Coronaviridae
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CHAPTER 3 Coronaviridae James A. Robb and Clifford W. Bond* Department of Pathology University of California, San Diego La Jol/a, California 92093 1. INTRODUCTION 1.1. Summary Corona viruses are widespread in nature, but relatively little atten tion has been given to them. They are pathogenic, enveloped RNA viruses whose large, single-stranded, nonsegmented genome is of a posi tive polarity. They cause a broad spectrum of disease in their natural hosts, including man, primarily by a cytocidal virus-cell interaction. The structure of the virion, the molecular mechanisms of their multi plication strategy, and their pathogenesis are just beginning to receive the attention they deserve. Our goals in this chapter are three: (1) to define the criteria for membership and the present members of the coronavirus family; (2) to define the structure of the virion and the multiplication strategy for this family; and (3) to describe the spectrum and pathogenesis of disease produced by this family of important pathogenic viruses in their natural hosts. The preprints and unpublished data supplied by many of our colleagues are appreciated. Our emphasis will be, whenever possible, on the molecular description of the multiplication strategy of these viruses * Present addresses: J. A. R., Department of Pathology, The Green Hospital of Scripps Clinic, La Jolla, California 92037. C. W. B., Department of Microbiology, Mon tana State University, Bozeman, Montana 59717. 193 H. Fraenkel-Conrat et al. (eds.), Comprehensive Virology © Plenum Press, New York 1979 194 Chapter 3 and on the molecular mechanisms involved in the pathogenesis of the disease produced by these viruses. There are at present large gaps in the description of the molecular virology of this family. In an attempt to bridge some of these gaps and make the review as up to date as possi ble, prudent speculation will be used and identified as such. 1.2 Definition and Members of the Coronavirus Family 1.2.1. Criteria for Membership and Tentative Members Until recently, enveloped, RNA-containing viruses that budded solely from the endoplasmic reticular membranes, that had a spherical shape with a 60-220 nm diameter, and that had a "corona" of widely spaced, bulbous peplomers 12-24 nm in length were defined as coronaviruses (McIntosh, 1974). Acceptable biochemical criteria were not available. Such biochemical criteria are now available using avian infectious bronchitis virus (IBV) as the prototype virus for this family (Schochetman et af., 1977; Lomniczi, 1977; Lomniczi and Kennedy, 1977). The virion genome of enveloped viruses of the above morphology must be a large (6-8 X 106 daltons) single piece of single-stranded infectious RNA of messenger (positive) polarity. In addition to IBV, the virions of mouse hepatitis virus (MHV), human coronavirus (HeV), and porcine transmissible gastroenteritis virus (TG EV) probably contain a genome of positive polarity as described in Section 2.2.2. In addition, several other viruses have satisfied the electron microscopic criteria for membership in this family and are listed in Table 1. Addi tional biochemical criteria will soon be forthcoming because the investi gation of the molecular virology of the coronavirus family is rapidly gaining momentum. 1.2.2. Serological Relatedness of Members The intraspecies and inter species serological relatedness of the various viruses are complex and not yet fully understood (Table 1). The production of useful vaccines against coronavirus diseases, many of which are economically important in agriculture and probably in the human economy as well, will require a more detailed understanding of the serological relatedness of these viruses than is currently available. The major problem in acquiring these data, as well as in understanding Coronaviridae 195 TABLE 1 Tentative Members of the Coronavirus Family and Their Serological Relatedness Natural Virus" host SerotypesC Serologically related to' Avian infectious bronchitis (IBV) Chicken Many' ? TGEV' Canine coronavirus (CCV) Dog ? One' TGEV',' Feline coronavirus (FCV) Cat ? One' TGEV, but not HEV' (infectious peritonitis) Huinan coronavirus (HCV) Man Several"···· HEV, MHV··7 Human enteric corona virus (HECV) Man ? One TGEV' Murine hepatitis virus (MHV) Mouse Many' RCV, SDA V, HCV, ?HEV··7 Neonatral calf diarrhea corona- Bovine Several· virus· (NCDCV) Porcine transmissible gastro- Pig One"'" CCV., FCV', ?HEV', ?IBV' enteritis virus (TGEV) Porcine hemagglutinating Pig ? One' encephalitis virus (HEV) Rat coronavirus (RCV) Rat ? Ones MHV' Rat sialodacryoadenitis Rat ? Ones MHV' virus (SDA V) Runde tick coronavirus (RTCV) ? Tick ? One'· Not to IBV or MHV'· ? Sea bird Turkey bluecomb disease Turkey ? One' virus (TCDV) " These names and abbreviations conform to those suggested by Tyrrell et al. (personal communi cation) in a revision to the previous recommendations (Tyrrell et al., 1975). • Mucosal disease virus (border disease, Plant et al., 1976) of sheep may belong in this group (Snowdon et al., 1975). C References: 1, Cowen and Hitchner (1975); 2, McIntosh (1974); 3, Kapikian (1975); 4, Binn et al. (1974); 5, Reynolds et al. (1977); 6, Monto (1974); 7, Kaye et al. (1977); 8, Hafez et al. (1976); 9, Kemeny (1976); 10, Traavik et al. (1977). the molecular virology, is the relative difficulty in isolating and growing these viruses in cell culture. This problem is discussed in Section 3. 