Project Directorate on Foot-and-Mouth Disease, Indian Veterinary Research Institute Campus, Mukteswar, Nainital-263 138 (Uttaranchal), India1
Author for correspondence: Chakradhar Tosh. Fax +91 5942 88005. e-mail tosh64{at}epatra.com
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() |
---|
![]() |
Main text |
---|
![]() ![]() ![]() ![]() |
---|
FMD remains a major animal health concern in India, and the disease situation is aggravated by the presence of large populations of susceptible animals, low vaccine coverage, prevalence of multiple serotypes (O, Asia 1 and A) and unrestricted movement of susceptible animals in the country. Added to this, there is endemic cocirculation of multiple genotypes of type A virus (Tosh et al., 2002 ), and this may lead to recombination, which readily occurs between closely related strains (intratypic recombination) if multiple genotypes co-infect the host. In a preliminary antigenic analysis of type A FMDV, one-way antigenic relationships (r values; Ouldridge et al., 1984
) revealed lower values (r<0·40) between the isolates of genotype VI and VII when tested with serum raised against one of the isolates from genotype VI (Anon., 19982001
).
In this paper, we present the results of an analysis of the capsid-coding (P1) region of FMDV type A isolates, which provide evidence of intergenotypic recombination in one of the isolates.
Fourteen FMDV type A isolates were used in the present study; the detailed history of 13 isolates has been described previously (Nayak et al., 2001 ; Tosh et al., 2002
). They include IND 6/92 and the type A vaccine strain (IND 17/77), whose partial and complete VP1-encoding (1D) gene sequences, respectively, are already available. Isolate IND 194/00 was recovered from cattle (on 13 March 2000) in a field outbreak in the state of Assam. The P1 nucleotide sequence data of the type A vaccine strain (IND 17/77) was taken from GenBank (accession no. AF204108) for comparison. Genotype designations were made on the basis of complete 1D gene sequences (Samuel & Knowles, 2001
; Gurumurthy et al., 2002
; Tosh et al., 2002
). All the isolates were passaged three to five times in BHK-21 monolayer cell cultures. RNA was extracted from BHK-21 cell culture-infected supernatants using a Total RNA Isolation Kit (Qiagen), following the manufacturer's recommendations. The nucleotide sequences and the source of the primers used in the study are given in Table 1
. cDNA was synthesized using the AMV RT Kit (Promega) and a reverse-sense primer (pNK61) at 48 °C, following the recommendations of the supplier. PCR amplification of the P1 region was performed using the HotstarTaq kit (Qiagen) and the primers pNK61 and L01F, essentially following the recommendations of the supplier. The following thermal conditions were used for amplification: 1 cycle of 95 °C for 15 min, 35 cycles at 94 °C for 30 s, 50 °C for 30 s and 72 °C for 3 min, followed by 1 cycle at 72 °C for 10 min. Amplicons of the expected size (approximately 3·0 kb) were cleaned using the QIAquick gel extraction kit. Cycle sequencing was performed using the fmol kit (Promega) and Cy5-labelled primers (Table 1
). The reactions were run on an ALF Express II automated sequencer (Amersham Pharmacia Biotech). Sequences were aligned using the CLUSTAL W algorithm (Thompson et al., 1994
), available in the OMEGA 2.0 package (Oxford Molecular). Phylogenetic analysis of the aligned sequence was performed using the neighbour-joining program (Kimura two-parameter method) in the PHYLIP package (Felsenstein, 1993
). Robustness of the trees was evaluated by bootstrap analysis on 1000 replicate datasets. Phylogenetic trees were depicted using the TreeView program (Page, 1996
).
|
|
|
The analyses identified the isolate IND 170/88 as a recombinant of genotypes VI and VII. Most of the earlier research on picornaviruses identified recombination break points in the region towards the 3' end of the genome encoding the non-structural proteins (Cammack et al., 1988 ; King, 1988
; Wilson et al., 1988
; Furione et al., 1993
; Santti et al., 1999
), and only one report is available on recombination within the FMDV coat proteins involving isogenic parents (King et al., 1982a
). This report provides direct evidence of recombination in the structural protein-coding region in type A FMDV involving two heterogenic parents.
