Mitigation of endemic GI-tract pathogen-mediated inflammation through development of multimodal treatment regimen and its impact on SIV acquisition in rhesus macaques

Abstract

Here, we assessed the efficacy of a short-course multimodal therapy (enrofloxacin, azithromycin, fenbendazole, and paromomycin) to eliminate common macaque endemic pathogens (EPs) and evaluated its impact on gastrointestinal (GI) microbiota, mucosal integrity, and local and systemic inflammation in sixteen clinically healthy macaques. Treatment combined with expanded practices resulted in successful maintenance of rhesus macaques (RM) free of common EPs, with no evidence of overt microbiota diversity loss or dysbiosis and instead resulted in a more defined luminal microbiota across study subjects. Creation of a GI pathogen free (GPF) status resulted in improved colonic mucosal barrier function (histologically, reduced colonic MPO+, and reduced pan-bacterial 16s rRNA in the MLN), reduced local and systemic innate and adaptive inflammation with reduction of colonic Mx1 and pSTAT1, decreased intermediate (CD14+CD16+) and non-classical monocytes (CD14-CD16+), reduced populations of peripheral dendritic cells, Ki-67+ and CD38+ CD4+ T cells, Ki-67+IgG+, and Ki-67+IgD+ B cells indicating lower levels of background inflammation in the distal descending colon, draining mesenteric lymph nodes, and systemically in peripheral blood, spleen, and axillary lymph nodes. A more controlled rate of viral acquisition resulted when untreated and treated macaques were challenged by low dose intrarectal SIVmac239x, with an ~100 fold increase in dose required to infect 50% (AID₅₀) of the animals receiving treatment compared to untreated controls. Reduction in and increased consistency of number of transmitted founder variants resulting from challenge seen in the proof of concept study directly correlated with post-treatment GPF animal’s improved barrier function and reduction of key target cell populations (Ki-67+ CD4+T cells) at the site of viral acquisition in the follow up study. These data demonstrate that a therapeutic and operational strategy can successfully eliminate varying background levels of EPs and their associated aberrant immunomodulatory effects within a captive macaque cohort, leading to a more consistent, better defined and reproducible research model. Simian Immunodeficiency virus in macaques remains the most translational model for HIV. Macaques are also key models for other infectious diseases and colitis, where background colon health and inflammation could confound experimental results. To address these issues SPF breeding colonies were established to exclude specific viruses detrimental to mucosal and systemic health. To further improve macaque models, we established a gastrointestinal pathogen free (GPF) colony by administration of a multimodal therapeutic regimen for common endemic pathogens (EPs) and established expanded operational practices for continued exclusion. Through extensive longitudinal microbiota, parasitology, microbiology, and tissue sampling we demonstrated that our treatment and exclusion practices successfully eliminated common EPs, improved mucosal barriers, and reduced mucosal and systemic inflammation resulting in less inter-animal variation without overt evidence of microbiota disruption. Finally, compared to treatment-naïve controls, GPF animals challenged with SIV intrarectally demonstrated a more controlled and consistent rate of SIV acquisition, further indicating underlying EPs even at subclinical levels may cause deleterious variations between study subjects. Collectively the GPF macaque represents a model that eliminates a significant source of morbidity, reduces inter-animal variability, and thus has the potential to improve both animal welfare and research outcomes.