV
Gut Lumen
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
• Severe immunodeficiency destabilizes enteric commensals
• Intestinal dysfunction and disease
• Treatment of enteric pathogens, in conjunction with ART, may limit HIV-associated enteropathy
• May minimize systemic immune activation
Shigella
Ruminococcus
Gut epithelium
Model of gut in HIV
Gut Lumen
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
• Severe immunodeficiency destabilizes enteric commensals
• Intestinal dysfunction and disease
• Treatment of enteric pathogens, in conjunction with ART, may limit HIV-associated enteropathy
• May minimize systemic immune activation
Shigella
Ruminococcus
Gut epithelium
Model of gut in HIV
Gut Lumen
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
• Severe immunodeficiency destabilizes enteric commensals
• Intestinal dysfunction and disease
• Treatment of enteric pathogens, in conjunction with ART, may limit HIV-associated enteropathy
• May minimize systemic immune activation
Shigella
Ruminococcus
Gut epithelium
What about the vaginal microbiome?
• Greater than 90% of HIV transmission worldwide occurs following heterosexual intercourse– Women are twice as likely to contract HIV as men
• Young African women have up to 8-fold increased HIV prevalence compared to young men
• The vaginal microbiota plays an important protective role in preventing– pre-term birth– bacterial vaginosis (BV)– yeast infections– sexually transmitted diseases including HIV
M. I. Petrova et al, FEMS microbiology reviews 37, 762-792 (2013); B. A. White et al, Trends in endocrinology and metabolism: TEM 22, 389-393 (2011);
What is the vaginal microbiome?
• There is no single core vaginal bacteriome, but instead several major community groupings distinguished by whether they are Lactobacillus dominant or deficient
• Lactobacilli inhibit invasive bacterial species through– reduction of vaginal pH
– Competing for nutrients and adherence to the vaginal epithelium
• Pathological changes such as the dysbiosis associated with bacterial vaginosis (BV)– Can occur rapidly
– Almost always accompanied by depletion of lactobacilli
M. I. Petrova et al, FEMS microbiology reviews 37, 762-792 (2013); B. A. White et al, Trends in endocrinology and metabolism: TEM 22, 389-393 (2011); J. J. Schellenberg, F. A. Plummer, International journal of inflammation 2012, 131243 (2012); H. Borgdorff et al., The ISME journal 8, 1781-1793 (2014).
Vaginal microbiome in HIV
• BV is directly associated with
– increased inflammation in the vagina
– increased risk of STIs including HPV, HSV, and HIV
• HIV-infected women with altered vaginal bacterial flora (BV)
– have higher concentrations of HIV-1 RNA in genital secretions
– associated with a 3-fold increased risk of HIV transmission to a discordant male partner
M. I. Petrova et al, FEMS microbiology reviews 37, 762-792 (2013); B. A. White et al, Trends in endocrinology and metabolism: TEM 22, 389-393 (2011); J. J. Schellenberg, F. A. Plummer, International journal of inflammation 2012, 131243 (2012); H. Borgdorff et al., The ISME journal 8, 1781-1793 (2014).
Lactobacillus-Deficient Vaginal Bacteriomes Are Associated with Increased HIV Acquisition
• FRESH cohort in South Africa
• 4 times higher rate of HIV acquisition in women with high diversity vaginal bacteriomes
• Higher levels of activated mucosal CD4 T cells are higher in women with high-risk bacteria (Prevotella and other anaerobes)
• Lower levels in women with Lactobacillus crispatus
Gosmann C et al, Immunity, 46(1): 29-37, 2017
Microbiome and PREP
• In clinical trials of women the efficacy of PREP was poor
• CAPRISA 004 trial showed 39% efficacy for coital vaginally-applied tenofovir gel
• Variable adherence = major contributing factor
• Why is higher adherence required in women?
Klatt, NR et al, Science 02 Jun 2017: Vol. 356, Issue 6341, pp. 938-945
HIV infection rate with vaginal Lactobacillus dominance and
non-Lactobacillus bacterial dominance
Klatt, NR et al, Science 02 Jun 2017: Vol. 356, Issue 6341, pp. 938-945
• Separated into Lactobacillus dominant (LD) or non-Lactobacillus dominant (non-LD) bacteriome
• LD group: HIV incidence rate was 61% lower in women assigned to tenofovir gel versus placebo gel [P = 0.013]
• Non-LD: HIV incidence rate was 18% lower with tenofovir gel versus placebo gel (P = 0.644)
• The efficacy of tenofovir gel in preventing HIV infection in the subgroup of women with >50% adherence: 78% (P = 0.003) in the LD group versus 26% (P = 0.558) in the non-LD group.
