Microbiome Composition and Function Drives Wound-Healing Impairment in the Female Genital Tract

The mechanism(s) by which bacterial communities impact susceptibility to infectious diseases, such as HIV, and maintain female genital tract (FGT) health are poorly understood. Evaluation of FGT bacteria has predominantly been limited to studies of species abundance, but not bacterial function. We therefore sought to examine the relationship of bacterial community composition and function with mucosal epithelial barrier health in the context of bacterial vaginosis (BV) using metaproteomic, metagenomic, and in vitro approaches. We found highly diverse bacterial communities dominated by Gardnerella vaginalis associated with host epithelial barrier disruption and enhanced immune activation, and low diversity communities dominated by Lactobacillus species that associated with lower Nugent scores, reduced pH, and expression of host mucosal proteins important for maintaining epithelial integrity. Importantly, proteomic signatures of disrupted epithelial integrity associated with G. vaginalis-dominated communities in the absence of clinical BV diagnosis. Because traditional clinical assessments did not capture this, it likely represents a larger underrepresented phenomenon in populations with high prevalence of G. vaginalis. We finally demonstrated that soluble products derived from G. vaginalis inhibited wound healing, while those derived from L. iners did not, providing insight into functional mechanisms by which FGT bacterial communities affect epithelial barrier integrity

Vaginal bacteria modify HIV tenofovir microbicide efficacy in African women.

CAPRISA, 2017.Abstract available in pdf

Increased levels of inflammatory cytokines in the female reproductive tract are associated with altered expression of proteases, mucosal barrier proteins, and an influx of HIV-susceptible target cells.

CAPRISA, 2016.Abstract available in pdf

Non-cationic proteins are associated with HIV neutralizing activity in genital secretions of female sex workers.

CAPRISA, 2015.Abstract available in pdf

A Comparative Proteomic Analysis of the Soluble Immune Factor Environment of Rectal and Oral Mucosa

<div><p>Objective</p><p>Sexual transmission of HIV occurs across a mucosal surface, which contains many soluble immune factors important for HIV immunity. Although the composition of mucosal fluids in the vaginal and oral compartments has been studied extensively, the knowledge of the expression of these factors in the rectal mucosa has been understudied and is very limited. This has particular relevance given that the highest rates of HIV acquisition occur via the rectal tract. To further our understanding of rectal mucosa, this study uses a proteomics approach to characterize immune factor components of rectal fluid, using saliva as a comparison, and evaluates its antiviral activity against HIV.</p><p>Methods</p><p>Paired salivary fluid (n = 10) and rectal lavage fluid (n = 10) samples were collected from healthy, HIV seronegative individuals. Samples were analyzed by label-free tandem mass spectrometry to comprehensively identify and quantify mucosal immune protein abundance differences between saliva and rectal fluids. The HIV inhibitory capacity of these fluids was further assessed using a TZM-bl reporter cell line.</p><p>Results</p><p>Of the 315 proteins identified in rectal lavage fluid, 72 had known immune functions, many of which have described anti-HIV activity, including cathelicidin, serpins, cystatins and antileukoproteinase. The majority of immune factors were similarly expressed between fluids, with only 21 differentially abundant (p<0.05, multiple comparison corrected). Notably, rectal mucosa had a high abundance of mucosal immunoglobulins and antiproteases relative to saliva, Rectal lavage limited HIV infection by 40–50% <i>in vitro</i> (p<0.05), which is lower than the potent anti-HIV effect of oral mucosal fluid (70–80% inhibition, p<0.005).</p><p>Conclusions</p><p>This study reveals that rectal mucosa contains many innate immune factors important for host immunity to HIV and can limit viral replication <i>in vitro</i>. This indicates an important role for this fluid as the first line of defense against HIV.</p></div

Average abundance of antiprotease proteins found in in rectal mucosa as determine by mass spectrometry, and relative expression of these proteins in rectal mucosa compared to saliva.

<p>Average abundance of antiprotease proteins found in in rectal mucosa as determine by mass spectrometry, and relative expression of these proteins in rectal mucosa compared to saliva.</p

Average abundance of proteins significantly underabundant in rectal mucosa relative to salivary fluid as determined by mass spectrometry, and relative expression of these proteins in rectal mucosa compared to saliva.

<p>Average abundance of proteins significantly underabundant in rectal mucosa relative to salivary fluid as determined by mass spectrometry, and relative expression of these proteins in rectal mucosa compared to saliva.</p

Selected list of immune proteins having described roles in HIV defense found in mucosal fluids.

<p>Selected list of immune proteins having described roles in HIV defense found in mucosal fluids.</p

Rectal lavage shows mild inhibitory activity against R5-tropic HIV <i>in vitro</i>.

<p>Inhibition assays of HIV BaL replication within the CCR5+/CXCR4+ TZM-bl reporter cells in the presence of varying concentrations of rectal lavage and salivary fluid protein were performed. Rectal lavage exhibited a significant, mild inhibitory effect on HIV infection in TZM-bl cells (∼40% inhibition) beginning at 2 µg/ml of protein relative to a negative control (p = 0.05, triplicate assays) (A). Parallel assays demonstrated that salivary fluid had a stronger anti-HIV capacity (∼70–80% inhibition) at as low as 0.3 µg/ml relative to the negative control (p<0.005, triplicate assays) (B). Mucosal fluids were determined to have a negligent effect on cell death based on a luciferase assay that indirectly measured the number of viable cells in each culture via their ATP production (data not shown).</p