Isolation and Characterization of Dromedary Camel Coronavirus UAE-HKU23 from Dromedaries of the Middle East: Minimal Serological Cross-Reactivity between MERS Coronavirus and Dromedary Camel Coronavirus UAE-HKU23.

Patrick Woo,Kwok-Yung Yuen,Rachel Fan,Ulrich Wernery,Candy Lau,Sunitha Joseph,Renate Wernery,Susanna Lau,Chi-Ching Tsang,Emily Wong,Alan Tsang,Cyril Yip

doi:10.3390/ijms17050691

Abstract

Recently, we reported the discovery of a dromedary camel coronavirus UAE-HKU23 (DcCoV UAE-HKU23) from dromedaries in the Middle East. In this study, DcCoV UAE-HKU23 was successfully isolated in two of the 14 dromedary fecal samples using HRT-18G cells, with cytopathic effects observed five days after inoculation. Northern blot analysis revealed at least seven distinct RNA species, corresponding to predicted subgenomic mRNAs and confirming the core sequence of transcription regulatory sequence motifs as 5′-UCUAAAC-3′ as we predicted previously. Antibodies against DcCoV UAE-HKU23 were detected in 58 (98.3%) and 59 (100%) of the 59 dromedary sera by immunofluorescence and neutralization antibody tests, respectively. There was significant correlation between the antibody titers determined by immunofluorescence and neutralization assays (Pearson coefficient = 0.525, p < 0.0001). Immunization of mice using recombinant N proteins of DcCoV UAE-HKU23 and Middle East respiratory syndrome coronavirus (MERS-CoV), respectively, and heat-inactivated DcCoV UAE-HKU23 showed minimal cross-antigenicity between DcCoV UAE-HKU23 and MERS-CoV by Western blot and neutralization antibody assays. Codon usage and genetic distance analysis of RdRp, S and N genes showed that the 14 strains of DcCoV UAE-HKU23 formed a distinct cluster, separated from those of other closely related members of Betacoronavirus 1, including alpaca CoV, confirming that DcCoV UAE-HKU23 is a novel member of Betacoronavirus 1.

Highlights

Coronaviruses (CoVs) are found in a wide variety of animals [1] in which they can cause respiratory, enteric, hepatic, and neurological diseases
We report the isolation of DcCoV UAE-HKU23 from the fecal sample of a dromedary and its characterization
Of the seven cell lines inoculated with dromedary fecal samples positive for DcCoV UAE-HKU23, viral replication was detected by RT-PCR in the supernatants of HRT-18G in two of the 14 dromedary fecal samples at day 7, with viral loads of 2.6 ˆ 1010 and 9.7 ˆ 106 copies/mL in HRT-18G cells in the presence of trypsin

Summary

Introduction

Coronaviruses (CoVs) are found in a wide variety of animals [1] in which they can cause respiratory, enteric, hepatic, and neurological diseases. As a result of their unique mechanism of viral replication, CoVs have a high frequency of recombination [2,6]. In 2005, we described the discovery, complete genome sequence, and molecular epidemiology of another novel HCoV, human CoV HKU1 (HCoV-HKU1), in the genus Betacoronavirus [20,21,22]. As for animal CoVs, we and others have described the discovery of SARS-CoV-like viruses in Chinese horseshoe bats in Hong Kong and other horseshoe bats in other provinces of China [23,24]. The discovery of SARS-CoV-like viruses in Chinese horseshoe bats in Yunnan has further highlighted the importance for hunting the animal origin of human infections [25]. From our studies it was shown that bats are the gene source for Alphacoronavirus and Betacoronavirus and birds are the gene source for Gammacoronavirus and Deltacoronavirus to fuel CoV evolution and dissemination [33]

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