• Tenofovir concentrations significantly lower in non-LD vs LD women (P = 0.0077).
Metabolism of tenofovir by G. vaginalis and BV-
associated bacteria
Klatt, NR et al, Science 02 Jun 2017: Vol. 356, Issue 6341, pp. 938-945
• Genital tenofovir concentrations negatively correlated with G. vaginalis protein abundance (correlation coefficient r = –0.19, P = 0.0014) and other anaerobic bacteria (Prevotella, r = –0.14, P = 0.023)
• Suggests relationship between BV-associated bacteria and tenofovir levels.• Used an in vitro culture system to assess potential metabolism of tenofovir by bacteria• Tenofovir concentrations in culture with G. vaginalis decreased rapidly by 50.6% compared
with marginal changes in either L. iners (P = 0.0037), L. crispatus (P = 0.0019), or control (P < 0.0001) at 4 hours
• Adenine, metabolite of tenofovir, only formed in presence of G. vaginalis• Microbiome metabolism of drugs has profound implications for treatment and prevention of
HIV
Limitations
• Main way used to examine bacterial microbiome is 16S rRNAsequencing, where small part of a region of the bacterial ribosome is amplified
• Sequencing different regions of 16S rRNA used may identify different bacteria (different results)
• Difficult to get species level identification of bacteria, usually more family level– Need species level for therapy
• All microbiome studies are associative – unless do in vivo or in vitrostudies to validate findings
• With shot-gun sequencing for viruses– Dependent on database used for comparison – need similar sequence
to identify– large amount of sequence data is “dark matter” – no homology to
known organisms– Sequencing depth vs cost issue
Conclusions
• Vaginal and enteric bacterial microbiomesdiffer in HIV infection from healthy control
• May offer novel therapeutic options to
– Prevent chronic inflammation in HIV
– Help prevent transmission and acquisition of infection
• Still a lot of work to be done
QUESTIONS?
Ways to measure intestinal microbiome
• Keep in mind that gut bacteria composition varies depending on location (Stomach vs upper small intestine vs middle vs lower small intestine vs colon)
• Fecal (stool or rectal swabs)– Easiest to collect– May not represent mucosal bacterial population
• Weck-cel (or similar)– Swab to rectal mucosal surface, have to leave in a little bit– Can get mucosal bacterial communities
• Biopsy from colonoscopy– Theoretically measures mucosal adherent bacteria– But only get after colonic prep – wash out a lot of bacteria
• Swab/fecal sample during colonoscopy– Same as above
Uganda Cohort CharacteristicsPatient Characteristics
HIV Neg(n = 40)
HIV on ART(n = 40)
HIV no treatment(n = 42) P value
16S rRNA sequencing (n) (total 110) 37 39 34VLP NGS sequencing (n) (total 65) 21 21 23Demographics, median (IQR)
Age (years) 43 (38-48) 44 (38-49) 29 (24-34) < 0.0001Males, n (%) 20 (50) 20 (50) 11 (26.2) 0.0404BMI (SD) 24 (22-28) 24 (21-27) 23 (20-27) 0.4305
Laboratory measures, median (IQR)CD4 T cell count (cells/µl) NA 396 (283-490) 225 (113-382) 0.0003
>500, n (%)* NA 8 (20) 4 (9.5)200-500, n (%)* NA 29 (72.5) 16 (38.1)<200, n (%) NA 3 (7.5) 22 (52.4)
CD4 Percent NA 25 (21-31) 15 (10-24) < 0.0001HIV Viral Load (copies/ml) NA 20 (20-20) 95,571 (24455-285548) < 0.0001CD4 nadir (cells/µl) NA 116 (58-167) 225 (110-382) 0.0001
Symptoms over last 30 days
Nausea/Vomiting, n (%) 13 (32.5) 5 (12.5) 15 (35.7) 0.0389
Diarrhea, n (%) 10 (25) 4 (10) 9 (21) 0.2000
Constipation, n (%) 17 (42.5) 13 (32.5) 10 (23.8) 0.1969
Loss of Appetite, n (%) 21 (52.5) 10 (25) 20 (47.6) 0.0286
Medications last 30dNRTI, n (%) NA 40 (100) NANNRTIn, n (%) NA 35 (85) NAYears on ART, median (IQR) NA 6.7 (6.1-7.1) NABactrim last 30d, n (%) 0 (0) 38 (95) 38 (90.5) < 0.0001Other Antimicrobials 30d, n (%) 15 (37.5) 14 (35) 4 (9.5) 0.0066
Monaco CL et al, Cell Host and Microbe, 2016