Research Article: Optimization of a Novel Non-invasive Oral Sampling Technique for Zoonotic Pathogen Surveillance in Nonhuman Primates

Date Published: June 5, 2015

Publisher: Public Library of Science

Author(s): Tierra Smiley Evans, Peter A. Barry, Kirsten V. Gilardi, Tracey Goldstein, Jesse D. Deere, Joseph Fike, JoAnn Yee, Benard J Ssebide, Dibesh Karmacharya, Michael R. Cranfield, David Wolking, Brett Smith, Jonna A. K. Mazet, Christine K. Johnson, Anne Rimoin.

Abstract: Free-ranging nonhuman primates are frequent sources of zoonotic pathogens due to their physiologic similarity and in many tropical regions, close contact with humans. Many high-risk disease transmission interfaces have not been monitored for zoonotic pathogens due to difficulties inherent to invasive sampling of free-ranging wildlife. Non-invasive surveillance of nonhuman primates for pathogens with high potential for spillover into humans is therefore critical for understanding disease ecology of existing zoonotic pathogen burdens and identifying communities where zoonotic diseases are likely to emerge in the future. We developed a non-invasive oral sampling technique using ropes distributed to nonhuman primates to target viruses shed in the oral cavity, which through bite wounds and discarded food, could be transmitted to people. Optimization was performed by testing paired rope and oral swabs from laboratory colony rhesus macaques for rhesus cytomegalovirus (RhCMV) and simian foamy virus (SFV) and implementing the technique with free-ranging terrestrial and arboreal nonhuman primate species in Uganda and Nepal. Both ubiquitous DNA and RNA viruses, RhCMV and SFV, were detected in oral samples collected from ropes distributed to laboratory colony macaques and SFV was detected in free-ranging macaques and olive baboons. Our study describes a technique that can be used for disease surveillance in free-ranging nonhuman primates and, potentially, other wildlife species when invasive sampling techniques may not be feasible.

Partial Text: The World Health Organization designated the assessment of the burden of zoonoses as a strategic area for action in their global plan to combat neglected tropical diseases [1]. Both domestic and wild animals contribute to the burden of zoonotic disease [2]. Viruses originating in wild animals however account for over 70% of emerging zoonotic infectious diseases in humans including viruses that have caused pandemics such as HIV/AIDS, epidemics such as Ebola hemorrhagic fever and yellow fever, as well as smaller outbreaks such as Marburg hemorrhagic fever [3–8]. Free-ranging nonhuman primates (hereafter referred to as primates) are of particular concern as sources or carriers of zoonotic viruses because of their close phylogenetic and physiologic relationship and, in many geographic regions, frequent and close contact with humans [9, 10]. Human and primate contact is common in equatorial Africa with human encroachment into forest and savannah habitats [10] and in parts of Asia where urban-dwelling primates are flourishing [11, 12]. Surveillance of free-ranging primates at these high-risk interfaces is critical and will facilitate improved understanding of disease ecology, identify human communities at risk for pathogen transmission, and can enable the detection of zoonotic pathogens before their spillover into humans [13–15].

Distributed ropes were accepted and chewed by primates in both laboratory and free-ranging settings. Among the laboratory colony trials, macaques accepted and chewed on the ropes 45 of the 55 times when the ropes were 6-inches in length, in contrast to 9 of 30 times when the ropes were 3-feet in length. These latter 9 macaques had however, already been exposed to 6-inch ropes. Among the free-ranging behavioral trials, oral samples were successfully collected from primates using ropes for 18 of 20 macaques, 18 of 22 olive baboons, 16 of 20 red-tailed guenons, and 8 of 10 l’hoest’s monkeys. All species accepted the ropes with fruit jam applied as an attractant (Figs 2 and 3) except baboons; the rope had to be completely disguised inside a banana in order for them to chew on it (Fig 4). Ropes with retrieval strings attached were not as effective due to macaques, baboons and l’hoest’s monkeys being fearful and/or distracted by the strings.

Non-invasive oral samples were successfully collected from arboreal and terrestrial dwelling free-ranging primate species for detection of DNA and RNA viruses. To use this technique, primates targeted for sampling must be willing to chew on ropes. We found that the optimal technique for recovering oral samples from laboratory colony macaques was to use ropes 6-inches in length with a retrieval string attached. Laboratory colony macaques were not willing to chew on the ropes when they were longer than 6-inches. We speculate that 3-foot length ropes resemble snakes, as fear behaviors are common among various primate species [52]. Similarly, the optimal technique for free-ranging primates was to use ropes 6-inches in length but with no retrieval strings attached. Baboons and l’hoest’s monkeys were fearful of any form of an attached string and macaques were more likely to become aggressive towards the handler. When strings were removed, primates were less distracted by the strings and spent more time chewing on ropes. In addition, macaques from the Pashupati Temple Complex were observed placing the ropes into cheek pouches where prolonged contact with oro-pharyngeal mucosa could occur, potentially increasing the opportunity for viral sampling.



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