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WHAT 2nd Microbiome workshop WHEN 17 & 18 November 2016 WHERE Masur Auditorium at NIH Campus, Bethesda MD Modulating the Vaginal Microbiome to Prevent HIV Infection Laurel Lagenaur For more information: www.virology-education.com Conflict of Interest I work for Osel Inc., a microbiome company developing live biotherapeutic products to prevent diseases in women The Vaginal Microbiome and HIV Infection What’s normal /healthy/ optimal and what’s not? • Ravel Community Groups • Lactobacillus dominant vs. dysbiosis Why are Lactobacilli important for HIV prevention? • Dysbiosis = Inflammation = Increased risk of HIV acquisition • Efficacy of Pre-Exposure Prophylaxis decreased Modulation of the vaginal microbiome • LACTIN-V, live biotherapeutic-Lactobacillus crispatus • Ongoing clinical trials How we can use Lactobacilli and go a step further • Genetically modified Lactobacillus to prevent HIV acquisition HIV is Transmitted Across Mucosal Surfaces • HIV infection in women occurs in the mucosa of the vagina and cervix vagina cervix • Infection of underlying target cells All mucosal surfaces are continuously exposed to a Herrera and Shattock, community of microorganisms Curr Top Microbiol Immunol 2013 Vaginal Microbiome: Ravel Community Groups Vaginal microbiomes clustered into 5 groups: Group V L. jensenii Group II L. gasseri 4 were dominated by Group I L. crispatus Lactobacillus, Group III L. iners whereas the 5th had lower proportions of lactic acid bacteria and Group 4 Diversity higher proportions of Prevotella, strictly anaerobic Sneathia, Megasphaera, organisms Atopobium Shannon Diversity: Ravel et al. PNAS 2011 low species high species richness/evenness richness/evenness Vaginal Microbiome Lactobacilli-dominated Healthy Vagina Dysbiosis • Low diversity, low vaginal pH • Antagonize pathogens Low Diversity High Diversity • Non-inflammatory Low Nugent High Nugent pH Bacterial vaginosis >5.5 • Decrease H O -producing Lactobacilli 2 2 pH • Increased bacterial diversity 3.6-4.5 • High pH, odor, discharge • Increased risk of STI and HIV Bacterial communities in O + Adapted from Srinivasan 2012 Vaginal Microbiota vs. Host Inflammatory Responses Inflammatory Host vs. Microbiota Vaginal • CT 1 CT Community types Community CT 4 CT Alpha diversity Alpha IFN-γ (pg/ml) IL-1α (pg/ml) 1β, TNF 1β, cytokines of proinflammatory levels Increased IL- IL-\) IL- IL-\) - \) IL-\) \) IL-\) IFN α, Anahtar - γ , IL , et al et - IL-10 (pg/ml) IL-1β (pg/ml) IL and 10 . Immunity . 2015 Immunity - IL- 8 IL- IL-\) IL-\) \) IL-\) \) IL-\) - IL-8 (pg/ml) TNF-α (pg/ml) IL - 1α , IL IL- IL- - IL-\) IL-\) \) IL-\) \) IL-\) Genital Inflammation is Linked to HIV infection Abundance% Abundance % Abundance % Abundance0 >50 % Abundance 0 0 0 0 >50>50 >50 >50 Lactobacillus crispatus Lactobacillus crispatus Lactobacillus crispatus Lactobacillus iners Lactobacillus iners South African womenLactobacillus iners in the Prevotella bivia Prevotella bivia Prevotella bivia LactobacilluSneathias crispatus sp 99 Sneathia sp 99 Sneathia sp 99 Shuttleworthia sp 9 [BVAB-1] Shuttleworthia sp 9 [BVAB-1] Shuttleworthia sp 9 [BVAB-1] LactobacilluAerococcuss iners christensenii Aerococcus CAPRISAchristensenii study withAerococcus christensenii P. bivia were Veillonella montpellierensis Veillonella montpellierensis Veillonella montpellierensis PrevotellaPrevotella bivia amnii Prevotella amnii Prevotella amnii Atopobium vaginae Atopobium vaginae Atopobium vaginae SneathiaMegasphaera sp 99 sp 9 Megasphaera sp 9 19 times moreMegasphaera likely sp 9 