JOURNAL OF CLINICAL MICROBIOLOGY, Oct. 1994, p. 2609-2612
Vol. 32, No. 10
0095-1137/94/$04.00+0 Copyright X 1994, American Society for Microbiology
Equine Rotaviruses with G14 Serotype Specificity Circulate among Venezuelan Horses MAX CIARLET,1 FELIPE REGGETI,1'2 CARMEN I. PINA,' AND FERDINANDO LIPRANDIl* Centro de Microbiologia y Biologia Celular, Instituto Venezolano de Investigaciones Cientificas, Caracas 1020-A,1 and Facultad de Ciencias Veterinarias, Universidad Central de Venezuela, Maracay7 Venezuela Received 25 March 1994/Accepted 11 July 1994
Two group A rotavirus strains isolated from diarrheic foals in Venezuela were classified as belonging to G14 serotype by cross-neutralization tests and on the basis of the homology of the sequenced VP7 gene. This report confirms that rotavirus strains of G14 serotype specificity circulate among equine populations.
Rotaviruses have emerged as an important group of viruses etiologically associated with neonatal diarrhea in a variety of animal species, including newborn horses (10). Since their first identification in young foals (11), rotaviruses have been isolated during epidemics of foal diarrhea in several parts of the world (2, 7, 8, 17, 22, 26). The two surface proteins of group A rotaviruses, VP4 and VP7, have been shown to independently elicit neutralizing antibodies and induce protective immunity (10). On the basis of neutralization assays, antigenic variants of VP4 and VP7 have been classified as P and G serotypes, respectively. Among group A rotaviruses, a total of 14 G serotypes have been identified in different species (2, 4, 10, 25). Reports of serotype analysis of equine rotavirus strains isolated to date are mainly limited to strains circulating in the United States and United Kingdom. Most strains have been classified as belonging to G3 serotype, whereas there are three instances of one isolate reported per serotype, namely, strains Hi (G5 [G serotype 5]), L338 (G13), and FI23 (G14) (1, 2, 17, 19, 20). During the course of a study on the incidence of rotavirus infections in six stud farms in north-central Venezuela in 1992 and 1993, of 42 diarrheic foals studied, samples from eight foals from two farms were found positive for group A rotavirus by enzyme-linked immunoassay (ELISA) and polyacrylamide gel electrophoresis analysis. All samples from both farms appeared to have the same electropherotype. Rotaviruses from three samples were successfully adapted to growth in tissue culture. The aim of this study was to characterize these rotavirus isolates to establish the G serotype specificity that circulates among Venezuelan horses. Equine rotaviruses FR4, FR5, and FR8 were isolated from fecal material of diarrheic foals after passage in rolling tubes of MA104 cells in the presence of 1 ,ug of trypsin per ml and were plaque purified three times before characterization. The electropherotypes of the cell-adapted strains were identical to those of the original fecal samples. The electropherotypes of strains FR4, FR5 (FR4 and FR5 isolated from one farm), and FR8 (isolated from another farm) were identical (Fig. 1). The isolated strains exhibited an electropherotype profile typical of many equine strains with segments 7, 8, and 9 clearly spaced and segments 3 and 4 migrating close together (1, 17). The subgroup of these equine strains was determined to be not subgroup I or II by ELISA with subgroup-specific monoclonal
antibodies (MAbs) 255/60 (subgroup I) and 631/9 (subgroup II) (16). Cross-neutralization tests were performed in 96-well plates by a fluorescent focus neutralization assay, essentially as described previously (21). Reference rotavirus strains Wa (Gl), DS-1 (G2), Rhesus (G3), Ch-2 (G7), 69M (G8), WI61 (G9), and their respective hyperimmune antisera (except for Ch-2), produced in guinea pigs, were kindly supplied by Y. Hoshino and M. Gorziglia (National Institute of Allergy and Infectious Diseases, Bethesda, Md.). Strains B223 (G10), L338 (G13), and F123 (G14) and their respective hyperimmune antisera (except for B223) were kindly provided by D. Snodgrass and P. Isa (Moredun Research Institute, Edinburgh, Scotland, United Kingdom). Hyperimmune antisera to strain FR4 and to the other standard rotavirus strains were produced as described previously (6). The results of the cross-neutralization assays between strains FR4, FR5, FR8 and standard rotavirus strains representative of 12 G serotypes are shown in Table 1. Different viral serotypes were defined by a >20-fold difference in neutralization titer. Strain FR4 showed a two-way neutralization reaction with G14 strain F123 and no reactivity with the remaining 11 strains. Strains FR5 and FR8, for which antisera were not produced, were efficiently neutralized by F123 and FR4 anti-
1
t11:5 a
1
Am
2 3 4
-
2
5
6
El
7
8
FIG. 1. Electrophoresis of genome RNAs of equine rotavirus strains. Lanes: 1, H1; 2, H-2; 3, FI14; 4, F123; 5, L338; 6, FR4; 7, FR8; 8, FR4 and FR8. Samples were subjected to electrophoresis in a 7% polyacrylamide gel, and genome segments were visualized by silver
* Corresponding author. Mailing address: C.M.B.C., IVIC, Apdo. 21827, Caracas 1020-A, Venezuela. Phone: (58 2) 501 1377. Fax: (58 2) 501 1382. Electronic mail address:
[email protected].
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NOTES
TABLE 1. Antigenic characterization of equine strains FR4 and FR8 by cross-neutralization of standard serotype G strains Virus (serotype)
Wa (Gl)
DS-1 (G2) RRV (G3) Gott (G4) OSU (G5) NCDV (G6) 69M (G8) W161 (G9) B223 (G1O) A253 (Gil) L338 (G13) F123 (G14) FR4
FR5 FR8
Titer' with antiserum to strain: Wa
6,400
