Public Health Laboratory1 and Department of Molecular Microbiology, University Medical School2, Southampton General Hospital, Tremona Road, Southampton SO16 6YD, UK
Regional Virus Laboratory, Public Health Laboratory, Myrtle Road, Bristol BS2 8EL, UK3
Author for correspondence: Vivienne James. Present address: Central Public Health Laboratory, 61 Colindale Avenue, London NW9 5HT, UK. Fax +44 20 8205 1488. e-mail vjames{at}phls.nhs.uk
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Abstract |
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Introduction |
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The rotavirus genome consists of 11 segments of double-stranded (ds) RNA that encode six structural and five non-structural proteins. The different rotavirus groups can be identified by the characteristic profile of the dsRNA on SDSPAGE and defined by terminal fingerprint analysis of the genome segments (Pedley et al., 1986 ). Sequence data are available for all eleven genome segments of the porcine group C rotavirus Cowden strain, although many of these are incomplete and do not have defined 5' and 3' termini. Sequences are also available for genome segments 36, 8 and 10 of the bovine group C rotavirus Shintoku strain. Genes corresponding to the structural proteins VP1 and VP2 and the non-structural proteins NSP1, 2 and 3 of human group C rotavirus have not been determined. The purpose of this work was to determine the coding assignments and sequences of the non-structural proteins NSP1, 2 and 3 of human group C rotavirus and to compare them with the corresponding porcine group C and mammalian group A rotavirus proteins.
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Methods |
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M13 clones for each of the 11 gene segments of human group C rotavirus Bristol strain have been constructed and stored as an ordered genomic library (Lambden et al., 1992 ). Briefly, the cloning method involved ligation of a single amino-linked modified oligonucleotide to the 3' termini of each dsRNA genome segment by using T4 RNA ligase. The tailed RNA was then converted to cDNA by using a complementary primer and reverse transcriptase. The resultant cDNA was annealed and repaired and then amplified by PCR by using a single complementary oligonucleotide primer. The PCR products were ligated into dephosphorylated, SmaI-cleaved M13 mp8 and transformed into E. coli JM101. This method increased the probability that the resultant clones would contain full-length inserts of the genes.
32P-labelled cDNA probes used for gene assignment studies were generated as described previously (Deng et al., 1995 ) and the coding assignments were determined by Northern blot analysis.
Sequence analysis.
Sequences were compiled from M13 clones and by direct sequencing of RTPCR amplicons. Recombinant M13 templates were prepared by standard techniques and sequenced initially with universal primer designed to hybridize 49 bp upstream of the SmaI cloning site by using the ABI PRISM terminator cycle sequencing kit (Applied Biosystems) and an Applied Biosystems model 373A automated sequencer. The complete sequences were determined by preparing primers and sequencing stepwise along the gene from each previous sequence. Internal primer sequences were designed approximately 150 bp upstream of the previously deduced sequence and resulted in read-lengths of 400500 bp. The oligonucleotides were synthesized on a Millipore Expedite 8909 automated synthesizer by using -cyanoethyl phosphoramidite chemistry. Computer analyses of the sequence data were performed by using the Lasergene software (DNASTAR).
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Results and Discussion |
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Gene segment 6
Genome segment 6, encoding the NSP3 (NS34) protein, is 1350 nucleotides long, the same length as the cognate bovine Shintoku gene and two nucleotides longer than the genome sequence for the porcine Cowden strain. The two extra nucleotides are at positions 1250 and 1251 in the human and bovine group C sequences in the 3' non-coding region. The NSP3 genome segment showed 78% identity to the porcine Cowden equivalent and 79% identity to the bovine Shintoku NSP3 gene. Similarly, the Cowden and Shintoku NSP3 genes share 78% sequence identity. The ATG translation initiation codon is at position 25, in a favourable context for translation initiation (Kozak, 1991 ). Computer analysis of the sequence revealed a single ORF of 1206 nucleotides and a 3' non-coding region of 120 nucleotides.
