Genetic diversity of human sapovirus across the Americas

Abstract

Background: Sapoviruses are responsible for sporadic and epidemic acute gastroenteritis worldwide. Sapovirus typing protocols have a success rate as low as 43% and relatively few complete sapovirus genome sequences are available to improve current typing protocols.

Objective/study design: To increase the number of complete sapovirus genomes to better understand the molecular epidemiology of human sapovirus and to improve the success rate of current sapovirus typing methods, we used deep metagenomics shotgun sequencing to obtain the complete genomes of 68 sapovirus samples from four different countries across the Americas (Guatemala, Nicaragua, Peru and the US).

Results: VP1 genotyping showed that all sapovirus sequences could be grouped in the four established genogroups (GI (n = 13), GII (n = 30), GIV (n = 23), GV (n = 2)) that infect humans. They include the near-complete genome of a GI.6 virus and a recently reported novel GII.8 virus. Sequences of the complete RNA-dependent RNA polymerase gene could be grouped into three major genetic clusters or polymerase (P) types (GI.P, GII.P and GV.P) with all GIV viruses harboring a GII polymerase. One (GII.P-GII.4) of the new 68 sequences was a recombinant virus with the hotspot between the NS7 and VP1 regions.

Conclusions: Analyses of this expanded database of near-complete sapovirus sequences showed several mismatches in the genotyping primers, suggesting opportunities to revisit and update current sapovirus typing methods.

Keywords: Genotypes; Next generation sequencing; Sapovirus; Viral gastroenteritis.

Fig. 1.

Pairwise sequence comparison of novel sapovirus strains (GII.8 Monterey6283, GII.8 Monterey6284, GII.8 Lima1859, and GII.8 Lima1867) with sapovirus GII.1-GII.7reference sequences. In the color-coded pairwise identity matrix each cell includes the percentage identity among two sequences (horizontally to the left and vertically at the bottom). The colored key indicates the correspondence between pairwise identities and the colors displayed in the matrix.

Fig. 2.

Phylogenetic analysis of 99 sapovirus RdRp (A) and 70 VP1 (B) sequences using Maximum Likelihood method. The tree was inferred by using the Maximum Likelihood method based on the Tamura-Nei model. The trees with the highest log likelihood are shown, on the basis of nucleotide sequence of the complete RdRp (A) and VP1 (B) genes. The trees are drawn to scale with branch lengths measured in the number of substitutions per site. Bootstrap values (100 replicates) are shown next to each branch. Branches have been compressed for clarity and only the GII branch is partially expanded to show how the four samples of GII.8 branch out from the closest GII.7 sapovirus.

Fig. 3.

Evidence of recombination for sapovirus GII.4 Lima1873 compared to previous sequenced GII.1 and GII.4 genomes. Sapovirus genomes were analyzed by SimPlot and similarity scores using a 200 nt sliding window are plotted. Three reference strains (A, B and C) were analyzed against the query sequence GII.4_Lima1873. Similarity scores of all three references (two GII.1 and one GII.4) were almost > 85% in the entire NS region, with a notable drop in similarity scores over the capsid region and ORF2 (VP2). The query sequence most closely related to previously described sapovirus recombinant strain with a GII.P polymerase and GII.4 capsid (KP067444). Red boxes indicate the approximate location of the hemi nested genotyping primers [29].

Fig. 4.

Alignment of 68 near complete genome sequences obtained in this study and the primers used for hemi-nested PCR [29]. Mismatches between individual strains and the primer binding regions are highlighted in yellow. ^aNucleotide positions based on Manchester strain (Accesion number X86560).