In conclusion, exchange of different genomic regions between viruses by recombination contributes directly to FMDV diversification and evolution. Recombination, particularly in the structural protein-coding region, as revealed in this study, may provide selective advantage to the virus, as the pre-existing immunity towards either of the parental viruses may not afford complete protection. This is of particular concern to countries where multiple FMDV genotypes co-circulate.
![]() |
Acknowledgments |
---|
![]() |
References |
---|
![]() ![]() ![]() ![]() |
---|
Anon. (19982001). Annual report: All India Coordinated Research Project for Epidemiological Studies on Foot-and-Mouth Disease. New Delhi: Indian Council of Agricultural Research.
Baxt, B., Vakharia, V., Moore, D. M., Franke, A. J. & Morgan, D. O. (1989). Analysis of neutralizing antigenic sites on the surface of type A12 foot-and-mouth disease virus. Journal of Virology 63, 2143-2151.[Medline]
Bolwell, C., Brown, A. L., Barnett, P. V., Campbell, R. O., Clarke, B. E., Parry, N. R., Ouldridge, E. J., Brown, F. & Rowlands, D. J. (1989a). Host cell selection of antigenic variants of foot-and-mouth disease virus. Journal of General Virology 70, 45-57.[Abstract]
Bolwell, C., Clarke, B. E., Parry, N. R., Ouldridge, E. J., Brown, F. & Rowlands, D. J. (1989b). Epitope mapping of foot-and-mouth disease virus with neutralizing monoclonal antibodies. Journal of General Virology 70, 59-68.[Abstract]
Borrego, B., Novella, I. S., Giralt, E., Andreu, D. & Domingo, E. (1993). Distinct repertoire of antigenic variants of foot-and-mouth disease virus in the presence or absence of immune selection. Journal of Virology 67, 6071-6079.[Abstract]
Cammack, N., Phillips, A., Dunn, G., Patel, V. & Minor, P. D. (1988). Intertypic genomic rearrangements of poliovirus strains in vaccines. Virology 167, 507-514.[Medline]
Domingo, E., Davila, M. & Ortin, J. (1980). Nucleotide sequence heterogeneity of the RNA from a natural population of foot-and-mouth disease virus. Gene 11, 333-346.[Medline]
Domingo, E., Martinez-Salas, E., Sobrino, F., de la Torre, J. C., Portela, A., Ortin, J., Lopez-Galindez, C., Perez-Brena, P., Villanueva, N., Najera, R., VandePol, S., Steinhauer, D., De Polo, N. & Holland, J. J. (1985). The quasispecies (extremely heterogeneous) nature of viral RNA genome populations: biological relevance a review. Gene 40, 1-8.[Medline]
Domingo, E., Mateu, M. G., Martinez, M. A., Dopazo, J., Moya, A. & Sobrino, F. (1990). Genetic variability and antigenic diversity of foot-and-mouth disease virus. In Applied Virology Research, vol. 2, Virus Variation and Epidemiology, pp. 233266. Edited by E. Kurstak, R. G. Marusyk, S. A. Murphy & M. H. V. Van Regenmortel. New York: Plenum.
Domingo, E., Escarmis, C., Martinez, M. A., Martinez-Salas, E. & Mateu, M. G. (1992). Foot-and-mouth disease virus populations are quasispecies. Current Topics in Microbiology and Immunology 176, 33-47.[Medline]
Domingo, E., Diez, J., Martinez, M. A., Hernandez, J. H., Holguin, A., Borrego, B. & Mateu, M. G. (1993). New observations on antigenic diversification of RNA viruses. Antigenic variation is not dependent on immune selection. Journal of General Virology 74, 2039-2045.[Abstract]
Felsenstein, J. (1993). PHYLIP (Phylogeny Inference Package), version 3.5c. University of Washington, Seattle, WA, USA.