The amino acid sequence predicted a protein 402 amino acids long, acidic in nature (Fig. 1), with a predicted molecular mass of 45·3 kDa and a calculated isoelectric point of 4·85. Eight cysteine residues were conserved between the human, porcine and bovine NSP3 proteins at positions 60, 171, 200, 280, 284, 294, 353 and 369. Three major functional domains described for the group A rotavirus SA11F NSP3 protein (Mattion et al., 1992
) were also present on the group C NSP3 protein. The regions are a basic region for single-stranded (ss) RNA binding, a heptad repeat region for oligomerization and a leucine zipper motif. The group A rotavirus NSP3 protein basic region extends from amino acids 81 to 150; this region is also charged in the group C rotaviruses, with a large number of lysine and arginine residues between amino acids 79 and 151. The region between amino acids 89 and 125 is conserved in the group C rotaviruses, with three amino acid changes between human and bovine strains and just two amino acid changes between the human and porcine strains. The consensus sequence for ssRNA-binding, (I/L)XXM(I/L)(S/T)XXG, seen in orbiviruses, reoviruses and group A rotaviruses (Rao et al., 1995
) is found in this region and is located at amino acids 104112 of NSP3 of SA11. In the human and bovine group C rotaviruses, the sequence 100LXXMLSXXG108 is conserved and is highlighted in Fig. 1
. This is at variance with the Cowden sequence, where the methionine residue is replaced by isoleucine (Qian et al., 1991
).
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Besides ssRNA binding, the NSP3 proteins of the group C rotaviruses have been shown to bind dsRNA (Langland et al., 1994 ). The group C rotavirus NSP3 proteins are 8789 amino acids longer than their group A counterparts. The carboxy-terminal region of the group C NSP3 protein contains the dsRNA-binding domain, with a consensus sequence LX39(G/A)XGXSKKXAKXXAAXX(A/I)LXXL (Langland et al., 1994
). This consensus region is also present in human group C rotavirus (amino acids 340400).
Gene segment 7
The non-structural protein NSP1 (NS53) is encoded by gene 7 in group C rotaviruses. The gene sequence is 1270 nucleotides long. Computer analysis revealed a single ORF of 1182 nucleotides with a 5' non-coding region of 37 bp and a 3' non-coding region of 48 bp. The nucleotide sequence GTCAACATGG in the region of the putative ATG codon (underlined), with A at -3 and G at +4, constitutes a strong signal for initiation of translation (Kozak, 1991 ). The human group C rotavirus gene 7 sequence is 35 nucleotides longer than the porcine equivalent (Bremont et al., 1993
), with an extra 31 nucleotides at the 5' terminus, one extra base at the 3' terminus and a novel extra codon at positions 135137. The sequence has 67% identity to the equivalent Cowden sequence.
The predicted protein sequence contains 394 amino acid residues, with a molecular mass of 46·6 kDa (Fig. 2). It is a basic protein with a calculated isoelectric point of 8·9. Human group C rotavirus NSP1 has 61% amino acid sequence identity to the equivalent Cowden sequence, which contains 393 amino acids. This is consistent with other reports that suggest that NSP1 is the most variable rotavirus protein and that diversity in sequences is evident between strains from different species (Hua et al., 1993
; Dunn et al., 1994
). Studies examining reassortants between rotavirus strains have implicated NSP1 in host-range selection (Graham et al., 1987
; Palombo & Bishop, 1994
) and pathogenic phenotype (Broome et al., 1993
).
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Gene segment 9
Gene segment 9 encodes the non-structural protein NSP2 (NS35). The nucleotide sequence is 1037 bp long and shares 88% identity (between nucleotides 21 and 1015) to the gene 9 sequence of the porcine Cowden strain (Bremont et al., 1993 ). The sequence contains a single ORF of 936 nucleotides with a 5' untranslated leader sequence of 42 bp and an initiator codon in a favourable context for translation initiation (Kozak, 1991
). The predicted protein sequence of 312 amino acids has a predicted molecular mass of 35·9 kDa and has 93·9% identity to the Cowden NSP2 protein (Fig. 3
).
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The first seven amino acids are absolutely conserved between all group A and group C rotavirus NSP2 proteins. Only one cysteine residue is absolutely conserved between the group A and group C NSP2 proteins, at position 6. Truncated NSP2 loses its ability to bind RNA when more than four amino acids are absent from the termini (Kattoura et al., 1992 ): therefore the conserved cysteine at position 6 could be involved in stabilizing the oligomerized NSP2 proteins.
