The emergence and evolution of the novel epidemic norovirus GII.4 variant Sydney 2012

Abstract

Norovirus is the leading cause of acute gastroenteritis with most infections caused by GII.4 variants. To understand the evolutionary processes that contribute to the emergence of GII.4 variants, we examined the molecular epidemiology of norovirus-associated acute gastroenteritis in Australia and New Zealand from 893 outbreaks between 2009 and 2012. Throughout the study GII.4 New Orleans 2009 was predominant; however, during 2012 it was replaced by an emergent GII.4 variant, Sydney 2012. An evolutionary analysis of capsid gene sequences was performed to determine the origins and selective pressures driving the emergence of these recently circulating GII.4 variants. This revealed that both New Orleans 2009 and Sydney 2012 share a common ancestor with GII.4 Apeldoorn 2007. Furthermore, pre-epidemic ancestral variants of each virus were identified up to two years before their pandemic emergence. Adaptive changes at known blockade epitopes in the viral capsid were also identified that likely contributed to their emergence.

Keywords: Emergence; Evolution; Gastroenteritis; Norovirus; Sydney 2012.

Fig. 1. Laboratory confirmed number of monthly norovirus outbreaks in Australia and New Zealand between 2009 and 2012.

The number of laboratory confirmed norovirus outbreaks per month is shown between January 2009 and December 2012 for New South Wales, Australia (A) and New Zealand (B). Annual peaks in activity were observed during the late winter period in Australia (A) and the late spring for New Zealand (B), except in 2011 where the peak in activity occurred during autumn.

Fig. 2. Prevalence of circulating norovirus genotypes in the Oceania region.

The genotyping results for Australia and New Zealand were combined then plotted by month between 2009 and 2012 to highlight the cause of each norovirus epidemic as well as the shift in prevalence between New Orleans 2009 and Sydney 2012 that occurred during late 2012. The prevalence of each GII.4 variants is shown with non-GII.4s grouped separately and coloured according to the key provided.

Fig. 3. Temporal evolutionary analysis of the GII.4 Apeldoorn-lineage capsid.

(A) Maximum likelihood phylogeny derived from capsid protruding domain sequences (n=84) showing the major GII.4 variants identified since 1996 (with a Camberwell 1994 outgroup). The Apeldoorn-lineage forms a distinct group of variants that included Apeldoorn 2007, New Orleans 2009 and Sydney 2012, highlighted green, red and blue, respectively. The scale bar indicates the number of nucleotide substitutions per site. (B) Bayesian Maximum Clade Credibility phylogeny of the Apeldoorn-lineage from 286 complete capsid sequences, of which 221 were derived from this study. The posterior probabilities for key nodes are shown for those >0.80. The node bars show the 95% highest probability densities interval for node ages. Clades are coloured as panel A with the pre-epidemic variants Orange 2008 and Auckland 2010 shown in lighter shades of red and blue, respectively. The x-axis is scaled to time (years) with individual tips representing time points of sample collection. Branches marked with an asterisk and arrows indicate that the sites listed are under significant positive selection specifically along these branches and match those shown in Fig. 4.

Fig. 4. Antigenic variation in the GII.4 Apeldoorn-lineage.

The GII.4 capsid sequence alignment of variants in the Apeldoorn-lineage used for the temporal evolutionary analysis was examined for antigenic clusters within and between different GII.4 variants as well as for evidence of positive selection. Asterisks indicate sites – 294, 368, 373 and 376 – that are under significant positive selection. Labelled boxes above each position indicate sites within the known blockade epitopes A–E that are important determinants of viral antigenicity. Amino acids have been coloured based on the properties of their side-chains: blue for positive charged – R, H, K; red for negative charged – D, E; Green for polar uncharged – S, T, N, Q; yellow for hydrophobic – A, V, I, L, M, F, Y, W; pink for special cases – C, U, G, P.