V. ter Meulen

The Biology and Pathogenesis of Coronaviruses

The coronavirases were first recognized and morphologically defined as a group by Tyrrell and co-workers (1968, 1975, 1978). Biochemical studies have recently provided additional information which allows better characterization of these agents. Presently, coronaviruses are defined as being particles which are pleomorphic to rounded with a diameter of 60–220 nm, surrounded by a fringe or layer of typical club-shaped spikes. The virion is composed of about four to six proteins and possesses a lipid bilayer. The genome consists of a single-stranded polyadenylated RNA which is infectious and of positive polarity. During maturation these viruses are released by internal budding into vesicles derived from the endoplasmatic reticulum. These viruses are widespread in nature and are associated with a great variety of diseases with an acute, subacute, or subclinical disease process.

The biology of coronaviruses.

Introduction. The Coronaviridae is a monogeneric family comprising 11 viruses which infect vertebrates. Members of the group are responsible for diseases of clinical and economic importance, in particular respiratory and gastrointestinal disorders (Table 1). The group was originally recognized on the basis of a characteristic virion morphology (Tyrrell et al., 1968), but can now be defined by biological and molecular criteria. Various aspects of coronavirus biology have been dealt with in recent reviews (Robb & Bond, 1979; Siddell et al., 1982; Wege et al., 1982). Structure. Morphology. Coronavirions are pleomorphic, although generally spherical, 60 to 220 nm in diameter and bear widely spaced, club-shaped surface projections about 20 nm in length. Complete virions have a density in sucrose of about 1.18 g/ml. In thin sections the virion envelope may be visualized as inner and outer shells separated by a translucent space. In negatively stained preparations of avian infectious bronchitis virus (IBV) an inner tongue-shaped membrane is visible (Bingham & Almeida, 1977; Fig. 1).

Sequence divergence of measles virus haemagglutinin during natural evolution and adaptation to cell culture

Phylogenetic analysis of the sequence of the H gene of 75 measles virus (MV) strains (32 published and 43 new sequences) was carried out. The lineage groups described from comparison of the nucleotide sequences encoding the C-terminal regions of the N protein of MV were the same as those derived from the H gene sequences in almost all cases. The databases document a number of distinct genotype switches that have occurred in Madrid (Spain). Well-documented is the complete replacement of lineage group C2, the common European genotype at that time, with that of group D3 around the autumn of 1993. No further isolations of group C2 took place in Madrid after this time. The rate of mutation of the H gene sequences of MV genotype D3 circulating in Madrid from 1993 to 1996 was very low (5 x 10(-4) per annum for a given nucleotide position). This is an order of magnitude lower than the rates of mutation observed in the HN genes of human influenza A viruses. The ratio of expressed over silent mutations indicated that the divergence was not driven by immune selection in this gene. Variations in amino acid 117 of the H protein (F or L) may be related to the ability of some strains to haemagglutinate only in the presence of salt. Adaptation of MV to different primate cell types was associated with very small numbers of mutations in the H gene. The changes could not be predicted when virus previously grown in human B cell lines was adapted to monkey Vero cells. In contrast, rodent brain-adapted viruses displayed a lot of amino acid sequence variation from normal MV strains. There was no convincing evidence for recombination between MV genotypes.

Identification of several different lineages of measles virus

The sequences of a region of the nucleocapsid protein gene, between nucleotides 1231 and 1686, encoding the C-terminal 151 amino acid residues of the nucleocapsid protein have been determined for 16 strains of measles virus. Analysis of this region showed that it is highly divergent (up to 7.2% divergence in the nucleotide sequence and 10.6% divergence in the amino acid sequence between most distant strains) and that several lineages of measles virus can be found to co-circulate at a given time. Some of the lineages show geographical restriction. The results for measles virus are similar to those reported for other human paramyxoviruses such as mumps virus, parainfluenza type 3 virus and the avian Newcastle disease virus.

The Structure and Replication of Coronaviruses

Coronaviruses were recognized as a group in 1968 primarily on the basis of their characte ristic morphology as seen in the electron microscope (Tyrrell et al. 1968). Since that time our knowledge of the structure and replication of these viruses has increased steadily and has been periodically reviewed (McIntosh 1974; Tyrrell et al. 1978; Robb and Bond 1979a). The basis for this review, which concentrates on the molecular biology of coronaviruses, is principally the new data which has become available in the last 2 years. The pathogeni-city of these viruses, which are associated with many diseases of clinical importance in animals and humans, is the subject of the accompanying article.

Parkinsonism in HIV dementia.