to have Lactobacillus gasseri Lactobacillus gasseri Lactobacillus gasseri ShuttleworthiaMycoplasma-like sp 9 [BVAB-1] sp 9 Mycoplasma-like sp 9 Mycoplasma-like sp 9 AerococcusMycoplasma christensenii hominis Mycoplasma hominis genital inflammationMycoplasma hominis and 13 Atopobium sp 6 Atopobium sp 6 Atopobium sp 6 Shuttleworthia sp 58 [BVAB-1] Shuttleworthia sp 58 [BVAB-1] Shuttleworthia sp 58 [BVAB-1] VeilloStreptococcusnella montpellierensis agalactiae Streptococcus agalactiae Streptococcus agalactiae Lactobacillus sp 91 Lactobacillus sptimes 91 more likelyLactobacillu tos sp 91acquire HIV PrevotellaStaphylococcus amnii epidermidis Staphylococcus epidermidis Staphylococcus epidermidis AtopobiumSneathia vaginae sanguinegens Sneathia sanguinegens Sneathia sanguinegens Prevotella timonensis Prevotella timonensis Prevotella timonensis MegasphaeraCoriobacteriaceae sp 9 sp 9 Coriobacteriaceae sp 9 Coriobacteriaceae sp 9 Prevotella sp 101 Prevotella sp 101 Prevotella sp 101 LactobacilluUreaplasmas gasseri sp 1 Ureaplasma sp 1 Ureaplasma sp 1 Granulicatella elegans Granulicatella elegans Granulicatella elegans Mycoplasma-likeWillia sp 9 msia muralis Williamsia muralis Williamsia muralis MycoplasmaHIV hominis status HIV status HIV status Cytokine group Cytokine group Cytokine group 150 150 150 Trichomonas vaginalis Trichomonas vaginalis Trichomonas vaginalis d d d Atopobium sp 6 s s s e e e e e e v v v i i i r r 100 100 r 100 Chlamydia trachomatis Chlamydia trachomatis Chlamydia trachomatis c c c e e e e e e s s s p p p b b b Neisseria gonorrhoeae Neisseria gonorrhoeae Neisseria gonorrhoeae s s 50 50 s 50 Shuttleworthia sp 58 [BVAB-1] O O O 0 0 0 HSV-2 HSV-2 HSV-2 StreptococcusHSV agalactiae -1 HSV-1 HSV-1 Lactobacillus sp 91 Staphylococcus epidermidis Sneathia sanguinegens Prevotella timonensis Coriobacteriaceae sp 9 Prevotella sp 101 Ureaplasma sp 1 Granulicatella elegans Williamsia muralis HIV status Cytokine group 150 Trichomonas vaginalis d s e e v i r 100 Chlamydia trachomatis c e e s p b Neisseria gonorrhoeae s 50 O 0 HSV-2 HSV-1 Vaginal Microbiota May Influence Tenofovir Efficacy • Tenofovir disoproxil - Nucleotide analog reverse-transcriptase inhibitor used in Pre-Exposure Prophylaxis • Delivered as a 1% intravaginal gel (before and after sex) Tenofovir gel effective against HIV with Lactobacillus- dominance Lactobacilli-dominated Non-Lactobacillus dominant 0.40 0.40 A. LaA.ct obaLacctobaillusc idllousm indoamntinant 0.40 0.40 B. NoB.n No-Lanc-Latobactobacilluscil ldusom dinomanintant 0.35 0.35 0.35 0.35 n 61% efficacy n 18% efficacy n EfficaEfcyf,i c a6c1y%, 61% n EfficEfacfyic,a 1c8y,% 18% o o 0.30o 0.30 0.30o i 0.30 i i i t t t t c 95% CI, 11 to 84% c 95% CI, -77 to 63% c 95% CI, 11 to 84% c 95% CI, -77 to 63% e e e e f f f f n n 0.25n 0.25n i 0.25 i 0.25 i i P=0.644 V V P=0.644 V I V I I I H H H H f 0.20 f 0.20 f 0.20 f 0.20 o o o o y y t y t y i t i t l i l i i l i 0.15 0.15l i 0.15 i 0.15 b b b b a a a a b b b b o o o r 0.10 P=0.013 o r 0.10 r 0.10 P=0.013 r 0.10 P P P P 0.05 0.05 0.05 0.05 HR = 0.39 (95% CI: 0.20; 0.83) HR = 0.82 (95% CI: 0.40; 1.65) 0.00 HR = 0.39 (95% CI: 0.20; 0.83) HR = 0.82 (95% CI: 0.40; 1.65) 0.00 0.00 0.00 0.0 0.5 1.0 1.5 2.0 2.5 0.0 0.5 1.0 1.5 2.0 2.5 0.0 0.0 0.5 0.5 1.0 1.0 1.5 1.5 2.0 2.0 2.5 2.5 Years in Study Years in Study Years in Study Years in Study Tenofovir Placebo Tenofovir Placebo Tenofovir Placebo Tenofovir Placebo Lactobacillus dominant women at risk (Cumulative number of infections) Non-Lactobacillus dominant women at risk (Cumulative number of infections) Lactobacillus dominant