Dromedary camels in northern Mali have high seropositivity to MERS-CoV

Falzarano, Darryl; Kamissoko, Badian; de Wit, Emmie; Maïga, Ousmane; Cronin, Jacqueline; Samaké, Kassim; Traoré, Abdalah; Milne-Price, Shauna; Munster, Vincent J.; Sogoba, Nafomon; Niang, Mamadou; Safronetz, David; Feldmann, Heinz

Serum from 562 dromedary camels from 16 different locations in Kidal region (north-eastern Mali) and from 9 dromedary camels on a single farm in Nara (central Mali) were collected as part of a survey for peste des petits ruminants virus between November 2009 and February 2010 according to local regulations. Samples were stored at − 25°C at Laboratoire Central Vétérinaire (LCV) in Bamako, Mali. For the MERS spike protein (S1) ELISA, serum obtained from LCV was diluted 1:1 in 0.2% Triton X − 100 (Sigma), heat inactivated for 15 min. at 65°C and subsequently stored at −20°C. NUNC MaxiSorp plates were coated with recombinant S1 antigen (1 μg/ml) (Sino Biological) in PBS overnight at 4°C. Unbound antigen was removed and plates were blocked with blocking solution (PBS + 5% non-fat skim milk, 0.05% Tween20). Serum was diluted in blocking solution and added to the plates for 1 h at 37°C. Plates were washed with PBS + 0.05% Tween20. A rabbit anti-llama IgG (H + L) HRP conjugated antibody (AgriSera) was used as the secondary antibody at 1:2000 in blocking buffer and incubated for 1 h at 37°C. Plates were washed, ABTS Peroxidase substrate (KPL) was added and plates were incubated in the dark for 30 min. Plates were subsequently read at 405 nm. It has been determined that this assay does not cross-react with antibodies to bovine coronavirus, OC43 or SARS-CoV.