Rotavirus strain diversity in Blantyre, Malawi, from 1997 to 1999

Abstract

In a 2-year study of viral gastroenteritis in children in Blantyre, Malawi, the diversity of rotavirus strains was investigated by using electropherotyping, reverse transcription-PCR amplification of the VP7 and VP4 genes (G and P genotyping), and nucleotide sequencing. Of 414 rotavirus strains characterized, the following strain types were identified: P[8], G1 (n = 111; 26.8%); P[6], G8 (n = 110; 26.6%); P[8], G3 (n = 93; 22.5%); P[4], G8 (n = 31; 7.5%); P[8], G4 (n = 21; 5.1%); P[6], G3 (n = 12; 2.9%); P[6], G1 (n = 7; 1.7%); P[6], G9 (n = 3; 0.7%); P[6], G4 (n = 3; 0.7%); P[4], G3 (n = 1; 0.2%); and mixed (n = 15; 3.6%). While all strains could be assigned a G type, seven strains (1.7%) remained P nontypeable. The majority of serotype G8 strains and all serotype G9 strains had short electropherotype profiles. All remaining typeable strains had long electropherotypes. Divergent serotype G1 rotaviruses, which contained multiple base substitutions in the 9T-1 primer binding site, were commonly identified in the second year of surveillance. Serotype G2 was not identified. Overall, G8 was the most frequently identified VP7 serotype (n = 144; 34.8%) and P[8] was the most frequently detected VP4 genotype (n = 227; 54.8%). Partial sequence analysis of the VP4 gene of genotype P[8] rotaviruses identified three distinct clusters, which predominantly (but not exclusively) comprised strains belonging to a distinct VP7 serotype (G1, G3, or G4). As a result of mutations in the 1T-1 primer binding site, strains belonging to each cluster required a separate primer for efficient typing. One cluster, represented by P[8], G4 strain OP354, was highly divergent from the established Wa and F45 VP4 P[8] lineages. As is the case for some other countries, the diversity of rotaviruses in Malawi implies that rotavirus vaccines in development will need to protect against a wider panel of serotypes than originally envisioned.

FIG. 1

(a) Nucleotide changes in VP7 of Malawi serotype G1 strain MW529 in the 9T-1 region compared to prototype serotype G1 strain Wa. (b) Nucleotide changes in VP4 of Malawi P[8] strains OP601, MW258, and OP354 in the IT-1 region compared to prototype genotype P[8] strain KU.

FIG. 2

Temporal distribution of rotavirus strains in Blantyre, Malawi, from 1997 to 1999. A total of 221 strains were detected in year 1 of the study (1997 to 1998 [shaded bars]), and 193 strains were detected in year 2 (1998 to 1999 [black bars with white stippling]). NT, nontypeable.

FIG. 3

Neighbor-joining nucleotide distance tree for residues representing nucleotides 11 to 887 of the VP4 gene of the indicated genotype P[8] strains. The G type is indicated for each strain. Sequences were aligned by using CLUSTALX and analyzed by using the DNADIST and NEIGHBOR programs in PHYLIP. The accession numbers of the reference strains used in this analysis were L34161 (for Wa) (46), U30716 (for F45) (37), and M21014 (for KU) (57).

FIG. 4

Comparison of conserved amino acids between residues 27 and 240 of VP4 of OP354-like strains compared to Wa-like and F45-like strains. (The data for Wa-like and F45-like strains are from a report by Gouvea et al. [25].)