women at risk (Cumulative number of infections) Non-Lactobacillus dominant women at risk (Cumulative number of infections) Tenofovir 205 (0) 204 (1) 183 (3) 129 (7) 46 (9) 0 (9) Tenofovir 140 (0) 137 (2) 123 (6) 87 (10) 32 (13) 0 (14) Tenofovir 205 (0) 204 (1) 183 (3) 129 (7) 46 (9) 0 (9) Tenofovir 140 (0) 137 (2) 123 (6) 87 (10) 32 (13) 0 (14) Placebo 202 (0) 196 (4) 173 (12) 123 (19) 51 (22) 0 (22) Placebo 141 (0) 137 (4) 116 (12) 84 (17) 28 (17) 0 (17) Placebo 202 (0) 196 (4) 173 (12) 123 (19) 51 (22) 0 (22) Placebo 141 (0) 137 (4) 116 (12) 84 (17) 28 (17) 0 (17) Tenofovir was rapidly depleted by Gardnerella but not Lactobacillus Tenofovir (supernatant) Tenofovir (cell) 1.5 Abiotic e g 0.003 L. iners L. iners n a ) h G. vaginalis G. vaginalis L C 1.0 m / d l g 0.002 o m F ( r i r i v 0.5 v o o f f o o 0.001 n n e e T T 0.0 0 10 20 30 0.000 Time (hours) 0 10 20 30 Time (hours) 4 hours: G. vag vs. L. iners: P=0.002 4 hours: G. vag vs Abiotic: P=0.005 G. vag vs. L. iners: P<0.001 24 hours: 24 hours: G. vag vs. L. iners: P<0.001 G. vag vs. L. iners: P<0.001 G. vag vs Abiotic: P<0.001 What Can We Do to Decrease HIV Infection in Women? • Treat their Bacterial vaginosis- Metronidazole Lactobacillus dominant Dysbiosis Abx Post-Abx Restoration ↓ H O + Lactobacillus 2 2 Live Biotherapeutic Product (LBP) Growth of diverse commensal and pathogenic bacteria LACTIN-V: Lactobacillus crispatus CTV-05 • Give women exogenous Lactobacillus Osel Product LACTIN-V (Lactobacillus crispatus) for Treatment of Bacterial Vaginosis LACTIN-V Clinical Studies Phase 1 • Safe and well-tolerated at all dose levels* * Hemmerling et al., S Trans Dis 2009 Phase 2a • Following Metronidazole treatment • Colonization up to 78% of women treated for BV** ** Hemmerling et al., S Trans Dis 2010 • Colonization correlated with reduced BV recurrence Phase 2b (sponsor NIAID/DMID ) • Prevention of recurrent BV • 1st patient enrolled Q2 2016 • Locations: University of California, San Diego, San Francisco General Hospital, Cook County Health and Hospitals System, Chicago, Washington University School of Medicine in St.
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    bioRxiv preprint doi: https://doi.org/10.1101/2020.03.10.985580; this version posted March 11, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 1 Differential vaginal Lactobacillus species metabolism of glucose, L and D-lactate by 13C-nuclear magnetic 2 resonance spectroscopy 3 Emmanuel Amabebea, [email protected] 4 Dilly O. Anumbaa, [email protected] 5 Steven Reynoldsb*, [email protected] 6 aDepartment of Oncology and Metabolism, University of Sheffield, Level 4, Jessop Wing, Tree Root Walk, 7 Sheffield S10 2SF, UK. 8 bDepartment of Infection, Immunity and Cardiovascular Disease, University of Sheffield, Royal Hallamshire 9 hospital, Glossop Road, Sheffield S10 2JF, UK 10 *Corresponding author 11 Dr Steven Reynolds, BSc (Hons), PhD 12 Academic Unit of Radiology, Department of Infection, Immunity and Cardiovascular Disease, University of 13 Sheffield, Royal Hallamshire hospital, Glossop Road, Sheffield S10 2JF, UK. +44 114 215 9596, 14 [email protected] 15 16 Acknowledgements: 17 We are grateful to the University of Sheffield, UK, for providing support and the 9.4T MRS spectrometer with 18 which this study was performed. 19 20 21 22 23 24 25 26 27 1 bioRxiv preprint doi: https://doi.org/10.1101/2020.03.10.985580; this version posted March 11, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 28 Abstract 29 Introduction 30 Cervicovaginal dysbiosis can lead to infection-associated spontaneous preterm birth.
