4 DISCUSSION
RVA is still the most common cause of non-bacterial AGE in children in
China27,28. The current study provides useful data in
the context of RVA infection and strain diversity in children ≤16 years
of age in Shanghai, highlighting the importance of routine surveillance
for the changing predominant strains to guide relevant vaccination
activities.
In this study, we closely monitored the trend of RVA infection in
Shanghai from May 2020 to December 2022. According to the available
data, RVA was still an important pathogen inducing AGE in Shanghai, with
the average detection rate of RVA being 16.6% during the study period.
The prevalence of RVA infection in children with AGE was similar to the
data from our previous study during 2012 and 2018 (13.6%)22. All these data suggested that the burden of AGE
attributed to RVA among outpatient children in Shanghai did not change
much during these years. Although LLR vaccine has been available in the
private market since 2000, vaccination coverage among children in
Shanghai is still low 29. Considering that the
children enrolled in this study were outpatients with mild symptoms and
the LLR vaccine showed high efficacy against severe and hospitalized AGE20,30,31, therefore, immune protection induced by RVA
vaccine may not yet be demonstrated in outpatients with AGE. In China,
recent large-scale national surveillance data showed that the prevalence
of RVA was 40.7%, 31.3%, and 11.2% in outpatient children in
lower-middle-income, upper-middle-income, and high-income regions,
respectively 32,33. The inclusion of appropriate RVA
vaccines in the non-pharmaceutical interventions (NPIs) should be
seriously considered as the burden of RVA-related AGE in children in
China is still serious.
Interestingly, the detection rate of RVA in 2021 (20.3%) was
significantly higher than that in 2020 (9.7%) and 2022 (14.5%). As we
all know, the coronavirus disease 2019 (COVID-19) epidemic spread
rapidly in China from the end of 2019, and the first COVID-19 case was
reported in Shanghai on January 20, 2020. Since then, different level of
NPIs (wearing masks, cordoning off, closing schools and day-care
centers, ensuring physical distance from others and washing hands) were
implemented in Shanghai until December 2022. Some studies also found
that the NPIs have a significant impact on the spread of many infectious
diseases, including gastroenteritis viruses 34-36.
This significant reduction in RVA infection observed in Shanghai in 2020
and 2022 maybe influenced by the strict implementation of the NPI
strategy. In contrast, the rebounded detection rate of RVA in 2021 may
be related to the relaxed implementation of the NPIs strategy because of
the low prevalence of COVID-19 in Shanghai during this period.
The temporal distribution of RVA infection among children with AGE in
Shanghai has a clear seasonal pattern, mainly concentrated in autumn and
winter, which was similar to the seasonal transmission pattern of RVA in
most regions of China 22. However, the analysis of the
monthly positive rate did not show a clear seasonal variation of RVA
infection influenced by the COVID-19 epidemic during the study period. A
relatively high RVA incidence was mainly concentrated in autumn in 2020,
winter and spring in 2020-2021 and 2021-2022. In addition, we also found
that the prevalence of RVA was at a low level in the winter of 2022.
However, in Japan, the peak infection of RVA has gradually shifted from
winter to spring since 1981, which may be influenced by warm weather37. Hence, continuous monitoring of the seasonal
epidemiologic characteristics of RVA in Shanghai in the future is
extremely important to identify the true cause of seasonal changes in
RVA prevalence. Similar to studies in different regions in China, the
prevalence of RVA in Shanghai was mainly concentrated in children aged
from 0-36 months 22. Meanwhile, we found that children
younger than 72 months and older children from 73 months to 16 years had
similar detection rates. We suspected this phenomenon may be related to
the decline in antibodies to RVA with age, which has been demonstrated
by several investigators 38-40.
In this study, the most common G genotypes of RVA in worldwide are G1,
G2, G3, G4, G9, and G12 8. Previous studies of
children infected with RVA in China showed that G3 and G1 were the most
common G genotypes before 2010, and G9 became a robust common G genotype
in children with acute diarrhea in China since 201141. Although G9 was still the most common G genotype
among the seven definite G genotypes, the proportion of this genotype
showed a significant decreasing trend from 2020 to 2022 and changed to
the second common G genotype in 2022. Surprisingly, G8 first appeared in
2020, but it eventually replaced G9 to become the most common genotype
in children with AGE in 2022. G8 is one of the more common RVA strains
of bovine origin and was first detected in humans in Indonesia between
1979 and 1981 42,43. Although it has been detected in
children in many countries and has become one of the dominant strains in
some sub-Saharan African countries, it was rare in China21,44. However, data from a study in Guangzhou showed
that a high proportion of infants with severe AGE were infected with G8
RVA recently 41. Altogether, the data indicate that G8
will be likely replace G9 as the predominant G genotype among
RVA-infected children with AGE in China. This hypothesis needs to be
supported by more comprehensive epidemiological data on RVA in different
regions of China.
