Emerging recombinant noroviruses identified by clinical and waste water screening

Abstract

Norovirus is estimated to cause 677 million annual cases of gastroenteritis worldwide, resulting in 210,000 deaths. As viral gastroenteritis is generally self-limiting, clinical samples for epidemiological studies only partially represent circulating noroviruses in the population and is biased towards severe symptomatic cases. As infected individuals from both symptomatic and asymptomatic cases shed viruses into the sewerage system at a high concentration, waste water samples are useful for the molecular epidemiological analysis of norovirus genotypes at a population level. Using Illumina MiSeq and Sanger sequencing, we surveyed circulating norovirus within Australia and New Zealand, from July 2014 to December 2016. Importantly, norovirus genomic diversity during 2016 was compared between clinical and waste water samples to identify potential pandemic variants, novel recombinant viruses and the timing of their emergence. Although the GII.4 Sydney 2012 variant was prominent in 2014 and 2015, its prevalence significantly decreased in both clinical and waste water samples over 2016. This was concomitant with the emergence of multiple norovirus strains, including two GII.4 Sydney 2012 recombinant viruses, GII.P4 New Orleans 2009/GII.4 Sydney 2012 and GII.P16/GII.4 Sydney 2012, along with three other emerging strains GII.17, GII.P12/GII.3 and GII.P16/GII.2. This is unusual, as a single GII.4 pandemic variant is generally responsible for 65–80% of all human norovirus infections at any one time and predominates until it is replaced by a new pandemic variant. In sumary, this study demonstrates the combined use of clinical and wastewater samples provides a more complete picture of norovirus circulating within the population.

Recombinant GII.P16/GII.4 Sydney 2012 Was the Dominant Norovirus Identified in Australia and New Zealand in 2017

“…Stool suspensions (10–20% in water) were prepared from clinical specimens, followed by viral RNA extraction, as described previously [ 13 , 29 ]. Ultracentrifugation was used to concentrate viruses in wastewater samples prior to viral RNA extraction, as described in [ 16 ]. Frozen aliquots of MS2 bacteriophage, with a concentration of 2.6 × 10 6 ± 1.6 × 10 5 genome copies/20 μL, were used as process control to validate RNA extraction and RT-PCR amplification [ 16 ].…”

“…However, a change in norovirus molecular epidemiology has been observed over the past three years. In 2016, we saw a decline of the pandemic Sydney 2012 variant, concomitant with the emergence of two novel GII.4 recombinant viruses, both of which retained the Sydney 2012 capsid but acquired new non-structural regions (GII.P4 New Orleans 2009/GII.4 Sydney 2012 and GII.P16/GII.4 Sydney 2012) [ 16 ]. The emergence of GII.P4 New Orleans 2009/GII.4 Sydney 2012 was first detected in 2013, and was created through a recombination event between the Sydney 2012 pandemic variant with its pandemic predecessor GII.4 New Orleans 2009 variant [ 17 ].…”

“…The emergence of GII.P4 New Orleans 2009/GII.4 Sydney 2012 was first detected in 2013, and was created through a recombination event between the Sydney 2012 pandemic variant with its pandemic predecessor GII.4 New Orleans 2009 variant [ 17 ]. Following its identification, this recombinant virus was also identified in Australia, New Zealand [ 16 , 18 ], Denmark [ 19 ], England [ 20 ], and South Africa [ 21 ]. The other novel GII.4 recombinant, GII.P16/GII.4 Sydney 2012, was first identified in the Oceania region in mid-2015, and was found circulating at a low prevalence in South Korea, Germany, and Japan in 2016 [ 16 , 22 , 23 , 24 ].…”

“…Following its identification, this recombinant virus was also identified in Australia, New Zealand [ 16 , 18 ], Denmark [ 19 ], England [ 20 ], and South Africa [ 21 ]. The other novel GII.4 recombinant, GII.P16/GII.4 Sydney 2012, was first identified in the Oceania region in mid-2015, and was found circulating at a low prevalence in South Korea, Germany, and Japan in 2016 [ 16 , 22 , 23 , 24 ]. In mid-late 2016, an increase of a third novel recombinant, GII.P16/GII.2, was observed in Australia and New Zealand [ 16 ].…”

“…The other novel GII.4 recombinant, GII.P16/GII.4 Sydney 2012, was first identified in the Oceania region in mid-2015, and was found circulating at a low prevalence in South Korea, Germany, and Japan in 2016 [ 16 , 22 , 23 , 24 ]. In mid-late 2016, an increase of a third novel recombinant, GII.P16/GII.2, was observed in Australia and New Zealand [ 16 ]. This virus was also detected around the globe in 2016, including in Japan [ 25 ], the United States [ 26 ], China (51% of outbreaks) [ 27 ], and Europe [ 24 , 28 ], where it was responsible for 14%–42% of all norovirus outbreaks.…”