Epidemiologic and genetic studies of paramyxoviruses

Abstract: Paramyxoviruses are viruses of the paramyxoviridae family of the mononegavirales order; they are negative-sense single-stranded RNA viruses responsible for a number of diseases in humans and animals. Mumps virus belongs to the Rubula virus genus. Mumps is a common childhood infection. The central nervous system (CNS) is a common site of mumps virus dissemination with or without signs of parotid gland involvement. In recent years the degree of neuropathogenicity has been estimated for specific mumps virus strains. The symptoms of mumps have been compared with individual virus strains /genotypes in clinical settings, and there has been evidence that certain virus strains are more neurovirulent than others. Thirteen different genotypes of mumps virus small hydrophobic (SH) gene, designated A to M, have been found in the beginning of the year 2008. Respiratory Syncytial virus (RSV) is a member of the subfamily pneumovirinae and genus pneumovirus. RSV is the most common viral pathogen for lower respiratory tract infection among infants and young children, and it is recognized as an important agent for respiratory disease in the elderly and in transplant patients. Antigenic characterization of RSV strains with monoclonal antibodies has identified two distinct groups of the virus, RSV group A and B, which circulate worldwide.The existence of eight group A genotypes , named GA1 to GA7 and SAA1 have been reported. Human metapneumovirus (hMPV) also belonging to the subfamily pneumovirinae and genus metapneumovirus has been identified in 2001 in Nederlands. It is the only member capable of infecting humans in the genus metapneumovirus. Genotyping studies have determined that hMPV can be classified into two main linages and four sublinges, named A1, A2, B1, B2. The frequency of the hMPV detection in patients with respiratory tract infection ranged from 1, 5 to 41%. The aims of this thesis was ( i ) to investigate the sequences of the HN and F protein genes for the neurovirulent C1 and non-neurovirulent C2 variants and compare the results with the HN and F proteins of known genotypes. As the Kilham strain is neurovirulent, but other genotype A viruses are not, it was considered of interest to see if a mutation in the HN protein could explain its neuropathogenic capacity. ( ii) to study the molecular epidemiology of RSV of group A isolated in Stockholm over a longer time period. (iii) to develop a simple and highly efficient method for genotyping group A of RS virus. ( iv) to study the epidemiology of hMPV in Stockholm, Sweden, and compare the epidemiology of hMPV with five other common respiratory viruses. During an epidemic in Lithuania in 1998-2000 mumps virus strains isolated were studied. Viruses of the C1 and C2 small hydrophobic (SH) genotypes variant were sequenced for the HN and F protein genes. Amino acid differences between C1 and C2 strains were found for both proteins. Two amino acid differences were of potential importance for the non-neurovirulent phenotype of the C2 virus. These amino acid differences have previously been reported to associate with a change in neurovirulence and fusion activity. In addition, the HN gene of the neurovirulent Kilham strain of genotype A was sequenced. The deduced amino acid sequence showed different amino acids compared to both genotype A and genotype C on some positions. The epidemiology of respiratory syncytial virus (RSV) group A was followed by nucleotide sequencing of the variable parts of the glycoprotein (G) gene. The amino acid sequences of an amino-terminal and carboxy-terminal amino acid portion of the G protein in 47 virus strains collected in Stockholm, between 1965 and 2004, were determined. Phylogenetic analysis jointly with previously described genotypes (GA1 to GA7 and SAA1), showed that 34 virus strains belonged to genotype GA5, seven to GA2, three to genotype GA1, one to genotype GA4 and two to genotype GA7. Genotype GA5 was predominant in four epidemics, between 2000/2001 and 2003/2004. Genotyping of respiratory syncytial (RS) virus group A by the use of a novel method based on reverse-transcriptase polymerase chain reaction (RT-PCR), FRET (fluorescence resonance energy transmission) based detection and two-dimensional melting curve analysis was applied on eighty RS virus samples of group A collected in Stockholm from 1976 to 2005. The Tm values were assessed for three different genotypes (GA2, GA5 and GA7) circulating in Sweden using two pairs of probes and subsequent data analysis by plotting results in a two-dimensional system. The results obtained were compared to genotyping by conventional nucleotide sequencing and phylogenetic tree analysis. It was found that the new assay was able to make a correct genotype identification in about 89% of the isolates and identified the remaining 11% as untypeable and candidates for conventional nucleotide sequencing. The human metapneumovirus (hMPV) was analyzed retrospectively, by RT-PCR in five epidemic seasons, 2002-2006. After respiratory syncytial virus (RSV), influenza A virus and parainfluenza virus hMPV was the fourth most common respiratory virus with a detection rate of 2,9% (n=143/4,989) in nasopharyngeal samples. genotype A dominated over genotype B. Approximately 2,6% (n=68/2,579) of the hMPV positive patients were <3 years, 3,1% (75/2,410) were ≥3 years and 2,5% (n=52/2,122) were <1 year. This age distribution differed from RSV, influenza A, B and parainfluenza virus. hMPV epidemics peaked in March, not coincident with RSV or parainfluenza virus. Large hMPV virus epidemics occurred biannually and were anticyclical to RSV epidemics.