Discussion
Previous studies have demonstrated that, besides dromedaries, several other members of the Camelidae family are susceptible to MERS-CoV infection, such as Bactrian camels, alpacas, and llamas , although human contact infections have only been reported from dromedary camels . However, the existing data on the susceptibility of non-camelids are conflicting. Our results are in line with a series of publications demonstrating that non-camelids do not play a major role in the transmission cycle . For instance, in a seroepidemiological study in Saudi Arabia (2010-2013), no antibodies were detected in sheep, goats, cattle, or chickens, but a high prevalence was found in camels . Similarly, after an experimental challenge of sheep, goats, and horses by intranasal MERS-CoV inoculation, none of the animals effectively shed the virus, and neutralizing antibodies were only found in goats . Further regarding the family Equidae , including horses, donkeys, and mules, although viral replication has been observed in primary horse kidney cells, no antibodies have been found in any of 1,053 equid sera collected in the UAE and Spain . A study on livestock cell lines from the Arabian Peninsula showed that MERS-CoV can replicate in goat, alpaca, and dromedary cells, in contrast to sheep, cattle, rodent, and insectivore cells . In a study experimentally challenging animals from France and Spain, MERS-CoV nucleic acid in the respiratory tract as well as seroconversion were detected only in llamas and pigs, but not in sheep and horses . A serosurvey in a MERS-CoV-affected area in Jordan identified MERS-CoV neutralising antibodies in the sera of all camels but none in goats and cattle, while 4.8 % of sheep sera reacted with MERS-CoV antigen . In a monitoring study of several mammalian species in direct contact with infected dromedaries in Egypt, Tunisia, and Senegal (2015-2017), antibodies were identified in an astonishing 55.6 % of sheep from Senegal, 1.8 % of sheep from Tunisia, and 0.9 % of goats from Egypt. MERS-CoV RNA was detected in nasal swabs from 1.2 % sheep and 4.1 % goats from Egypt and Senegal, as well as 1.9 % cattle and 7.1 % donkeys from Egypt . In conclusion, it seems that several non-camelid species may become infected with MERS-CoV during viral spillover events when they are in direct contact with virus-shedding dromedaries. Maybe this susceptibility is due to the close relatedness of the Betacoronaviruses (specifically their receptors) infecting camels and especially bovids, but also humans, porcines, and equids . However, whether these animals are implicated in the MERS-CoV transmission cycle remains unknown, but seems to be highly unlikely.
The fact that none of the sheep, goats, and cattle in our survey tested positive for antibodies indicates that none of them had previously experienced a MERS-CoV infection, although with an age range of three months to six years, all animals could have encountered the virus in the past. Moreover, as the average time of stay at the market for goats is usually 7-10 days, 10-14 days for sheep, and between 14 and 30 days for cattle, there is also a not negligible risk of infection at the market. However, the nasal swab of one sheep (1.8 %) contained MERS-CoV nucleic acid (Supplementary Table 1), suggesting either a current infection, or a spillover event from nearby dromedaries, which appears to be more likely since no antibodies were detected and the sheep was kept in a pen in close proximity to the camel pens (approximately 25 m away; Fig. 1). Of note, we additionally sampled 90 dromedaries at the same market and at the same time as the non-camelids, of which 35.6 % were positive for MERS-CoV nucleic acid, and another 24.4 % tested borderline positive (Ct values 38.6-41.2). Moreover, 91.1 % of the 90 camel sera were MERS-CoV antibody-positive by ELISA, and 96.7 % by IFA (data can be provided upon request), further supporting the high prevalence of MERS-CoV infections in dromedaries. This hypothesis is also supported by our latest study, in which we screened 76 dromedaries during spring and autumn 2019 at the same market, and found MERS-CoV nucleic acid in 57.9 % of their nasal swabs .
The positive sheep from this study was a two-year-old female, presumably locally raised Nuaimi breed. However, due to the fact that sex was not equally distributed across the animals we sampled (115 females, 75.7 %, compared to 37 males, 24.3 %), no conclusions can be drawn regarding the susceptibility of the different sexes. The same is true for the different breeds, with only one positive animal, we cannot infer any breed-specific genetic predispositions to MERS-CoV infection.
In conclusion, our study shows that non-camelids such as sheep, goats, and cattle do not play a major role in MERS-CoV epidemiology. The one sheep that tested RNA-positive in our study most likely reflects a viral spillover event from infected dromedaries in nearby pens. Furthermore, serology revealed that the sheep did not develop an antibody response, suggesting that the infection was very recent, or the sheep only carried viral particles on its nasal mucosa without actually being infected.