Differential cell line susceptibility to the emerging novel human betacoronavirus 2c EMC/2012: implications for disease pathogenesis and clinical manifestation

Abstract

The emerging novel human betacoronavirus 2c EMC/2012 (HCoV-EMC) was recently isolated from patients with severe pneumonia and renal failure and was associated with an unexplained high crude fatality rate of 56%. We performed a cell line susceptibility study with 28 cell lines. HCoV-EMC was found to infect the human respiratory tract (polarized airway epithelium cell line Calu-3, embryonic fibroblast cell line HFL, and lung adenocarcinoma cell line A549), kidney (embryonic kidney cell line HEK), intestinal tract (colorectal adenocarcinoma cell line Caco-2), liver cells (hepatocellular carcinoma cell line Huh-7), and histiocytes (malignant histiocytoma cell line His-1), as evident by detection of high or increasing viral load in culture supernatants, detection of viral nucleoprotein expression by immunostaining, and/or detection of cytopathic effects. Although an infected human neuronal cell line (NT2) and infected monocyte and T lymphocyte cell lines (THP-1, U937, and H9) had increased viral loads, their relatively lower viral production corroborated with absent nucleoprotein expression and cytopathic effects. This range of human tissue tropism is broader than that for all other HCoVs, including SARS coronavirus, HCoV-OC43, HCoV-HKU1, HCoV-229E, and HCoV-NL63, which may explain the high mortality associated with this disease. A recent cell line susceptibility study showed that HCoV-EMC can infect primate, porcine, and bat cells and therefore may jump interspecies barriers. We found that HCoV-EMC can also infect civet lung fibroblast and rabbit kidney cell lines. These findings have important implications for the diagnosis, pathogenesis, and transmission of HCoV-EMC.

Figure 1.

Expression of human betacoronavirus 2c EMC/2012 (HCoV-EMC) nucleoprotein as intense apple green cytoplasmic fluorescence in different cell lines on day 3 after infection stained by monospecific polyclonal serum obtained from a guinea pig infected with His6-tagged HCoV-EMC nucleoprotein (original magnification ×200). A, Infected lower airway (Calu-3) cells. B, Uninfected Calu-3 control. C, Infected intestinal (Caco-2) cells. D, Uninfected Caco-2 control. E, Infected liver (Huh-7) cells. F, Uninfected Huh-7 control. G, Infected histiocytes (His-1). H, Uninfected His-1 control. I, Infected African green monkey kidney (Vero) cells. J, Uninfected Vero control.

Figure 2.

Cytopathic effects of cell rounding, detachment, and syncytia formation (black arrows) in the Calu-3 and Caco-2 cell lines and aggregates of infected cells with marked shrinkage (white arrows) in the Huh-7 cell line (hematoxylin-eosin stain) on day 3 after infection by human betacoronavirus 2c EMC/2012, observed under inverted microscopy (original magnification ×40–100). A, Infected lower airway (Calu-3) cells. B, Uninfected Calu-3 control. C, Infected intestinal (Caco-2) cells. D, Uninfected Caco-2 control. E, Infected liver (Huh-7) cells. F, Uninfected Huh-7 control.