Furione, M., Guillot, S., Otelea, D., Balanant, J., Candrea, A. & Crainic, R. (1993). Polioviruses with natural recombinant genomes isolated from vaccine-associated paralytic poliomyelytis. Virology 196, 199-208.[Medline]
Gebauer, F., de la Torrre, J. C., Gomes, I., Mateu, M. G., Barahona, H., Tiraboschi, B., Bergmann, I., Auge de Mello, P. & Domingo, E. (1988). Rapid selection of genetic and antigenic variants of foot-and-mouth disease virus during persistence in cattle. Journal of Virology 62, 2041-2049.[Medline]
Giraudo, A. T., Sagedahl, A., Bergmann, I. E., La Torre, J. L. & Scodeller, E. A. (1987). Isolation and characterization of recombinants between attenuated and virulent aphthovirus strains. Journal of Virology 61, 419-425.[Medline]
Gurumurthy, C. B., Sanyal, A., Venkataramanan, R., Tosh, C., George, M. & Hemadri, D. (2002). Genetic diversity in the VP1 gene of foot-and-mouth disease virus serotype Asia 1. Archives of Virology 147, 85-102.[Medline]
Holguin, A., Hernandez, J., Martinez, M. A., Mateu, M. G. & Domingo, E. (1997). Differential restrictions on antigenic variation among antigenic sites of foot-and-mouth disease virus in the absence of antibody selection. Journal of General Virology 78, 601-609.[Abstract]
King, A. M. Q. (1988). Recombination in positive strand RNA viruses. In RNA Genetics , pp. 149-165. Edited by E. Domingo, J. J. Holland & P. Ahlquist. Boca Raton, FL:CRC Press.
King, A. M. Q., McCahon, D., Slade, W. R. & Newman, J. W. I. (1982a). Biochemical evidence of recombination within the unsegmented RNA genome of aphthovirus. Journal of Virology 41, 66-77.[Medline]
King, A. M. Q., McCahon, D., Slade, W. R. & Newman, J. W. I. (1982b). Recombination in RNA. Cell 29, 921-928.[Medline]
King, A. M. Q., McCahon, D., Saunders, K., Newman, J. W. I. & Slade, W. R. (1985). Multiple sites of recombination within the RNA genome of foot-and-mouth disease virus. Virus Research 3, 373-384.[Medline]
Kirkegaard, K. & Baltimore, D. (1986). The mechanism of RNA recombination in poliovirus. Cell 47, 433-443.[Medline]
Knowles, N. J. & Samuel, A. R. (1995). Polymerase chain reaction amplification and cycle sequencing of the 1D (VP1) gene of foot-and-mouth disease viruses. In Report of the Session of the Research Group of the Standing Technical Committee of the European Commission for the Control of Foot-and-Mouth Disease, Vienna, Austria, September 1994, pp. 4553. Rome: FAO.
Krebs, O. & Marquardt, O. (1993). Identification and characterization of foot-and-mouth disease virus O1 Burgwedel/1987 as an intertypic recombinant. Journal of General Virology 73, 613-619.[Abstract]
Lai, M. M. C. (1992). Genetic recombination in RNA viruses. Current Topics in Microbiology and Immunology 176, 21-32.[Medline]
McCahon, D. & Slade, W. R. (1981). A sensitive method for the detection and isolation of recombinants of foot-and-mouth disease virus. Journal of General Virology 53, 333-342.[Abstract]
McCahon, D., King, A. M. Q., Roe, D. S., Slade, W. R., Newman, J. W. I. & Cleary, A. M. (1985). Isolation and biochemical characterization of intertypic recombinants of foot-and-mouth disease virus. Virus Research 3, 87-100.[Medline]
Malirat, V., Auge de Mello, P., Tiraboschi, B., Beck, E., Gomes, I. & Bergmann, I. E. (1994). Genetic variation of foot-and-mouth disease virus during persistent infection in cattle. Virus Research 34, 31-48.[Medline]
Nayak, B., Pattnaik, B., Tosh, C., Sanyal, A., Hemadri, D., Patil, S. S. & Venkataramanan, R. (2001). Genetic and antigenic analysis of type A foot-and-mouth disease viruses isolated in India during 19871996. Acta Virologica 45, 13-21.[Medline]
Ouldridge, E. J., Barnett, P. V., Hingley, P. J., Head, M. & Rweyemamu, M. M. (1984). The differentiation of foot-and-mouth disease virus strains using an indirect sandwich enzyme-linked immunosorbent assay saturation model. Journal of Biological Standardization 12, 367377.[Medline]
Page, R. D. M. (1996). TREEVIEW: an application to display phylogenetic trees on personal computers. Computer Applications in the Biosciences 12, 357-358.[Medline]
Pereira, H. G. (1977). Subtyping of foot-and-mouth disease virus. Developments in Biological Standardization 35, 167-174.
