Lactobacillus-depleted vaginal microbiota in pregnant women living with HIV-1 infection are associated with increased local inflammation and preterm birth

Background: Pregnant women living with HIV-1 infection (PWLWH) have an elevated risk of preterm birth (PTB) of unknown aetiology, which remains after successful suppression of HIV. Women at high risk for HIV have a common bacterial profile which has been associated with poor birth outcomes. We set out to explore factors associated with gestational age at delivery of PWLWH in a UK population. Methods: Prospective study of PWLWH (n = 53) in whom the vaginal microbiota and cervicovaginal cytokine milieu were assessed using metataxonomics and multiplexed immunoassays, respectively. Cross-sectional characterisation of vaginal microbiota in PWLWH were compared with 22 HIV uninfected pregnant women (HUPW) at a similar second trimester timepoint. Within PWLWH the relationships between bacterial composition, inflammatory response, and gestational age at delivery were explored. Findings: There was a high rate of PTB among PWLWH (12%). In the second trimester the vaginal microbiota was more diverse in PWLWH than in HUPW (Inverse Simpson Index, p = 0.0004 and Species Observed, p = 0.009). PWLWH had a lower prevalence of L. crispatus dominant vaginal microbiota group (VMB I, 15 vs 54%) than HUPW and higher prevalence of L. iners dominant (VMB III, 36 vs 9% and VMB IIIB, 15 vs 5%) and mixed anaerobes (VMB IV, 21 vs 0%). Across the second and third trimesters in PWLWH, VMB III/IIIB and IV were associated with PTB and with increased local inflammation [cervicovaginal fluid (CVF) cytokine concentrations in upper quartile]. High bacterial diversity and anaerobic bacterial abundance were also associated with CVF pro-inflammatory cytokines, most notably IL-1β. Interpretation: There is an association between local inflammation, vaginal dysbiosis and PTB in PWLWH. Understanding the potential of antiretroviral therapies to influence this cascade will be important to improve birth outcomes in this population

Comparative analysis of vaginal microbiota sampling using menstrual cups and high vaginal swabs in pregnant women living with HIV-1 infection

Background: Menstrual cups (MCs) are increasingly used to collect cervicovaginal secretions to characterise vaginal mucosal immunology, in conjunction with high vaginal swabs (HVS) for metataxonomics, particularly in HIV transmission studies. We hypothesised that both methods of collecting bacterial biomass are equivalent for 16S rRNA gene sequencing. Material and Methods: Cervicovaginal fluid (CVF) samples from 16 pregnant women with HIV-1 (PWWH) were included to represent the major vaginal bacterial community state types (CST I-V). Women underwent sampling during the second trimester by liquid amies HVS followed by a MC (Soft disc™) and samples were stored at -80°C. Bacterial cell pellets obtained from swab elution and MC (500 µL, 1 in 10 dilution) were resuspended in 120 µL PBS for DNA extraction. Bacterial 16S rRNA gene sequencing was performed using V1-V2 primers and were analysed using MOTHUR. Paired total DNA, bacterial load, amplicon read counts, diversity matrices and bacterial taxa were compared by sampling method using MicrobiomeAnalyst, SPSS and R. Results: The total DNA eluted from one aliquot of diluted CVF from an MC was similar to that of a HVS (993ng and 609ng, p=0.18); the mean bacterial loads were also comparable for both methods (MC: 8.0 log10 16S rRNA gene copies versus HVS: 7.9 log10 16S rRNA gene copies, p=0.27). The mean number of sequence reads generated from MC samples was lower than from HVS (MC: 12730; HVS:14830, p=0.05). The α-diversity metrices were similar for both techniques; MC Species Observed: 41 (range 12-96) versus HVS: 47 (range 16-96), p=0.15; MC Inverse Simpson Index: 1.98 (range 1.0-4.0) versus HVS: 0.48 (range 1.0-4.4), p=0.22). The three most abundant species observed were: Lactobacillus iners, Lactobacillus crispatus and Gardnerella vaginalis. Hierarchical clustering of relative abundance data showed that samples obtained using different techniques in an individual clustered in the same CST group. Conclusion: These data demonstrate that despite sampling slightly different areas of the lower genital tract, there was no difference in bacterial load or composition between methods. Both are suitable for characterisation of vaginal microbiota in PWWH. The MC offers advantages, including a higher volume of sample available for DNA extraction and complimentary assays

Vaginal microbiota, genital inflammation and extracellular matrix remodelling collagenase: MMP-9 in pregnant women with HIV, a potential preterm birth mechanism warranting further exploration

Background: Pregnant women living with HIV infection (PWLWH) have elevated rates of preterm birth (PTB) in which HIV and cART are implicated. PWLWH also have a high prevalence of adverse vaginal microbiota, which associate with genital tract inflammation. The mechanism underlying PTB in PWLWH is unknown. We present the first data in PWLWH on genital-tract matrix-metalloproteinase-9(MMP-9), an important collagenase implicated in labour onset, and tissue inhibitor of metalloproteinases-1(TIMP-1) and explore correlations with local inflammation and vaginal bacteria

Inflammatory profiles across the spectrum of disease reveal a distinct role for GM-CSF in severe COVID-19

