Lack of Zika Virus and Other Recognized Flaviviruses among the Mosquito Vectors during and Post the Hajj Mass Gathering

Makkah city, Kingdom of Saudi Arabia (KSA), contains many of the world’s mosquito vectors of parasitic and arboviral disease and is the site of the Hajj mass gathering. As such there is a risk of exportation and globalization of vector-borne viruses, including the re-emerging Zika virus (ZIKV). There was international concern regarding the introduction of ZIKV to KSA and potential international spread of the virus following the 2016 Hajj which took place few days after the Rio summer Olympics at the height of the ZIKV pandemic. We aimed to detect flaviviruses, including ZIKV, circulating among mosquito hosts in the city of Makkah during and post the 2016 Hajj pilgrimage. Mosquitos (adults and larvae) were sampled from 15 sites in Makkah city during and post the 2016 Hajj and identified to species by morphological keys. Mosquitos were pooled according to date of collection, location, and species. A Pan-Flaviviruses RT-PCR assay that enables identification of 51 flaviviruses species and three tentative species was used to detect flavivirus RNA directly from mosquito homogenates. Between the 10 September and 6 October 2016, 9412 female mosquitos were collected. Of these, 81.3% were Aedes aegypti, 18.6% were Culex species, and 0.1% were Anopheles species. Of the total 493 mosquito pools generated, 242 (49%) were positive by the Pan-Flaviviruses primer set. Sequence analysis revealed that none of the mosquitos carried a pathogenic flavivirus, including ZIKV, but were infected with a novel insect-specific flavivirus. We found no pathogenic flaviviruses circulating in Makkah city during and post the 2016 Hajj and no evidence of introduction of ZIKV through the pilgrimage. Enhanced vector-borne diseases surveillance, prevention, and control are crucial in KSA especially during international mass gatherings such as the annual Hajj to prevent outbreaks and the spread of viruses with epidemic and pandemic potentials.