Antibodies against MERS Coronavirus in Dromedary Camels, United Arab Emirates, 2003 and 2013

Benjamin Meyer, Marcel A. Müller, Victor M. Corman, Chantal B.E.M. Reusken, Daniel Ritz, Gert-Jan Godeke, Erik Lattwein, Stephan Kallies, Artem Siemens, Janko van Beek, Jan F. Drexler, Doreen Muth, Berend-Jan Bosch, Ulrich Wernery, Marion P.G. Koopmans, Renate Wernery, and Christian Drosten

Author affiliations: University of Bonn Medical Centre, Bonn, Germany (B. Meyer, M.A. Müller, V.M. Corman, D. Ritz, S. Kallies, A. Siemens, J.F. Drexler, D. Muth, C. Drosten); National Institute for Public Health and the Environment, Bilthoven, the Netherlands. (C.B.E.M. Reusken, G.-J. Godeke, J. van Beek, M.P.G. Koopmans);Erasmus Medical Centre, Rotterdam, the Netherlands (C.B.E.M. Reusken, J.F. Drexler, M.P.G. Koopmans); EUROIMMUN AG, Lübeck, Germany (E. Lattwein); Utrecht University, Utrecht, the Netherlands (B.-J. Bosch); Central Veterinary Research Laboratory, Dubai, United Arab Emirates (U. Wernery, R. Wernery)

Abstract

Middle East respiratory syndrome coronavirus (MERS-CoV) has caused an ongoing outbreak of severe acute respiratory tract infection in humans in the Arabian Peninsula since 2012. Dromedary camels have been implicated as possible viral reservoirs. We used serologic assays to analyze 651 dromedary camel serum samples from the United Arab Emirates; 151 of 651 samples were obtained in 2003, well before onset of the current epidemic, and 500 serum samples were obtained in 2013. Recombinant spike protein–specific immunofluorescence and virus neutralization tests enabled clear discrimination between MERS-CoV and bovine CoV infections. Most (632/651, 97.1%) camels had antibodies against MERS-CoV. This result included all 151 serum samples obtained in 2003. Most (389/651, 59.8%) serum samples had MERS-CoV–neutralizing antibody titers >1,280. Dromedary camels from the United Arab Emirates were infected at high rates with MERS-CoV or a closely related, probably conspecific, virus long before the first human MERS cases.

Middle East respiratory syndrome coronavirus (MERS-CoV) is an emerging pathogen associated with severe respiratory symptoms and renal failure in infected patients (1,2). Globally, 156 laboratory-confirmed cases of infection with MERS-CoV, including 65 deaths, were reported as of early November 2013. All human cases were linked to the Arabian Peninsula (Saudi Arabia, Jordan, Oman, Qatar, Kuwait, and the United Arab Emirates). Imported cases were detected in countries in Europe and Africa (United Kingdom, Germany, Italy, France, and Tunisia) (3).

Transmission patterns, including the putative zoonotic source of the virus, remain unclear. Hypotheses include frequent zoonotic infections with limited subsequent human-to-human transmission chains and existence of a self-sustained epidemic in humans (4). A recent study found evidence to support the existence of epidemiologically unlinked cases in a large outbreak in the al-Hasa region, Saudi Arabia (5). It was speculated that zoonotic introductions of MERS-CoV from an unknown reservoir might occur at high rates, in addition to obvious human-to-human transmission.

Coronaviruses (CoV) are positive-sense RNA viruses. Viruses in the genera Alphacoronavirus andBetacoronavirus are associated with mammals and show a particularly high level of diversification in bats. Viruses in the genera Gammacoronavirus and Deltacoronavirus are mostly avian-associated viruses (6,7). MERS-CoV belongs to Betacoronavirus phylogenetic lineage C that, in addition to MERS-CoV, contains 2 distinct bat-associated CoV species (HKU4 and HKU5) (1,8).

Insectivorous bats of the family Vespertilionidae were recently shown to carry viruses that are probably conspecific with MERS-CoV (9). However, the limited rate of contact between humans and insectivorous bats makes a continuous and frequent acquisition of MERS-CoV from bats an unlikely scenario. In a manner similar to observations regarding severe acute respiratory syndrome CoV (SARS-CoV), an intermediate reservoir host might exist from which human infections are acquired. Dromedary camels from different regions in Africa and the Arabian Peninsula have been shown to have antibodies against MERS-CoV (10,11). Animals from the Arabian Peninsula had high neutralizing serum activities overall and reciprocal antibody titers ≤320–1,280, which support recent infection with MERS-CoV or a highly related virus. Thus, dromedary camels might serve as intermediate hosts. However, detailed serologic studies in countries with actual incidence of MERS-CoV infections in humans have not been conducted.

Serologic analysis of CoVs is challenging because of cross-reactivity between CoVs infecting the same host and the broad distribution of CoVs in diverse mammalian species (6,7,12–14). Antibodies directed against some of the major antigens of different CoVs are known to cross-react in standard serologic assays (15,16). Potential cross-reactivity is a diagnostic challenge because camelids are known to be infected with bovine CoV (BCoV), a distinct betacoronavirus of phylogenetic lineage A unrelated to the MERS-CoV (17,18). As an additional challenge, camel immunoglobulins lack a light chain peptide, which affects specific physical properties, such as altered size and stability, compared with immunoglobulins of other mammals (19,20). The influence of this feature on serologic assays has not been thoroughly investigated. Thus, serologic assays should be applied with caution, and different assay formats should be tested concurrently.

We reported a 2-staged approach for MERS-CoV serologic analysis in humans (15,16). Expanding upon these studies, we used in the present study a recombinant MERS-CoV spike protein immunofluroescence assay (rIFA) augmented by a validated protein microarray (10,21), followed by MERS-CoV–specific neutralization assay, to screen 651 dromedary camel serum samples from the United Arabian Emirates. Cross-reactivity against clade A betacoronaviruses was assessed by using a immunofluorescence assay (IFA) and a BCoV-specific neutralization assay. Serum samples obtained in 2003 and 2013 were compared to obtain information for the time in which MERS-related CoV has been circulating in camels.