While it is now widely accepted that host inflammatory responses contribute to lung injury, the pathways that drive severity and distinguish coronavirus disease 2019 (COVID-19) from other viral lung diseases remain poorly characterized. We analyzed plasma samples from 471 hospitalized patients recruited through the prospective multicenter ISARIC4C study and 39 outpatients with mild disease, enabling extensive characterization of responses across a full spectrum of COVID-19 severity. Progressive elevation of levels of numerous inflammatory cytokines and chemokines (including IL-6, CXCL10, and GM-CSF) were associated with severity and accompanied by elevated markers of endothelial injury and thrombosis. Principal component and network analyses demonstrated central roles for IL-6 and GM-CSF in COVID-19 pathogenesis. Comparing these profiles to archived samples from patients with fatal influenza, IL-6 was equally elevated in both conditions whereas GM-CSF was prominent only in COVID-19. These findings further identify the key inflammatory, thrombotic, and vascular factors that characterize and distinguish severe and fatal COVID-19

The immunological basis of preterm birth in HIV-1 infected pregnant women

Hypotheses HIV infection and combination antiretroviral therapy (cART) used to prevent mother-to-child-transmission of HIV in pregnancy increase the risk of preterm birth (PTB). We hypothesise that the use of cART, particularly protease inhibitor (PI) regimes, induce a pro-inflammatory environment at the fetal maternal unit, which triggers labour to occur at an earlier point in gestation than would occur without HIV-1 infection or cART. We also hypothesise that local cervicovaginal fluid (CVF) will be more informative regarding immune function than plasma. Specific aims -To understand the inflammatory mechanisms underlying PTB in HIV1infected and uninfected women -To elucidate if PI-based cART regimes are associated with greater concentrations of inflammatory cytokines compared to non-PI-based cART regimes in pregnancy -To characterise inflammatory and immune proteins in the CVF of HIV-1 infected pregnant women and compare by ART exposure and prematurity with a view to identifying potential PTB biomarkers. Results PTB in HIV-1 infected women was associated with greater circulating activated T cells and abundance of vaginal Gardnerella and Prevotella species. HIV-1 infected pregnant women have a high prevalence of L. iners and diverse anaerobic bacteria dominant vaginal community state types. Pre-conception cART was associated with greater abundance of Gardnerella vaginalis and a reduction in circulating activated T cells but not to the same level as observed in uninfected pregnant women. Length of time on cART was associated with an increase in plasma IL-12, the macrophage activating cytokine, indicating a pro-inflammatory 11 shift in cytokine environment. HIV-1 infected pregnant women had much higher inflammatory cytokines in their CVF. In HIV-1 infected pregnant women vaginal bacterial diversity was positively correlated with CVF pro-inflammatory cytokine IL-1E, known to be associated with PTB. Directed exploration of the CVF proteome in HIV-1 infected pregnant women revealed high abundance of immune proteins associated with macrophage and neutrophil activation, and Extracellular Matrix (ECM) modifiers such as matrix metalloproteinases (MMPs). These proteins were especially abundant in women receiving PI-based cART. Significance These data indicate that ART may modulate the local bacterial species; perhaps through selective pressure and that the vaginal bacterial communities observed in HIV-1 infected women are pro-inflammatory and associated with PTB. Local CVF inflammation and immune activation is exaggerated in these HIV-1 infected women and not fully reversed with cART. The presence of bacterial antigen in the context of enhanced local innate and adaptive inflammatory immune response is likely responsible for up-regulating downstream NFκB pathways such as induction of MMPs, known to be association with induction of labour. Contribution to academic area A greater understanding of the mechanisms behind this phenomenon enable us to greater define the toxicity of certain antiretroviral classes or specific drugs in pregnancy and assess newer drugs using the same techniques. Optimising the risk stratification in these women, perhaps through application of improved PTB biomarkers or regular vaginal bacterial screening, may enable improved targeted risk reduction strategies e.g. progesterone, antibiotics or probiotics.Open Acces

Transcriptional reprogramming from innate immune functions to a pro-thrombotic signature by monocytes in COVID-19

Although myeloid cell dysfunction has been observed in COVID-19, the underlying mechanisms remain incompletely understood. Here, the authors demonstrate that monocytes from patients with mild to moderate COVID-19 show a blunted innate immune response and a pro-thrombotic signature following secondary SARS-CoV-2 challenge

Elevated antiviral, myeloid and endothelial inflammatory markers in severe COVID-19

Introductory paragraph The mechanisms that underpin COVID-19 disease severity, and determine the outcome of infection, are only beginning to be unraveled. The host inflammatory response contributes to lung injury, but circulating mediators levels fall below those in classical ‘cytokine storms’. We analyzed serial plasma samples from 619 patients hospitalized with COVID-19 recruited through the prospective multicenter ISARIC clinical characterization protocol U.K. study and 39 milder community cases not requiring hospitalization. Elevated levels of numerous mediators including angiopoietin-2, CXCL10, and GM-CSF were seen at recruitment in patients who later died. Markers of endothelial injury (angiopoietin-2 and von-Willebrand factor A2) were detected early in some patients, while inflammatory cytokines and markers of lung injury persisted for several weeks in fatal COVID-19 despite decreasing antiviral cytokine levels. Overall, markers of myeloid or endothelial cell activation were associated with severe, progressive, and fatal disease indicating a central role for innate immune activation and vascular inflammation in COVID-19