1987;8:221C228

1987;8:221C228

1987;8:221C228. P typing was carried out by nested amplification of variable sequences of the VP7 and the VP4 genes with six G- and five P-type-specific primers (multiplex PCR). Results obtained by this method showed the prevalence of the following G and P types: G1, 39%; Emtricitabine G2, 43%; G4, 4%; P[8], 16%; P[4], 71%. Unexpectedly, the G-P type combination most frequently found was G2P[4] (43%) rather than G1P[8] (12%), which is the most commonly found worldwide. Unusual strains of the type G1P[4] accounted for 14% of the total, while mixed infections with more Emtricitabine than one type were found in 10% of the samples. Detection of fecal rotavirus-specific immunoglobulin M (IgM) and IgA antibodies in consecutive samples of two patients taken at daily intervals exhibited that high levels of IgM and IgA antibodies were detected on day 1 after the onset of disease and that the samples remained positive for about 10 days, after which virus shedding was no longer observed. Multiplex PCR offers a sensitive and specific alternative to determine the prevalence of group A rotavirus G and P types and to identify the emergence of uncommon strains, whereas detection of fecal IgM and IgA antibodies represents a useful supplement to virus detection for the diagnosis of current or recently acquired infections. Rotaviruses have been recognized as the major etiologic brokers of acute gastroenteritis in infants and young children worldwide (14, 20, 35). Rotavirus serotypes are specified by two outer capsid proteins, VP4 and VP7, encoded by different genome segments (13, 32). VP7 and VP4 proteins elicit, independently, neutralizing antibodies and specify the virus G (outer shell glycoprotein) and P (for protease-susceptible protein) serotypes, respectively. VP4, the product of gene 4, is the viral hemagglutinin and appears to be responsible for restriction of growth in tissue culture and virulence in experimental animals (50). Proteolytic cleavage of this protein enhances rotavirus infectivity (41). Rotavirus serotypes have been established on the basis of a 20-fold or higher difference in reciprocal neutralization titers with hyperimmune homologous and heterologous antisera (57C59). Because the genes encoding these proteins segregate independently of each other during reassortment, a dual-serotyping system to account for the specificities of both VP7 and VP4 has been adopted (32, 44). On the basis of the VP7 protein, 14 different G types have been described so far; among these, 10 serotypes were associated with acute gastroenteritis in humans (31). Four of these rotavirus serotypes (G1 to G4) are the most common etiologic brokers of childhood diarrhea worldwide for which vaccines have been developed (36, 37). Typing of human group A rotavirus by molecular and immunological methods has been reported (6, 15, 18, 26, 27, 40, 57). P serotypes have been defined by Gorziglia et al. (25) by using polyclonal antibodies to baculovirus-expressed VP4 protein. They showed that rotavirus serotype P[8] with G1, G3, and G4 specificities (prototype strains Wa, Ku, P, and VA70) was present in isolates from children with acute diarrhea whereas type P[4] combined with virulent G2 (DS-1-like strain) and P[6] with G1 to G4 specificities (prototype strains M37, 1076, McN13, and STE) were isolated from asymptomatic newborns excreting rotavirus. These strains were classified into three genetic and three Emtricitabine antigenic types, designated P1A, P1B, and P2, respectively. Since VP4 is usually a minor outer protein with only 250 copies of the molecule per viral particle, monoclonal antibodies to this protein are rather difficult to obtain (5) for the average laboratory. In addition, preparation of the necessary reagents is usually laborious and time-consuming (21, 23). To overcome these problems, the typing of rotavirus P strains can be Rabbit Polyclonal to SPTA2 (Cleaved-Asp1185) accomplished by identification of genetically different VP4 genes by reverse transcription-PCR (RT-PCR), as previously reported (9, 17, 33, 52, 53). Analysis of prevalent VP7 and VP4 genes is usually important for evaluating candidate rotavirus vaccines. The prevalence of G types in Argentine children infected with group A rotavirus was previously assessed with monoclonal antibodies to types G1 to G4 by an enzyme-linked immunosorbent assay (ELISA) (24). Many studies using VP4 (P) genotyping methods demonstrated a worldwide combination of one P genotype, P[8], with G1, G3, and G4, whereas P[4] was frequently associated with G2 (16, 17, 52). Studies carried out in India revealed the prevalence of a different G-P combination: genotype P[6] frequently associated with an unusual G serotype G9.