Summary. A great number of human immunodeficiency virus (HIV)-infected patients develop a central nervous system disorder, commonly called HIV dementia or AIDS dementia complex (ADC). HIV dementia is independent of opportunistic infections and is due to the virus itself. Symptoms include psychomotor slowing, apathy and motor disorders similar tothe bradykinesia and postural and gait abnormalities observed in late Parkinsons disease. Consequently, HIV has been discussed during the last few years as an additional cause for parkinsonism, and parkinsonian syndromes as manifestations of HIV dementia. Moreover, the early phase of HIV infection gains increasing interest because of studies which report subtle neurological symptoms at this stage. Accordingly, we found in SIV-infected monkeys that dopamine is reduced by 44% within as few as two months of infection, indicating that changes during early infection must be thoroughly evaluated. In this short review, we discuss alterations in the nigrostriatal dopaminergic system during early and late immunodeficiency virus infection and the common clinical and biochemical features shared by HIV dementia and Parkinsons disease.

Corona virus induced subacute demyelinating encephalomyelitis in rats: a morphological analysis.

SummaryThirty percent of weanling rats infected with JHM murine corona virus developed a subacute demyelinating encephalomyelitis approximately 3 weeks after intracerebral inoculation. Small demyelinating foci were located in the deep cerebral white matter and large, sharply demarcated demyelinating lesions were detectable in optic chiasma, pons and spinal cord. Axons as well as neurons were well preserved in the demyelinating plaques in areas where the lesions extended to the gray matter. Perivascular cuffings, consisting of plasma cells and mononuclear cells, were frequently found.Viral antigen was found mostly in the white matter and in glial cells, leaving neurons unstained. Electron microscopic studies of the early lesions of white matter disclosed two different kinds of cell degeneration which developed prior to the myelin disruption and mononuclear cell infiltration. One was a small pyknotic cell, which is thought to be an oligodendrocyte and the other is a ballooned cell containing abundant microtubules. Virus particles could be demonstrated only in the latter cell type. Discussion about astrocytes as well as oligodendrocytes was made in relation to the initial stage of demyelination caused by virus infection. This animal model may be useful in the analysis of the mechanisms leading to demyelination in subacute or chronic infections.

Coronavirus JHM: Intracellular Protein Synthesis

Coronavirus JHM contained six major proteins, four of which were glycosylated. The proteins were gp170, gp98, gp65, p60, gp25 and p23. Sac(-) cells infected with JHM shut off host cell protein synthesis, and the synthesis of three major (150K, 60K and 23K) and three minor (65K, 30K and 14K) polypeptides was detected by pulse-labelling with 35S-methionine. Antiserum directed against purified virus proteins specifically immunoprecipitated the three major intracellular species and also the 65K minor species. During a chase period, species 150K and 23K were processed and three new immunoprecipitable species, 170K, 98K and 25K appeared. The intracellular species 170K, 98K, 65K, 60K, 25K and 23K co-electrophoresed with virion proteins. Two-dimensional gel electrophoresis of infected cell polypeptides showed that the 60K, 23K, 25K and 14K species were relatively basic polypeptides whilst the 98K and 170K were relatively acidic and heterogeneously charged polypeptides. Additionally, a charge-size modification of the 23K species during processing was detected, which was not apparent using one-dimensional gel analysis.

Differential receptor usage by measles virus strains.

Recently, we demonstrated that infection of cells with all measles virus (MV) strains tested was inhibited by antibodies against CD46, although not all strains caused downregulation of the MV receptor CD46 from the surface of human cells. We now show that infection of cells with MV strain WTFb, a variant of wild-type isolate WTF which has been isolated and propagated on human BJAB cells, is not inhibited by antibodies against CD46. In contrast, infection of cells with the closely related strain WTFv, a Vero cell-adapted variant of WTF, is inhibited by antibodies against CD46. This observation led us to investigate the interaction of these viruses and the vaccine strain Edmonston (Edm) with CD46 and target cells. Cellular receptors with high affinity binding for WTFb are present on BJAB cells, but not on transfected CD46-expressing CHO cells. In contrast to the Edm strain, virus particles and solubilized envelope glycoproteins of WTFb have a very limited binding capacity to CD46. Furthermore, we show that recombinant soluble CD46 either does not bind, or binds very weakly, to WTFb glycoproteins expressed on the cell surface. Our findings indicate that wild-type MV strain WTFb and vaccine strain Edm use different binding sites on human cells. In addition, the results suggest that MV strains may alternatively use CD46 and an unknown molecule as receptors, and that the degree of usage of both receptors may be MV strain-specific.

Detection and identification of virus-specific, oligoclonal IgG in unconcentrated cerebrospinal fluid by immunoblot technique*

A technique is described which allows the detection of virus-specific oligoclonal IgG in unconcentrated cerebrospinal fluid (CSF) from patients with virus infections of the central nervous system. CSF is isoelectrically focused in agarose gels and immunoglobulins are blotted to nitrocellulose filters, passively loaded with either anti-human IgG or viral antigen. Transferred total IgG, as well as virus-specific IgG, is identified by the use of peroxidase-labelled anti-human IgG and 4-chloro-1-naphthol as a precipitating peroxidase substrate. Application of this assay in cases of SSPE, mumps meningitis and herpes simplex encephalitis demonstrates sensitivity and possible suitability of this technique for use in diagnosis of virus infections of the CNS.