Simian Immunodeficiency Virus and Storage Buffer: Field-friendly preservation methods for RNA viral detection in primate feces

Abstract

Wild non-human primates carry many types of RNA viruses, including simian immunodeficiency virus (SIV), simian foamy virus, simian T-cell leukemia virus, and hepatitis C virus. These viruses can also infect humans via zoonotic transmission through handling and consumption of primate bushmeat. Characterizing viral prevalence and shedding in natural hosts is critical to understand infection and transmission risks within and between primate species. Here, we sought to identify a robust "field-friendly" method (i.e., without freezing or refrigeration) for preserving viral RNA, specifically SIV, in primate fecal samples. Fecal samples were collected from a mantled guereza colobus (Colobus guereza) housed at the Columbus Zoo and Aquarium. Samples were homogenized and inoculated with three concentrations (low, medium, high) of inactivated SIV virus and preserved in four different storage buffers (DNA/RNA Shield, RNAlater, 95% Ethanol, and Viral Transport Medium). SIV viral RNA was then extracted from samples at four time points (1 week, 4 weeks, 8 weeks, and 12 weeks) to determine the efficacy of each buffer for preserving SIV RNA. Quantitative RT-PCR was used for detection and quantification of viral RNA. At all concentrations, DNA/RNA Shield yielded the highest average SIV virion concentrations. We then successfully validated this approach using fecal samples from known SIV-positive and SIV-negative sooty mangabeys (Cercocebus atys) housed at Emory National Primate Research Center. Our results indicate that DNA/RNA shield is an optimal "field-friendly" buffer for preserving SIV RNA in fecal samples over time, and may also be effective for preserving other RNA viruses in feces. ImportanceHuman immunodeficiency virus (HIV) was introduced into human populations through zoonotic transmission of SIV from African primates, leading to a global epidemic and ongoing worldwide public health issue. SIV occurs naturally in over 40 primate species in sub-Saharan Africa and these viruses have crossed species barriers on multiple occasions, leading to the spread of HIV-1 and HIV-2. Quantifying RNA viruses in wild primate populations can be challenging as invasive sampling is often not feasible, and many field stations lack ready access to a freezer for storing biological samples. This study compares SIV RNA preservation and recovery across multiple storage buffers to identify a robust field-friendly option for RNA viral detection in noninvasively collected feces. Our results will inform future fieldwork and facilitate improved approaches to characterizing prevalence, shedding, and transmission of RNA viruses like SIV in natural hosts including wild-living nonhuman primates.