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. 2012 Apr 27;30 Suppl 1(0 1):A140-51.
doi: 10.1016/j.vaccine.2011.09.119.

Molecular characterization of rotavirus strains detected during a clinical trial of a human rotavirus vaccine in Blantyre, Malawi

Toyoko Nakagomi  1 , Osamu Nakagomi, Winifred Dove, Yen Hai Doan, Desiree Witte, Bagrey Ngwira, Stacy Todd, A Duncan Steele, Kathleen M Neuzil, Nigel A Cunliffe
Affiliations

Molecular characterization of rotavirus strains detected during a clinical trial of a human rotavirus vaccine in Blantyre, Malawi

Toyoko Nakagomi et al. Vaccine. .

Abstract

The human, G1P[8] rotavirus vaccine (Rotarix™) significantly reduced severe rotavirus gastroenteritis episodes in a clinical trial in South Africa and Malawi, but vaccine efficacy was lower in Malawi (49.5%) than reported in South Africa (76.9%) and elsewhere. The aim of this study was to examine the molecular relationships of circulating wild-type rotaviruses detected during the clinical trial in Malawi to RIX4414 (the strain contained in Rotarix™) and to common human rotavirus strains. Of 88 rotavirus-positive, diarrhoeal stool specimens, 43 rotaviruses exhibited identifiable RNA migration patterns when examined by polyacrylamide gel electrophoresis. The genes encoding VP7, VP4, VP6 and NSP4 of 5 representative strains possessing genotypes G12P[6], G1P[8], G9P[8], and G8P[4] were sequenced. While their VP7 (G) and VP4 (P) genotype designations were confirmed, the VP6 (I) and NSP4 (E) genotypes were either I1E1 or I2E2, indicating that they were of human rotavirus origin. RNA-RNA hybridization using 21 culture-adapted strains showed that Malawian rotaviruses had a genomic RNA constellation common to either the Wa-like or the DS-1 like human rotaviruses. Overall, the Malawi strains appear similar in their genetic make-up to rotaviruses described in countries where vaccine efficacy is greater, suggesting that the lower efficacy in Malawi is unlikely to be explained by the diversity of circulating strains.

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Figures

Figure 1
Figure 1
Electropherotypes on 10% polyacrylamide gels of cell-culture adapted rotavirus strains used in this study. The control RNAs were obtained from strains Wa (G1P[8]), KUN (G2P[4]), and RIX4414 (G1P[8]). Gels were stained with ethidium bromide and illuminated under UV light. The name of the strain is indicated on the top of each lane. G8P[4] (MAL01, MAL02, MAL33, MAL47, MAL55, MAL60, MAL70, and MAL81); G8P[6] (MAL43); G12P[6] (2 short pattern viruses MAL39 and MAL88, and 2 long RNA pattern viruses MAL12 and MAL40); G9P[8] (MAL80 and MAL82); G1P[8] (MAL23, MAL38, and MAL50); G1P[6] (MAL63); G12P[8] (MAL65); and G2P[4] (MAL66).
Figure 2
Figure 2
Hybridization of RIX4414 probe with genomic RNAs from various Malawian rotavirus strains. 1: Wa; 2: RIX4414; 3: RIX4414; 4: MAL23, G1P[8]; 5: MAL38, G1P[8]; 6: KUN, G2P[4]; 7: MAL80, G9P[8]; 8: MAL12, G12P[6]; 9: RIX4414; 10: MAL88, G12P[6]; 11: MAL43, G8P[6]; 12: MAL60, G8P[4]; 13: MAL70, G8P[4]; 14: MAL66, G2P[4]. Top panel: ethidium bromide staining followed by ultraviolet illumination. Bottom panel: corresponding autoradiograph.
Figure 3
Figure 3
Hybridization of MAL60 probe (G8P[4]) with genomic RNAs from various Malawian rotavirus strains. 1: Wa; 2: RIX4414; 3: MAL23, G1P[8]; 4: MAL38, G1P[8]; 5: MAL50, G1P[8]; 6: MAL63, G1P[6]; 7: MAL82, G9P[8]; 8: MAL65, G12P[8]; 9: MAL40, G12P[6]; 10: MAL88, G12P[6]; 11: MAL43, G8P[6]; 12: MAL60, G8P[4]; 13: MAL70, G8P[4]; 14: MAL66, G2P[4]. 15: KUN, G2P[4]. Top panel: ethidium bromide staining followed by ultraviolet illumination. Bottom panel: corresponding autoradiograph.
Figure 4
Figure 4
Hybridization of MAL88 probe (G12P[6], short RNA pattern]) with genomic RNAs from various Malawian rotavirus strains. 1: MAL88, G12P[6]; 2: MAL39, G12P[6]; 3: MAL88, G12P[6]; 4: MAL12, G12P[6]; 5: MAL40, G12P[6]; 6: MAL65, G12P[8]. Top panel: ethidium bromide staining followed by ultraviolet illumination. Bottom panel: corresponding autoradiograph.
Figure 5
Figure 5
Phylogenetic trees for the G1 VP7 genes (a), the G8 VP7 genes (b), the G9 VP7 genes (c), and the G12 VP7 genes (d). The phylogenetic trees were constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the VP7 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591403-7 respectively.
Figure 5
Figure 5
Phylogenetic trees for the G1 VP7 genes (a), the G8 VP7 genes (b), the G9 VP7 genes (c), and the G12 VP7 genes (d). The phylogenetic trees were constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the VP7 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591403-7 respectively.
Figure 5
Figure 5
Phylogenetic trees for the G1 VP7 genes (a), the G8 VP7 genes (b), the G9 VP7 genes (c), and the G12 VP7 genes (d). The phylogenetic trees were constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the VP7 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591403-7 respectively.
Figure 5
Figure 5
Phylogenetic trees for the G1 VP7 genes (a), the G8 VP7 genes (b), the G9 VP7 genes (c), and the G12 VP7 genes (d). The phylogenetic trees were constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the VP7 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591403-7 respectively.
Figure 6
Figure 6
Phylogenetic trees for the P[8] VP4 genes (a), the P[6] VP4 genes, and the P[4] VP4 genes (c) . The phylogenetic trees were constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the VP4 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591398-402 respectively.
Figure 6
Figure 6
Phylogenetic trees for the P[8] VP4 genes (a), the P[6] VP4 genes, and the P[4] VP4 genes (c) . The phylogenetic trees were constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the VP4 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591398-402 respectively.
Figure 6
Figure 6
Phylogenetic trees for the P[8] VP4 genes (a), the P[6] VP4 genes, and the P[4] VP4 genes (c) . The phylogenetic trees were constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the VP4 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591398-402 respectively.
Figure 7
Figure 7
Phylogenetic tree for the VP6 genes. The phylogenetic tree was constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the VP6 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591408-12 respectively.
Figure 8
Figure 8
Phylogenetic tree for the NSP4 genes. The phylogenetic tree was constructed using the neighbour-joining method and Kimura’s 2-parameter model. Percentage bootstrap values are given at branch nodes except values <70 which are not shown. The nucleotide sequences of the NSP4 genes of the Malawi strains used in this study have been deposited in GenBank as follows: MAL12, MAL23, MAL81, MAL82, MAL88 have been assigned accession numbers JN591393-7 respectively.
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