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Human adenovirus (HAdV-7) is a highly contagious pathogen that causes severe respiratory illnesses. However, the epidemic patterns and genetic variability of HAdV-7 circulating in mainland China have not been well elucidated. In this study, we used Chinese HAdV sentinel surveillance data obtained from 2012-2015 to investigate the clinical features of 122 HAdV-7-positive cases and performed amplification and sequence determination of three capsid genes (penton base, hexon, and fiber) from 69 isolated viruses covering from seven provinces of China. Additionally, we compared with data from representative sequences of 21 strains covering seven more provinces in China and 32 international HAdV-7 strains obtained from GenBank database to determine the phylogenetic, sequence variations, and molecular evolution of HAdV-7. The results indicated that HAdV-7 infection occurred throughout the year, and a high proportion of severe cases (27 cases, 22.1%) exhibited infantile pneumonia. Moreover, phylogenetic analysis showed that all HAdV-7 strains could be divided into two major evolutionary branches, including subtype 1 and subtype 2, and subtype 3 was also formed according to analysis of the penton base gene. Subtypes 1 and 2 co-circulated in China before 2008, and HAdV-7 strains currently circulating in China belonged to subtype 2, which was also the predominant strain circulating worldwide in recent years. Further sequence variation analysis indicated that three genes of HAdV-7 were relatively stable across time and geographic space, particularly for viruses within subtypes, which shared almost the same variation sites. Owing to continuous outbreaks caused by HAdV-7, resulting in increased illness severity and fatality rates in China, the establishment of a national HAdV surveillance system is urgently needed for the development of effective preventive and infection-control interventions for adenovirus respiratory infections in China.

[This corrects the article DOI: 10.1371/journal.pone.0232092.].

Due to the Expanded Program on Immunization (EPI) and supplementary immunization activities (SIAs) in China, the incidence of measles in China has decreased extensively. The incidence reached its lowest levels in contemporary history in 2012 and 2017, with incidence rates of 4.6 and 4.3 per million population, respectively. However, more than 147,000 measles cases were reported from 2013 to 2016. Furthermore, the proportions of cases in infants < 8 months and adults have been increasing since 2013, representing a considerable challenge for measles elimination in China. A total of 14,868 measles viruses were isolated from confirmed measles cases from 2011 to 2017, of which 14,631 were identified as the predominant endemic genotype, H1; 87 were identified as genotype A viruses that were vaccine associated strains; and 150 were identified as non-H1 genotype viruses. The non-H1 genotype viruses included 62 D8 viruses, 70 D9 viruses, 3 D11 viruses, 14 B3 viruses, and 1 G3 virus, which were identified as imported or import-related viruses that caused sporadic cases or small outbreaks. Most of the transmission chains detected during the period 2011-2012 were interrupted and were followed by many new transmission chains of unknown origin that spread, causing a large measles resurgence in China during 2013-2016. After 4 years of measles resurgence and continuous implementation of the routine immunization program and SIAs, the population immunity reached a sufficiently high level to interrupt most of the transmission chains; only a few strains survived, which continued to be sporadically detected in China in 2017. In the present study, the results from the combined epidemiological and molecular virological data demonstrated the great progress towards measles elimination in China by the further analysis of circulation dynamics for the endemic H1 genotype measles virus from 2011 to 2017. And this study accumulated critical baseline data on circulating wild-type measles viruses in China and provides comprehensive information to the world. These comprehensive baseline data provide evidence to support measles elimination in the future, not only in China but also in other countries worldwide. In addition, the information will be very useful to other countries for tracing their sources of measles cases and for identifying transmission links, which can help prevent potential measles outbreaks.

In addition to high vaccination coverage, timely and accurate laboratory confirmation of measles cases is critical to interrupt measles transmission. To evaluate the role of real-time reverse transcription-polymerase chain reaction (RT-PCR) in the diagnosis of measles cases, 46,363 suspected measles cases with rash and 395 suspected measles cases without rash were analyzed in this study; the cases were obtained from the Chinese measles surveillance system (MSS) during 2014-2017 and simultaneously detected by measles-specific IgM enzyme-linked immunosorbent assay (ELISA) and real-time RT-PCR. However, some IgM-negative measles cases were identified by real-time RT-PCR. The proportion of these IgM-negative and viral nucleic acid-positive measles cases was high among measles cases with measles vaccination history, cases without rash symptoms, and cases within 3 days of specimen collection after onset. The proportion of IgM-negative and viral nucleic acid-positive measles cases in the 0-3 day group was up to 14.4% for measles cases with rash and 40% for measles cases without rash. Moreover, the proportions of IgM-negative and nucleic acid-positive measles cases gradually increased with the increase in the measles vaccination dose. Therefore, integrated with IgM ELISA, real-time RT-PCR would greatly improve the accurate diagnosis of measles cases and avoid missing the measles cases, especially for measles cases during the first few days after onset when the patients were highly contagious and for measles cases with secondary vaccine failure. In conclusion, our study reconfirmed that IgM ELISA is the gold-standard detection assay for measles cases confirmation. However, real-time RT-PCR should be introduced and used to supplement the laboratory diagnosis, especially in the setting of pre-elimination and/or elimination wherever appropriate.

