Molecular characteristics of norovirus in sporadic and outbreak cases of acute gastroenteritis and in sewage in Sichuan, China - Virology Journal - Virology Journal

Norovirus is highly diverse and multiple genotypes are co-circulating worldwide to cause AGE. However, the distribution of norovirus genotypes varies by region, age, season, and setting. In this study, we investigated the molecular characteristics of norovirus circulating in Sichuan by analyzing surveillance data from multiple years of norovirus positive sporadic and outbreak specimens. A total of 11 distinct genotypes were detected. Among them, GII.4 and GII.3 were the most common genotypes in sporadic infections while GII.2 and GII.17 were most common in outbreak infections. These findings revealed that the dominant genotypes were different between sporadic cases and outbreak cases. In addition, these data suggested that vaccines for preventing norovirus gastroenteritis should protect against these four genotypes at minimum to induce broad protective immunity.

GII.4 Sydney 2012 norovirus has been circulating globally since 2012. From 2014 to 2015, non-GII.4 strains, such as GII.17 and GII.2, were prevalent in Asian regions. However, from 2015 to 2019, GII.4 Sydney 2012 remained as the predominant genotype in Sichuan. As shown in our previous report, norovirus had the highest detection rate in 2018 [10]. There may be three reasons that explain this trend. First, the number of norovirus genotypes was the highest in 2018. The increase in gene diversity may cause an increase in incidence. Second, GII.4 Sydney 2012 was the predominant genotype in 2018, indicating that this virus may be the main genotype leading to this increase in incidence. Third, the nucleotide homology of GII.4 norovirus was lower than other genotypes circulating in the same time period. GII.4 strains collected in 2018 had the lowest nucleotide homology, indicating that GII.4 was more variable in 2018, which may have also led to an increase in incidence. Our previous study showed that autumn was the predominant season of norovirus gastroenteritis in Sichuan [10]. In the current study, GII.4 norovirus was responsible for the majority of norovirus infections in the autumn, further confirming that it continues to be the predominant genotype in Sichuan.

Our previous study indicated that norovirus was more likely to infect children 0–12 months of age than rotavirus in Sichuan [10]. The current study further demonstrated that children 0–6 months of age were primarily infected by GII.3 while children older than 6 months were mainly infected by GII.4. Similar results were reported from Novosibirsk, Russia where GII.3-positive children were younger than GII.4-positive children [11]. These findings revealed that the median age of susceptible children was different depending on the norovirus genotype. In conclusion, in addition to GII.4, GII.3 should be considered for the development of vaccines for infants and young children.

In late 2014, an emergent variant of a previously rare norovirus genotype, GII.17, became dominated in Asian regions, including mainland China [12,13,14]. GII.17 was detected in 2015 in sporadic AGE cases but not subsequently identified in 2016 or 2017 in Sichuan. The sequences of all strains detected from 2015 to 2019 were highly homologous. However, GII.17 was the second etiology responsible for norovirus outbreaks. It was reported that GII.17 was most prevalent in adult patients and these patients were significantly older than those patients infected with GII.4 [13]. Our current study also showed that GII.17 was not detected in children 0–12 months of age while the detection rate increased with age in children older than 13 months. In addition, five out of seven GII.17 outbreaks occurred in hospitals, companies, building sites and communities. These locations were typically occupied by adults, supporting that GII.17 was a genotype that primarily infects adults.

In the winter of 2016–2017, GII.2 [P16] replaced GII.17 as the predominate genotype in China and accounted for 81.2% of norovirus outbreaks from 2016 to 2018 [15,16,17,18]. Interestingly, GII.2 [P16] was also the predominant genotype detected in sewage in Sichuan. However, GII.2 [P16] was rarely detected in sporadic norovirus cases. It was reported that GII.2 [P16] resulted in a higher viral load than GII.17 and GII.4 [19]. In the current study, none of the genotypes detected were found to have a higher viral load.

Surveillance of norovirus was largely based on clinical cases of AGE in sentinel hospitals in China and there was only one sentinel hospital in Sichuan. This hospital-based surveillance system was not adequate to fully evaluate the epidemiology of norovirus. First, this system primarily covered populations under five years of age. Second, cases included in this surveillance system were patients with severe AGE symptoms who sought medical care. Mild or asymptomatic norovirus infections remained unknown. Also, detection rates of some norovirus genotypes were low in clinical cases. Therefore, it is crucial to improve the current surveillance system to obtain comprehensive epidemiological data on norovirus.

Environmental sewage surveillance systems have been established in some developed countries to serve as an effective supplement to the current clinical case surveillance [20,21,22,23]. These new systems could yield epidemiological data of norovirus on the view of whole population. And a variety of norovirus genotypes, including genotypes not commonly found in clinical cases, were detected by sewage surveillance. Besides, establishing of sewage surveillance could not only provide information about norovirus disease but also information about other contagious diseases, especially some emerging or re-merging diseases. This surveillance system will likely be more effective and more economic than case surveillance after taking full advantage of these data. Thus, this study set up environmental sewage surveillance system and described the molecular characteristics of norovirus in sewage in Sichuan. Using amplicon sequencing, 15 norovirus genotypes were identified, of which eight genotypes had not been detected in clinical cases in the same period. Notably, GII.3 was secondary to GII.4 in sporadic AGE cases while the relative abundance of GII.3 in sewage had exceeded GII.4 in Sichuan. In China from 2015 to 2016, the number of GII.3 norovirus outbreaks increased compared with previous years [24]. In Beijing in 2017, GII.3 was found to have a higher detection rate than GII.4 in hospitalized children with AGE [25]. To determine whether the detection rate of GII.3 would exceed that of GII.4 in clinical cases, long-time surveillance is need, which should be a focus of future monitoring work. A slight majority (50.37%) of norovirus sequences found in sewage were typed as genogroup I. However, in a five-year period, only two sporadic norovirus cases were typed as GI. This may suggest that the current surveillance system was not sensitive enough to detect GI infections or GI noroviruses result more frequently in asymptomatic infections.