Two other regions are well conserved between the group A and group C rotavirus NSP2 proteins. The first is a basic region (amino acids 4760) with a run of three proline residues with the consensus sequence IXYGXAPPPXF(K/N/R)(K/N/R)R. Proline-rich domains have been identified in several transcription regulators that both activate and repress gene transcription (Hanna-Rose & Hansen, 1996 ). NSP2 localizes in the viroplasm (Petrie et al., 1984
) and NSP2 has been shown in vivo to form part of a viral enzyme complex with replicase activity (Aponte et al., 1996
). Proline-rich activation motifs are frequently associated with serine and threonine residues (Hanna-Rose & Hansen, 1996
). NSP5, which also localizes in the viroplasm, interacts with NSP2 (Poncet et al., 1997
) and has a high serine and threonine content. The interaction between NSP2 and NSP5 could regulate rotavirus RNA synthesis.
Between amino acid residues 109 and 139, a second, conserved, basic region is located followed by the first of the hydrophobic heptad repeats with consensus sequence (V/I)RHLENLX2RX5D(V/I)LX5LX5(L/M)I. NSP2 has been shown to form a complex with VP1, the putative RNA polymerase, in infected cells (Kattoura et al., 1994 ). Furthermore, NSP2 has an affinity for dsRNA (Kattoura et al., 1992
) and associates with all 11 dsRNA segments, although Aponte et al. (1996)
showed that, in the replicase complexes, some regions of the RNAs were single stranded, supporting the theory of a regulatory role in RNA replication.
Phylogenetic analyses
Genomic reassortment during co-infection in vivo and in vitro with distinct strains of group A rotaviruses is well documented (Ramig, 1997 ). Evolution of the different rotavirus serogroups could have arisen because of reassortment events and divergent evolution. Alternatively, the different rotavirus serogroups may have arisen from a common ancestor, in which case all genes would display equal divergence. To test these hypotheses, unrooted phylogenetic trees were constructed for groups A, B and C rotavirus NSP1, 2, 3, 4 and 5 gene sequences. Multiple alignments were performed with CLUSTAL X (Thompson et al., 1997
) and unrooted trees were generated by using the neighbour-joining method (Saitou & Nei, 1987
). Trees were subjected to a bootstrap analysis (Felsenstein, 1985
) using 1000 data sets and output as a graphic representation by using DRAWTREE in the PHYLIP package (Felsenstein, 1993
).
Non-structural proteins have been considered to be more useful than structural proteins in determining the ancestral relationships of different viruses (Clewley, 1998 ). Expression of the non-structural proteins is confined within the host cell and non-structural proteins are not subject to the conformational structural constraints that affect the structural proteins, suggesting they would be subject to less evolutionary pressure than the structural proteins. However, Xu et al. (1994)
demonstrated that divergence of gene 5 of group A rotaviruses encoding NSP1 exceeded that of the divergence of the major neutralizing antigen, VP7. Phylogenetic analysis of NSP3 proteins (Rao et al., 1995
) revealed that the rodent group B rotavirus (IDIR), porcine group C rotavirus (Cowden) and various mammalian group A rotavirus strains belonged to three different groups with a distinct ancestral origin. In the study presented here, the phylogenetic trees for each of the five genes encoding the non-structural proteins were essentially similar (for NSP3 comparison see Fig. 4
), with three major branches, of similar length, for the three serogroups analysed. This branching pattern suggests that the groups may have diverged at the same time, possibly from a common ancestor, rather than as a result of reassortment of the genes. No reassortants have been detected between strains of different rotavirus groups (AG) (Taniguchi & Urasawa, 1995
) and divergence of the rotavirus serogroups is so great that reassortment between viruses from different serogroups does not appear to occur (Yolken et al., 1988
). Comparison of the group C branch of the phylogenetic trees suggests that the human, porcine and bovine group C rotaviruses also developed from a common ancestor at a later time. It may still be possible for reassortment to occur between group C rotaviruses from different animal species in vitro, although there is no evidence for naturally occurring reassortment (Grice et al., 1994
; Jiang et al., 1996
).
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Acknowledgments |
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Footnotes |
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References |
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Bishop, R. F., Davidson, G. P., Holmes, I. H. & Buck, B. J. (1973). Virus particles in epithelial cells of duodenal mucosa from children with acute non-bacterial gastroenteritis. Lancet ii, 12811283.
