In the last few decades, HFMD has contributed to several large outbreaks and thousands of HFMD fatal cases worldwide, which has been a growing public health problem[20-22]. After the first report of HFMD in Shanghai in 1981, alternating epidemic of EV71 and CV-A16 occurred in the following years. HFMD poses a significant threat to public health in China and was classified as a C-class notifiable communicable disease in China in 2008. Our previous research revealed that EV71 and CV-A16 represented the two major epidemic enterovirus serotypes in Hangzhou between 2012 and 2013. However, the predominant types of enteroviruses have changed since 2013, and the positive rates of the EV positive HFMD cases that were both non-EV71 and non-CV-A16 have grown rapidly. The other EVs replaced EV71 and CV-A16 as the major serotype in Hangzhou between 2014 and 2015. This study continues to analyze the characteristics of HFMD epidemic that occurred in Hangzhou in 2016, 2017 and 2018.
In this study, we reported that other EVs had replaced EV71 as the primary viral serotype responsible for HFMD onset in Hangzhou between 2016 and 2018. Concurrently, the same trend was observed in other cities near Hangzhou in China[23,24]. The cases of EV71 decreased year by year in the past three years. And compared to 2016 and 2017, only 13 cases of EV71 were detected in 2018, which meant a significant decrease in the proportion. Therefore, 1297 cases randomly selected of the other EVs were tested in 2017 and 2018. There were 835 (64.4%) cases of CV-A6 infections and 177 (13.6%) cases of CV-A10 infections in 2017 and 2018. Less mixed infections was detected among different types in this study. The cases of CV-A6 infections also increased dramatically and CV-A6 became one of the main causative agents of HFMD in Hangzhou. CV-A10 also overtook CV-A16 to become the second most common viral serotype. Hangzhou is the capital city of Zhejiang province, and its high population mobility made it a HFMD-prone area and likely more conducive to virus faster evolution. Some epidemiologists suggest three primary factors were involved in the regionally increased CV-A6 infection rates and decreased EV71 infection rates. Firstly, after a long period of the EV71 epidemic, elevated antibody and thus immunity levels against EV71 in the population might impart a selection pressure for other EV serotypes to arise, because antibodies against EV71 and CV-A16 cannot protect the susceptible population from the other serotypes. Secondly, the popularity of EV71 vaccination may play a role in this change in recent years. Mass EV71 vaccination is not expected to substantially reduce the total number of HFMD cases because the vast majority of HFMD cases are mild and more than half of the mild cases were due to CV-A16 and other EV serotypes infections. However, most of severe and death HFMD cases could be prevented by EV-A71 vaccination. Thirdly, CV-A6 might have transformed into a more virulent strain to escape human immune protection. Yoshitomi H’s study showed that the phylogenetic analysis of the entire VP1 gene revealed the diversity of the prevalent CV-A6 between 2013 and 2017 in Fukuoka. The CV-A6 strains were classified into seven genetic clades (A–G) and subgroups of clade A (subclades A1–A4) based on the entire VP1 sequences.The phylogenetic analysis revealed that all the CV-A6 strains detected in Fukuoka between 2013 and 2017 were classified into clade A（A3 and A4）,which were different from Hong Kong(D4 and D5)[27,28]. The emergence of CV-A6 strains with high genetic diversity was believed to be a factor associated with the epidemics. More studies including whole genome sequencing of the epidemic strains are warranted to determine whether these variations affect the epidemic incidence of CV-A6. We think more attention should also be paid to the other EV serotypes, such as CV-A6 and CV-A10, in the future surveillance and control of HFMD in Hangzhou and even in Zhejiang Province. We speculate that the year of 2018 may become a turn point in the EV serotypes causing HFMD, and we should pay more attention to this phenomenon.
Additionally, Hangzhou shared similar seasonal epidemic patterns of EV-predominant serotypes with other southern cities in China. Our data showed two seasonal peaks in 2016 and 2017 in the total HFMD cases. Similar finding was reported in Shenzhen, Wuxi and Hong Kong[28-30]. In these three districts, the season alternations from spring to summer and from autumn to winter are relatively mild, which might be in favor of the spread of certain EV viral serotypes. This is consistent with the results of some studies that found the number of patients with HFMD is affected by environmental factors such as temperature, humidity, sunshine time and so on. In this study, we reported that other EV infection peaks mainly occurred during October to December in 2016 and 2017. Biao Di’s study also found CV-A6 infection mainly occurred during the autumn and winter, and the outbreak of CV-A6 infection might contribute to the second peak of the HFMD. In 2018, the main infection was other EV infection in Hangzhou, and there was no EV71 infection peak, which might also be the reason why the second peak was not obvious in 2018. CV-A16 displayed a low detection rate throughout the year.
In our study, we investigated whether the etiological spectrum of mild and severe HFMD changed. EV71 was still the leading serotype responsible for severe HFMD cases, while the other EV infection accounted for a increase in severe cases. Between 2016 and 2018, other EV infection cases accounted for 46.8% of EV-positive specimens in severe HFMD cases in Hangzhou. This proportion was much higher than the result of our study in the previous several years. This may be due to the change in predominant types of the prevalent EV serotype distributions throughout the year. In our study, the severe case rate of EV71 infections (69.2% of the total EV71-positive cases) was significantly higher than that of CV-A16 infections (13.7%, 34 of 249) and other EV infections (10.1%, 290 of 2884), which suggested that EV71 infected patients had still a greater chance of developing severe cases of HFMD than other EV serotypes infected patients. However, the pathogenic mechanism of EV71 severity and fatality remained unclear. Ya-Ping Li et al’s report suggested that the different genetic background of host innate immunity may lead to different clinical outcomes. They found that RIG-1 rs3739674 and RIG-1 rs9695310 polymorphisms are associated with EV71 HFMD risk. RIG-1 rs3739674 and TLR3 rs5743305 polymorphisms are associated with disease severity. Previous reports showed that the levels of Serum interleukin-6, B-type natriuretic peptide, hyperglycemia and leukocytosis in children with severe neurological symptoms caused by EV71 infection significantly increased, causing a systemic inflammatory factor storm and leading to heart and lung failure, which might indicate a strong relationship between proinflammatory cytokines and severity in EV71 infection[34,35].
Statistical analysis of the gender distribution of all cases in the present study showed that more infection in males than in females in our study. Male predominance of HFMD was observed, possibly because of higher physical activity and less attention to hygiene of boys. However, there was no gender difference in the ratio of each different enterovirus infection. Statistical analysis of the age in the present study showed that children under age five were the most susceptible to HFMD, which is consistent with previous reports. We found the age of patients with other EV infection was younger than those with EV71 or CV-A16 infection. With the popularity of other EV serotypes, the small-aged infants become the focus of the prevention and treatment of HFMD in Hangzhou.We also found that the average age of EV71 infection gradually increased from 2016 to 2018, especially in 2018, indicating that the protection of EV71 vaccine in low age groups has been obvious effective. In this study, enteroviruses were detected in throat swab and feces, and there was no significant difference in the detection rate of enteroviruses in throat swab and feces (X2=0.127,P=0.722) .No enterovirus was detected in cerebrospinal fluid specimens in our study. Relevant reports in abroad also showed that the detection rate of enterovirus nucleic acid in cerebrospinal fluid was lower than in throat swab, feces or bleb fluid.