key: cord-0330550-yy80q9i3 authors: Sereme, Youssouf; Zarza, Sandra Madariaga; Medkour, Hacène; Amona, Inestin; Fenollar, Florence; Akiana, Jean; Mezouar, Soraya; Orain, Nicolas; Vitte, Joana; Davoust, Bernard; Raoult, Didier; Mediannikov, Oleg title: Stool serology: development of a non-invasive immunological method for the detection of Enterovirus-specific antibodies in Congo gorilla feces date: 2020-11-28 journal: bioRxiv DOI: 10.1101/2020.11.28.402230 sha: 20db6f12ef5a27c9ec840f4d5d12cf98b4ddf481 doc_id: 330550 cord_uid: yy80q9i3 The incidence of poliovirus has been significantly reduced by 99.9% globally meanwhile vaccine-associated paralytic poliomyelitis emerges. Recently, a new recombinant virus (Enterovirus C/Poliovirus) was identified in humans working as eco-guards and gorillas in Democratic Republic of Congo, including one gorilla with polio-like sequeler. A strain of this recombinant virus (Ibou002) was also isolated from feces. In order to assess the potential role of poliovirus infection, we have developed and optimized a protocol, based on the lyophilization and solubilization of stool in a small volume, for the detection of specific antibodies. A high level of total immunoglobulins was first detected in the concentrated stool extracts. Then, specific antibodies were detected in 4/16 gorilla samples and 2/3 human samples by western blot using both poliovaccine antigen and Ibou002 antigen and by ELISA using polio vaccine antigen. The humoral responses were greater with the Ibou002 antigen both in number and intensity of bands. Thus, we suggest that this recombinant virus could cause polio-like disease in the endangered western lowland gorilla. The development of a non-invasive method for the detection of microorganism-specific immunoglobulins from fecal samples opens up new perspectives for the exploration of humoral responses of pathogens in animals and knowledge in zoonotic infectious diseases. Poliovirus is a virus belonging to the Picornaviridae family and the Enterovirus genus 42 (Enterovirus C), which causes poliomyelitis in humans 1 . The poliovirus genome is a 7.5 43 kilobase single-stranded RNA surrounded by a capsid, which is composed by four proteins 44 (VP1, VP2, VP3, and VP4) 2 . There are three poliovirus types (1, 2, and 3); the immunity to one 45 type would not protect against the two others 3 . Mediterranean Region with endemic circulation of wild poliovirus type 1 in 2 countries 54 (https://www.who.int/wer/2020/wer9541/en/) 6,11-13 . In parallel, another challenge arose with the 55 and the emergence of infectious diseases in humans 16 . It has been shown under experimental 66 conditions that hominoids and monkeys can be infected with poliovirus 17, 18 . A 2005 study in 67 Madagascar demonstrated the circulation of vaccine-derived recombinant poliovirus strains, 68 which are recombinant Enterovirus-C viruses, and their ability to give "polio-like" in humans 19 . The polio antigen derived from the human pathogenic poliovirus types, specifically type 1 134 (Brunhilde), type 2 (Lansing), and type 3 (Leon) was bound in the wells of the plate. The 135 diluted samples or standards were then deposited. After 1-hour incubation at room temperature, 136 the plate was rinsed with wash buffer. The anti-human IgG peroxidase conjugate was then 137 added and incubated for 30 min, followed by a wash step. Afterwards, the substrate solution Supernatant was filtered by serial filtration steps to eliminate cells and cellular debris, going 156 throw 0.8 µm, 0.45 µm, and 0.2 µm filters. Then, virus particles were precipitated in 10% PEG-then fractionated with TS buffer (7 M Urea, 2 M Thiourea, 4% CHAPS) to release the antigen. 165 The released antigen was then concentrated with the Amicon 3 kDa filter (Merck KGaA, 166 Darmstadt, Germany) before being analyzed by SDS gel electrophoresis, and the western blot. 167 The concentration of the antigen was measured. Then, 30 µg of purified were loaded onto 12% (Figure 4) . It is therefore the latter that was selected to continue analyzes on the stool. 218 The protein profile was performed for both antigens (Imovax vaccine and strain Ibou002). In this study, we developed a non-invasive method for screening animal immune 234 responses using fresh stool samples. We also provided a proof of concept on the feasibility of 235 exploring immunity from feces. 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