Bremont, M., Charpilienne, A., Chabanne, D. & Cohen, J. (1987). Nucleotide sequence and expression in Escherichia coli of the gene encoding the nonstructural protein NCVP2 of bovine rotavirus. Virology 161, 138-144.[Medline]
Bremont, M., Chabanne-Vautherot, D. & Cohen, J. (1993). Sequence analysis of three non structural proteins of a porcine group C (Cowden strain) rotavirus. Archives of Virology 130, 85-92.[Medline]
Broome, R. L., Vo, P. T., Ward, R. L., Clark, H. F. & Greenberg, H. B. (1993). Murine rotavirus genes encoding outer capsid proteins VP4 and VP7 are not major determinants of host range restriction and virulence. Journal of Virology 67, 2448-2455.[Abstract]
Brown, D. W. G., Beards, G. M., Chen, G. M. & Flewett, T. H. (1987). Prevalence of antibody to group B (atypical) rotavirus in humans and animals. Journal of Clinical Microbiology 25, 316-319.[Medline]
Caul, E. O., Ashley, C. R., Darville, J. M. & Bridger, J. C. (1990). Group C rotavirus associated with fatal enteritis in a family outbreak. Journal of Medical Virology 30, 201-205.[Medline]
Clewley, J. P. (1998). A users guide to producing and interpreting tree diagrams in taxonomy and phylogenetics. Communicable Disease and Public Health 1, 64-66.[Medline]
Cohen, C. & Parry, D. A. D. (1986). Alpha-helical coiled coils a widespread motif in proteins. Trends in Biochemical Sciences 11, 245-248.
Deng, Y., Fielding, P. A., Lambden, P. R., Caul, E. O. & Clarke, I. N. (1995). Molecular characterization of the 11th RNA segment from human group C rotavirus. Virus Genes 10, 239-243.[Medline]
Dunn, S. J., Cross, T. L. & Greenberg, H. B. (1994). Comparison of the rotavirus nonstructural protein NSP1 (NS53) from different species by sequence analysis and northern blot hydridization. Virology 203, 178-183.[Medline]
Felsenstein, J. (1985). Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39, 783-791.
Felsenstein, J. (1993). PHYLIP (phylogeny inference package), version 3.5c. Distributed by the author. Department of Genetics, University of Washington, Seattle, WA, USA.
Flewett, T. H., Bryden, A. S. & Davies, H. (1973). Virus particles in gastroenteritis. Lancet ii, 1497.
Graham, A., Kudesia, G., Allen, A. M. & Desselberger, U. (1987). Reassortment of human rotavirus possessing genome rearrangements with bovine rotavirus: evidence for host selection. Journal of General Virology 68, 115-122.[Abstract]
Grice, A. S., Lambden, P. R., Caul, E. O. & Clarke, I. N. (1994). Sequence conservation of the major outer capsid glycoprotein of human group C rotaviruses. Journal of Medical Virology 44, 166-171.[Medline]
Hanna-Rose, W. & Hansen, U. (1996). Active repression mechanisms of eukaryotic transcription repressors. Trends in Genetics 12, 229-234.[Medline]
Hua, J. & Patton, J. T. (1994). The carboxyl-half of the rotavirus nonstructural protein NS53 (NSP1) is not required for virus replication. Virology 198, 567-576.[Medline]
Hua, J., Mansell, E. A. & Patton, J. T. (1993). Comparative analysis of the rotavirus NS53 gene: conservation of basic and cysteine-rich regions in the protein and possible stemloop structures in the RNA. Virology 196, 372-378.[Medline]
Hua, J., Chen, X. & Patton, J. T. (1994). Deletion mapping of the rotavirus metalloprotein NS53 (NSP1): the conserved cysteine-rich region is essential for virus-specific RNA binding. Journal of Virology 68, 3990-4000.[Abstract]
Hung, T., Chen, G. M., Wang, C. G., Chou, Z. Y., Chao, T. X., Ye, W. W., Yao, H. L. & Meng, K. H. (1983). Rotavirus-like agent in adult non-bacterial diarrhoea in China. Lancet ii, 10781079.