Pringle, C. R. (1965). Evidence of genetic recombination in foot-and-mouth disease virus. Virology 25, 48-54.[Medline]
Ray, S. C. (1999). SimPlot for Windows 95, version 2.5. Distributed by the author (www.med.jhu.edu/deptmed/sray/download).
Reuckert, R. R. (1996). Picornaviridae: the viruses and their replication. In Fields Virology , pp. 609-654. Edited by B. N. Fields, D. M. Knipe & P. M. Howley. Philadelphia:LippincottRaven.
Roberts, P. J. & Belsham, G. J. (1995). Identification of critical amino acids within the foot-and-mouth disease virus leader protein, a cysteine protease. Virology 213, 140-146.[Medline]
Robertson, D. L., Sharp, P. M., McCutchan, F. E. & Hahn, B. H. (1995). Recombination in HIV-1. Nature 374, 124-126.[Medline]
Sabarinath, G. P. (2001). Comparison of nucleotide sequence of structural protein encoding (P1) region of foot-and-mouth disease virus serotype O isolates. MVSc thesis (submitted to the Deemed University, Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh, India).
Salminen, M. O., Carr, J. K., Burke, D. S. & McCutchan, F. E. (1995). Identification of breakpoints in intergenotypic recombinants of HIV type 1 by bootscanning. AIDS Research and Human Retroviruses 11, 1423-1425.[Medline]
Salt, J. S. (1993). The carrier state in foot-and-mouth disease an immunological review. British Veterinary Journal 149, 207-223.[Medline]
Samuel, A. R. & Knowles, N. J. (2001). Foot-and-mouth disease type O viruses exhibit genetically and geographically distinct evolutionary lineages (topotypes). Journal of General Virology 82, 609-621.
Santti, J., Hyypia, T., Kinnunen, L. & Salminen, M. (1999). Evidence of recombination among enteroviruses. Journal of Virology 73, 8741-8749.
Sobrino, F., Davila, M., Ortin, J. & Domingo, E. (1983). Multiple genetic variants arise in the course of replication of foot-and-mouth disease virus in cell culture. Virology 128, 310-318.[Medline]
Thomas, A. A. M., Woortmeijer, R. J., Puijk, W. & Barteling, S. J. (1988). Antigenic sites on foot-and-mouth disease virus type A10. Journal of Virology 62, 2782-2789.[Medline]
Thompson, J. D., Higgins, D. G. & Gibson, T. J. (1994). CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice. Nucleic Acids Research 22, 4673-4680.[Abstract]
Tosh, C., Venkataramanan, R., Hemadri, D., Sanyal, A., Samuel, A. R., Knowles, N. J. & Kitching, R. P. (2000). Nucleotide sequence of the structural protein-encoding region of foot-and-mouth disease virus A22-India. Virus Genes 20, 269276.
Tosh, C., Sanyal, A., Hemadri, D. & Venkataramanan, R. (2002). Phylogenetic analysis of serotype A foot-and-mouth disease virus isolated in India between 1977 and 2000. Archives of Virology 147, 493-513.[Medline]
Wilson, V., Taylor, P. & Desselberger, U. (1988). Crossover regions in foot-and-mouth disease virus (FMDV) recombinants correspond to regions of high local secondary structure. Archives of Virology 102, 131-139.[Medline]
Woodbury, E. L. (1995). A review of the possible mechanisms for the persistence of foot-and-mouth disease virus. Epidemiology and Infection 114, 1-13.[Medline]
Received 22 February 2002;
accepted 10 May 2002.