Compared with the prevalent G genotypes in this study, only four P
genotypes (P[4], P[6], P[8], and P[10]) were detected.
Consistent with many other studies worldwide in recent years, P[8]
(97.2%) was the absolute most prevalent genotype in Shanghai from 2020
to 2022 8. The other three P genotypes were prevalent
at low levels and were only detected in 2021.
In this study, 13 RVA genotypes were identified in children with AGE.
Differences in the predominance of RVA strains and emerging strains were
observed. The circulation of G9P[8] strain from 2020 to 2021 was
overall predominant and similar to the rest of the world and our
previous data 22. Interestingly, the G8P[8]
unexpectedly became the most prevalent in 2022 with the proportion of
68.8%, which emerged in 2020. To our knowledge, this is the first time
that an epidemic of G8P[8] was found in Shanghai as early as 202022,41. Given that two licensed RVA vaccines have been
introduced in Shanghai, whether this change is due to the natural immune
pressure mechanisms or the RVA vaccination pressure is still unknown45. While universal RVA vaccination has progressed
worldwide, its impact on the emergence of escape mutants or even the
more efficient spread of previously known unusual strains due to
selective pressure is still a concern 37,46,47. Some
studies have reported a shift in the proportion of dominant RVA
genotypes before and after the introduction of the RVA vaccine37,46,48. However, continuous monitoring of RVA
genotypes in the pre- and post-vaccine era is essential to refine future
vaccine strategy and to understand the true nature of the immune
pressure exerted by RVA vaccines against other circulating wild-type
strains. Considering that the vaccine efficacy of LLR and RV5 against
AGE caused by G8P[8] is still unclear in China, the efficacy and
immunogenicity of these vaccines in Chinese children were urgently
needed to be clarified.
The circulation of G3P[8], which were the main epidemic strains in
Shanghai before 2011, showed a significant decrease during this study22. In addition, some other rare RVA genotypes were
identified during this study, including G1P[8], G1P[10],
G2P[8], G4P[8], G6P[8], G9P[4], and G9P[6] strains.
Although 7.0% of RVA samples were inconclusive for G or P genotypes,
this was much lower than our previous study (nearly 30.0%)22. We have improved the detection method to identify
G or P genotype in this study, which may be the main reason. Data from
this study imply that appropriate monitoring methods for RVA are
essential to find relatively comprehensive prevalent genotypes.
Additionally, phylogenetic analysis revealed that VP7 nucleotide
sequences of G8 strains detected in the study clustered together with
DS-8-like G8 strains isolated from Japan, Singapore, Thailand, and Korea
and shared high nucleotide sequence identity. We also found that the
Shanghai strains and bovine G8 strains shared 88.5%-96.8% aa sequence
identity with each other. Although some research had confirmed that the
G8P[8] strains in Southeast Asia were generated by reassortment of
bovine G8 strains and human DS-1-like strains occurred between 2007 and
2012 49, considering that RVA has 11 gene fragments
and we only analyzed the VP7 nucleotide sequence of G8 in this study, we
cannot yet conclude that the G8P[8] strains originated from
reassortment events between bovine and human strains. Therefore, more
and detailed whole genome sequence analysis is needed to clarify the
origin of the G8P[8] strain prevalent in Shanghai.
We also analyzed the distribution of RVA genotypes among children in
different age groups. We found that the greatest diversity of RVA
genotypes was found in children aged 0-72 months. This phenomenon
suggests that children aged 0-72 months are susceptible to multiple
genotypes of RVA because the immune response to RVA is not well
developed in this age group. Notably, G8P[8] was significantly more
common in children aged 73 months to 16 years than in children aged 0 to
72 months. This may also be one of the reasons for the higher detection
rate of RVA infection in this age group that we found in this study. Our
research suggested that there may be a difference in the age
distribution of G8P[8] infection, with prevalence mainly in older
age groups. However, this phenomenon still needs to be clarified
according to much more detailed epidemiologic data on RVA in people of
different ages.
In conclusion, the present study demonstrated the rich diversity of RVA
strains circulating in Shanghai. We reported for the first time that the
G8P[8] strain emerged in 2020, replacing G9P[8] as the
predominant cause of AGE in children with AGE in Shanghai in 2022.
Furthermore, the G8P[8] strain was more common in older children
than in younger children. Therefore, it is important to continue
monitoring RVA genotypes to inform future vaccine strategy to maintain
the success and effectiveness of the RVA vaccination program in
Shanghai.