Hung, T., Chen, G. M., Wang, C. A., Fan, R., Yong, R., Chang, J., Dan, R. & Ng, M. H. (1987). Seroepidemiology and molecular epidemiology of the Chinese rotavirus. In Novel Diarrhoea Viruses, pp. 49-62. Edited by G. Bock & J. Whelan. Chichester: John Wiley.
James, V. L. A., Lambden, P. R., Caul, E. O. & Clarke, I. N. (1998). Enzyme-linked immunosorbent assay based on recombinant human group C rotavirus inner capsid protein (VP6) to detect human group C rotaviruses in fecal samples. Journal of Clinical Microbiology 36, 3178-3181.
Jiang, B., Tsunemitsu, H., Dennehy, P. H., Oishi, I., Brown, D., Schnagl, R. D., Oseto, M., Fang, Z. Y., Avendano, L. F., Saif, L. J. & Glass, R. I. (1996). Sequence conservation and expression of the gene encoding the outer capsid glycoprotein among human group C rotaviruses of global distribution. Archives of Virology 141, 381-390.[Medline]
Kapikian, A. Z. & Chanock, R. M. (1996). Rotaviruses. In Fields Virology, pp. 1657-1708. Edited by B. N. Fields, D. M. Knipe & P. M. Howley. Philadelphia: LippincottRaven.
Kattoura, M. D., Clapp, L. L. & Patton, J. T. (1992). The rotavirus nonstructural protein, NS35, possesses RNA-binding activity in vitro and in vivo.Virology 191, 698-708.[Medline]
Kattoura, M. D., Chen, X. & Patton, J. T. (1994). The rotavirus RNA-binding protein NS35 (NSP2) forms 10S multimers and interacts with the viral RNA polymerase. Virology 202, 803-813.[Medline]
Kozak, M. (1991). Structural features in eukaryotic mRNAs that modulate the initiation of translation. Journal of Biological Chemistry 266, 19867-19870.
Krishnan, T., Sen, A., Choudhury, J. S., Das, S., Naik, T. N. & Bhattacharya, S. K. (1999). Emergence of adult diarrhoea rotavirus in Calcutta, India. Lancet 353, 380-381.[Medline]
Lambden, P. R. & Clarke, I. N. (1995). Cloning of viral double-stranded RNA genomes by single primer amplification. In Methods in Molecular Genetics, pp. 359-372. Edited by S. D. Aldolph. London: Academic Press.
Lambden, P. R., Cooke, S. J., Caul, E. O. & Clarke, I. N. (1992). Cloning of noncultivatable human rotavirus by single primer amplification. Journal of Virology 66, 1817-1822.[Abstract]
Langland, J. O., Pettiford, S., Jiang, B. & Jacobs, B. L. (1994). Products of the porcine group C rotavirus NSP3 gene bind specifically to double-stranded RNA and inhibit activation of the interferon-induced protein kinase PKR. Journal of Virology 68, 3821-3829.[Abstract]
Mattion, N. M., Cohen, J., Aponte, C. & Estes, M. K. (1992). Characterization of an oligomerization domain and RNA-binding properties on rotavirus nonstructural protein NS34. Virology 190, 68-83.[Medline]
Mitchell, D. B. & Both, G. W. (1990). Conservation of a potential metal binding motif despite extensive sequence diversity in the rotavirus nonstructural protein NS53. Virology 174, 618-621.[Medline]
Nakata, S., Estes, M. K., Graham, D. Y., Wang, S. S., Gary, G. W. & Melnick, J. L. (1987). Detection of antibody to group B adult diarrhea rotaviruses in humans. Journal of Clinical Microbiology 25, 812-818.[Medline]
Okada, J., Kobayashi, N., Taniguchi, K. & Urasawa, S. (1999). Analysis on reassortment of rotavirus NSP1 genes lacking coding region for cysteine-rich zinc finger motif. Archives of Virology 144, 345-353.[Medline]
Palombo, E. A. & Bishop, R. F. (1994). Genetic analysis of NSP1 genes of human rotaviruses isolated from neonates with asymptomatic infection. Journal of General Virology 75, 3635-3639.[Abstract]
Patton, J. T., Salter-Cid, L., Kalbach, A., Mansell, E. A. & Kattoura, M. (1993). Nucleotide and amino acid sequence analysis of the rotavirus nonstructural RNA-binding protein NS35. Virology 192, 438-446.[Medline]
