THE MICROBIOME IN · Alterations in the enteric microbiome are associated with disease •Metabolic...
Transcript of THE MICROBIOME IN · Alterations in the enteric microbiome are associated with disease •Metabolic...
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THE MICROBIOME IN HIV
Cindy Monaco, MD, PhDUniversity of Rochester School of
Medicine and Dentistry
No Disclosures
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Outline
• What is the microbiome?
• Why does it matter?
• What is the relationship between the microbiome and HIV?
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What is human?
• We are comprised of more than just human cells
http://scalar.usc.edu/works/lope/diagram-of-human-body
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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
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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.
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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
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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
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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
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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
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Microbiome components - Definitions
• Fungus: multicellular organisms like yeast or molds
• Other prokaryotes: i.e amoeba, helminth, tapeworms, etc
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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
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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
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The bacteriome varies by age
Ottman, N et al. Aug 2012 Frontiers in Cellular and Infection Microbiology
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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;
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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
Bu
rkina Faso
Euro
pe
Prevotella
Bacteroides
De Filippo et al., PNAS, 2010
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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)
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Why Does the microbiome matter?
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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
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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
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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
Div
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Au
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Co
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Control Augmentin
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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)
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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)
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Inflammatory Bowel Disease linked to alterations in enteric bacteriome and virome
Norman JM,…, Monaco CL, et al. Cell, 2015
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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
ersi
ty
Diversity
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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.
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Cardiovascular disease and the gut microbiome
Blum HE. Adv in Med Sci. 62 (2), 2017.
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THE MICROBIOME AND HIV
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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
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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;
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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
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The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vesselCD4Tcell
CD4Tcell
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
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The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vesselCD4Tcell
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
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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
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The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
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The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
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The gut during HIV infection
Gut Lumen
Gut epithelium
Blood vessel
CD4Tcell
GALTLamina Propria
CD4Tcell HIV
HIV
Adenovirus
Bacteria
Bacteriophage
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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
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HIV gut microbiome studies
Gootenberg DB et al, Current Opinion in Infectious Diseases, 2017
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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
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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
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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
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CD4 T cell count correlates with sCD14
Monaco CL et al, Cell Host and Microbe, 2016
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Adenoviridae and Anelloviridae are expanded in AIDS
Monaco CL et al, Cell Host and Microbe, 2016
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Bacterial richness and diversity differ by CD4 T cell count
Richness Diversity
Monaco CL et al, Cell Host and Microbe, 2016
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Discriminant Bacterial Families
Monaco CL et al, Cell Host and Microbe, 2016
HIV Negative versus CD4 T cell count < 200
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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
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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
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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
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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
![Page 50: THE MICROBIOME IN · Alterations in the enteric microbiome are associated with disease •Metabolic disorders such as obesity and diabetes mellitus –Fecal transplant from obese](https://reader034.fdocuments.net/reader034/viewer/2022050217/5f632278fee278709d635e4a/html5/thumbnails/50.jpg)
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
![Page 51: THE MICROBIOME IN · Alterations in the enteric microbiome are associated with disease •Metabolic disorders such as obesity and diabetes mellitus –Fecal transplant from obese](https://reader034.fdocuments.net/reader034/viewer/2022050217/5f632278fee278709d635e4a/html5/thumbnails/51.jpg)
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
![Page 52: THE MICROBIOME IN · Alterations in the enteric microbiome are associated with disease •Metabolic disorders such as obesity and diabetes mellitus –Fecal transplant from obese](https://reader034.fdocuments.net/reader034/viewer/2022050217/5f632278fee278709d635e4a/html5/thumbnails/52.jpg)
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
![Page 53: THE MICROBIOME IN · Alterations in the enteric microbiome are associated with disease •Metabolic disorders such as obesity and diabetes mellitus –Fecal transplant from obese](https://reader034.fdocuments.net/reader034/viewer/2022050217/5f632278fee278709d635e4a/html5/thumbnails/53.jpg)
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
![Page 54: THE MICROBIOME IN · Alterations in the enteric microbiome are associated with disease •Metabolic disorders such as obesity and diabetes mellitus –Fecal transplant from obese](https://reader034.fdocuments.net/reader034/viewer/2022050217/5f632278fee278709d635e4a/html5/thumbnails/54.jpg)
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);
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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).
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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).
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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
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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
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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).
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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
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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
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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
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QUESTIONS?
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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
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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