  • Antibiotic Resistance in Poultry Gastrointestinal Microbiota and Targeted Mitigation

    Antibiotic Resistance in Poultry Gastrointestinal Microbiota and Targeted Mitigation

    Antibiotic Resistance in Poultry Gastrointestinal Microbiota and Targeted Mitigation Dissertation Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Yang Zhou, B.S. Graduate Program in Food Science and Technology The Ohio State University 2016 Dissertation Committee: Hua Wang, Advisor Monica Giusti Michael Lilburn Zhongtang Yu Copyright by Yang Zhou 2016 Abstract The rapid emergence and spread of antibiotic resistance (AR) is a major public health concern. The poultry industry worldwide represents the largest segment in food animal production. The prevalence and abundance of antibiotic resistant (ART) bacteria in poultry and poultry products have been a recognized food safety challenge. The large amount of ART bacteria-rich feces from industrial poultry production and its release further contaminate water and soil, impacting the environmental AR gene pool. Therefore, revealing contributing factors to AR in poultry production and developing targeted control strategies has critical impacts on AR mitigation in the ecosystem. This study examined 1) the natural occurrence of AR in gastrointestinal (GI) tract of chicken without antibiotic exposure; 2) the impact of antibiotic administration route on AR ecology in chicken GI microbiota, and 3) the efficacy of commensal Lactobacillus crispatus CG-2 inocula in AR mitigation in poultry rearing. The results of this study contributed to an improved understanding of AR ecology in food-producing animals. ii The first chapter is a literature review covering the origin, propagation, and dissemination of AR, as well as AR mitigation strategies. AR status in food-producing animals, particularly poultry, and recent achievements in AR mitigation were also reviewed to lay the foundation for research presented in this dissertation.
  • Vaginal Microbiota and the Potential of Lactobacillus Derivatives in Maintaining Vaginal Health Wallace Jeng Yang Chee , Shu Yih Chew and Leslie Thian Lung Than*

    Vaginal Microbiota and the Potential of Lactobacillus Derivatives in Maintaining Vaginal Health Wallace Jeng Yang Chee , Shu Yih Chew and Leslie Thian Lung Than*

    Chee et al. Microb Cell Fact (2020) 19:203 https://doi.org/10.1186/s12934-020-01464-4 Microbial Cell Factories REVIEW Open Access Vaginal microbiota and the potential of Lactobacillus derivatives in maintaining vaginal health Wallace Jeng Yang Chee , Shu Yih Chew and Leslie Thian Lung Than* Abstract Human vagina is colonised by a diverse array of microorganisms that make up the normal microbiota and mycobiota. Lactobacillus is the most frequently isolated microorganism from the healthy human vagina, this includes Lactobacil- lus crispatus, Lactobacillus gasseri, Lactobacillus iners, and Lactobacillus jensenii. These vaginal lactobacilli have been touted to prevent invasion of pathogens by keeping their population in check. However, the disruption of vaginal ecosystem contributes to the overgrowth of pathogens which causes complicated vaginal infections such as bacterial vaginosis (BV), sexually transmitted infections (STIs), and vulvovaginal candidiasis (VVC). Predisposing factors such as menses, pregnancy, sexual practice, uncontrolled usage of antibiotics, and vaginal douching can alter the microbial community. Therefore, the composition of vaginal microbiota serves an important role in determining vagina health. Owing to their Generally Recognised as Safe (GRAS) status, lactobacilli have been widely utilised as one of the alterna- tives besides conventional antimicrobial treatment against vaginal pathogens for the prevention of chronic vaginitis and the restoration of vaginal ecosystem. In addition, the efectiveness of Lactobacillus as prophylaxis has also been well-founded in long-term administration. This review aimed to highlight the benefcial efects of lactobacilli deriva- tives (i.e. surface-active molecules) with anti-bioflm, antioxidant, pathogen-inhibition, and immunomodulation activi- ties in developing remedies for vaginal infections.