Pedley, S., Bridger, J. C., Chasey, D. & McCrae, M. A. (1986). Definition of two new groups of atypical rotaviruses. Journal of General Virology 67, 131-137.[Abstract]
Petrie, B. L., Greenberg, H. B., Graham, D. Y. & Estes, M. K. (1984). Ultrastructural localization of rotavirus antigens using colloidal gold. Virus Research 1, 133-152.[Medline]
Poncet, D., Lindenbaum, P., LHaridon, R. & Cohen, J. (1997). In vivo and in vitro phosphorylation of rotavirus NSP5 correlates with its localization in viroplasms. Journal of Virology 71, 34-41.[Abstract]
Qian, Y., Jiang, B. M., Saif, L. J., Kang, S. Y., Ojeh, C. K. & Green, K. Y. (1991). Molecular analysis of the gene 6 from a porcine group C rotavirus that encodes the NS34 equivalent of group A rotaviruses. Virology 184, 752-757.[Medline]
Ramig, R. F. (1997). Genetics of the rotaviruses. Annual Review of Microbiology 51, 225-255.[Medline]
Rao, C. D., Das, M., Ilango, P., Lalwani, R., Rao, B. S. & Gowda, K. (1995). Comparative nucleotide and amino acid sequence analysis of the sequence-specific RNA-binding rotavirus nonstructural protein NSP3. Virology 207, 327-333.[Medline]
Rodger, S. M., Bishop, R. F. & Holmes, I. H. (1982). Detection of a rotavirus-like agent associated with diarrhea in an infant. Journal of Clinical Microbiology 16, 724-726.[Medline]
Saif, L. J., Bohl, E. H., Theil, K. W., Cross, R. F. & House, J. A. (1980). Rotavirus-like, calicivirus-like, and 23-nm virus-like particles associated with diarrhea in young pigs. Journal of Clinical Microbiology 12, 105-111.[Medline]
Saitou, N. & Nei, M. (1987). The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution 4, 406-425.[Abstract]
Taniguchi, K. & Urasawa, S. (1995). Diversity in rotavirus genomes. Seminars in Virology 6, 123-131.
Taniguchi, K., Kojima, K. & Urasawa, S. (1996a). Nondefective rotavirus mutants with an NSP1 gene which has a deletion of 500 nucleotides, including a cysteine-rich zinc finger motif-encoding region (nucleotides 156 to 248), or which has a nonsense codon at nucleotides 153155. Journal of Virology 70, 4125-4130.[Abstract]
Taniguchi, K., Kojima, K., Kobayashi, N., Urasawa, T. & Urasawa, S. (1996b). Structure and function of rotavirus NSP1. Archives of Virology Supplementum 12, 53-58.[Medline]
Thompson, J. D., Higgins, D. G. & Gibson, T. J. (1994). CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Research 22, 4673-4680.[Abstract]
Thompson, J. D., Gibson, T. J., Plewniak, F., Jeanmougin, F. & Higgins, D. G. (1997). The CLUSTAL X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Research 25, 4876-4882.
Tian, Y., Tarlow, O., Ballard, A., Desselberger, U. & McCrae, M. A. (1993). Genomic concatemerization/deletion in rotaviruses: a new mechanism for generating rapid genetic change of potential epidemiological importance. Journal of Virology 67, 6625-6632.[Abstract]
Tsunemitsu, H., Saif, L. J., Jiang, B. M., Shimizu, M., Hiro, M., Yamaguchi, H., Ishiyama, T. & Hirai, T. (1991). Isolation, characterization, and serial propagation of a bovine group C rotavirus in a monkey kidney cell line (MA104). Journal of Clinical Microbiology 29, 2609-2613.[Medline]
Xu, L., Tian, Y., Tarlow, O., Harbour, D. & McCrae, M. A. (1994). Molecular biology of rotaviruses. IX. Conservation and divergence in genome segment 5. Journal of General Virology 75, 3413-3421.[Abstract]
Yolken, R., Arango-Jaramillo, S., Eiden, J. & Vonderfecht, S. (1988). Lack of genomic reassortment following infection of infant rats with group A and group B rotaviruses. Journal of Infectious Diseases 158, 1120-1123.[Medline]
Received 29 March 1999;
accepted 4 August 1999.