2. VIRIONS 2.1. Morphology Coronavirus virions are pleomorphic spherical particles of 80-160 nm diameter with characteristic large, widely spaced, 12- to 24-nm-Iong spikes or peplomers that form a corona around the particle and provide the name for this virus family. Figures 1 and 2 show the novel mor- 196 Chapter 3 Fig. 1. A group of IBV virions treated with formaldehyde before staining with phos photungstic acid. Almost all reveal the internal component. The majority display the tongue or flask orientation, but others show a circular structure or even two concentric rings. Figure 2 shows that all of these patterns are compatible with an internal membra nous sac continuous with the outer membrane. Magnification x 135,000 (reduced 20% for reproduction). From Bingham and Almeida (1977); used by permission. phology of the infectious bronchitis virion (Bingham and Almeida, 1977). Whether this is the only internal morphology available to a coronavirus virion remains to be determined. The topology of the ribonucleocapsid within a virion is not yet known. The nucleocapsid is composed of a 9-nm-wide ribonucleoprotein that forms a helical struc ture (Kennedy and Johnson-Lussenburg, 1975/1976; Pocock and Garwes, 1977) and probably corresponds to the flask like structure within the virion (Fig. 2). Using transmission electron microscopy, we have observed in mouse hepatitis virus (JHMV and A59V) infected 17CL-16 cells and BALB/c brains that the apparent nucleocapsids in cytoplasmic factories and in early stages of budding have horseshoe or flask like configurations (Robb et af., 1979a,b). Purification of nucleocapsids from virions and infected cells will be necessary to clarify the internal structure of the coronavirus virion. The mass of a human coronavirus (OC43) is 390 ± 5 X 106 daltons as determined by analytical ultracentrifugation (Hierholzer et al., 1972). Similar experiments with another human coronavirus (229E) did Coronaviridae 197 not give reliable data because the virus particles disintegrated too rapidly. The mass of other coronaviruses has not been determined. 2.2. Composition 2.2.1. Overall Chemical Composition The virions of coronaviruses contain RNA, protein, carbohydrate, and lipid. The buoyant density of the virion in sucrose or potassium tar trate is in the range of 1.17-1.19 g/cm3 [IBV: MacNaughton and Madge, 1977a; NCDCV: La Porte, personal communication; HCV (299E): Hierholzer, 1976; FCV: Horzinek et al., 1977; MHV (A59V, lHMV): Lai and Stohlman, 1978; Stohlman, personal communication; Leibowitz, Bond, and Robb, manuscript in preparation]. 2.2.2. RNA The genome of coronaviruses has been amply demonstrated to be single-stranded RNA. Lomniczi (1977) and Schochetman et al. (1977) have shown that the genome of infectious bronchitis virus (lBV) is Fig. 2. Three different transilluminated orientations of a glass model are compared to four individual virus particles printed at x330,000 (reduced 20% for reproduction). Although both refraction and reflection will occur with the glass model, the overall cor relation is good. From Almeida (1977); used by permission. 198 Chapter 3 infectious, and Lomniczi (personal communication) has shown that the genome of transmissible gastroenteritis virus (TGEV) is infectious. The molecular size of the virion RNA of IBV has been variously reported to be 0.5-3.0 million daltons (4-40 S) by Tannock (1973), 9.0 million daltons (50 S) by Watkins et al. (1975), 8 million daltons (64 S) by Lomniczi and Kennedy (1977), 8 million daltons (58 S) by MacNaughton and Madge (1977b), and 5.5-5.7 million daltons (48 S) by Schochetman et al., (1977). These variable data reflect the fact that the genome of corona viruses is the largest viral RNA genome known and, as a result, no standards exist to size the RNA by traditional techniques. Lomniczi and Kennedy (1977) used the elegant method of RNase T1 oligonu cleotide fingerprinting to confirm the molecular size of 8.1 ± 0.2 million daltons which had been determined on methyl mercury gels and isokinetic sucrose gradients. Tannock and Hierholzer (1977) have shown that the RNA of a human coronavirus (HCV-OC43)