With the implementation of mumps virus (MuV) vaccination in the expanded program on immunization (EPI) in mainland China since 2008, the incidence of mumps has decreased, and the natural epidemic pattern of mumps has slightly changed during 2013-2015. The two epidemic peaks (April-July and November-December) became less obvious than those observed from 2004 to 2012. Children and adolescents younger than 15, particularly in the five-to-nine-year-old age group, remain the target group and should be the focus of high-quality immunization activities in mainland China. However, it was also found that the incidence and reported cases of mumps decreased in each age group during 2013-2015, particularly in the five-to-nine-year-old and ten-to-fourteen-year-old age groups. The proportion of mumps cases among adults in some provinces also increased. Unlike the changes in the epidemiological characteristics of mumps affected by vaccination, the data of MuV virology surveillance indicated that most of the MuV transmission chains have not yet been effectively interrupted, and MuV remains a natural epidemic pattern in mainland China. In the MuV virology surveillance, 194 MuV strains during 2013-2015 were isolated from 10 of 31 provinces in mainland China. Based on the phylogenetic analysis of the small hydrophobic (SH) gene, both genotype F (99.0%) and G (1.0%) were identified, and genotype F was still the predominant genotype continuously circulating in mainland China. Representative genotype F and G strains isolated in China from 1995 to 2012 were selected for further analysis. The results indicated that there were multiple transmission chains within genotype F, with no obvious geographical or time differences. The high genetic diversity of genotype F strains could be a result of the continuous transmission and evolution of the MuV in mainland China. Genotype G was also detected in four provinces in mainland China. Because of the limited epidemiological data, it was uncertain whether the genotype G MuV strains found in 2011 and 2013 were imported from other countries. Therefore, combined high-quality epidemiological and virological surveillance is necessary for mumps control; it can also be used to observe the changes in epidemiological characteristics and viral transmission of mumps over time after mumps-containing vaccine (MuCV) implementation and to provide a comprehensive epidemiological and genetic baseline for mumps elimination in mainland China.

BACKGROUND: Acute respiratory infections (ARIs) are the leading cause of children and their leading killer. ARIs are responsible for at least six percent of the world's disability and death. Viruses are one of the most common agents causing ARIs. Few studies on the viral etiology and clinical characteristics of ARIs have been performed in the northwest region of China, including Gansu Province. METHODS: Clinical and demographic information and throat swabs were collected from 279 patients from January 1st to December 30st, 2011. Multiplex RT-PCR was performed to detect 16 respiratory viral pathogens. RESULTS: 279 patients were admitted for ARIs. The patients aged from 1 month to 12 years, with the median age of 2 years. Of which, 105 (37.6%) were positive for at least one pathogen. A total of 136 respiratory viral pathogens were identified from the 105 patients. Respiratory syncytial virus (RSV) was the most frequently detected pathogen (26.5%, 36/136), followed by parainfluenza virus (PIV) 1-3 (22.1%, 30/136), human rhinovirus (HRV) (21.3%, 29/136), human coronavirus (CoV) (10.3%, 14/136) and human adenovirus (HAdV) (9.6%, 13/136). Influenza A (Flu A), human metapneumovirus (hMPV) and human bocavirus (BoCA) were found 4.4%, 3.7% and 2.2%, respectively. Influenza B (Flu B) and seasonal influenza A H1N1(sH1N1) were not detected. Single-infections were detected in 30.5% (85/279) of cases. RSV was the most common pathogens in patients under 1 year and showed seasonal variation with peaks during winter and spring. CONCLUSIONS: This paper presents data on the epidemiology of viral pathogens associated with ARIs among children in Gansu Province, China. RSV is most frequently detected in our study. The findings could serve as a reference for local CDC in drawing up further plans to prevent and control ARIs.

BACKGROUND: China experienced several large measles outbreaks in the past two decades, and a series of enhanced control measures were implemented to achieve the goal of measles elimination. Molecular epidemiologic surveillance of wild-type measles viruses (MeV) provides valuable information about the viral transmission patterns. Since 1993, virologic surveillnace has confirmed that a single endemic genotype H1 viruses have been predominantly circulating in China. A component of molecular surveillance is to monitor the genetic characteristics of the hemagglutinin (H) gene of MeV, the major target for virus neutralizing antibodies. PRINCIPAL FINDINGS: Analysis of the sequences of the complete H gene from 56 representative wild-type MeV strains circulating in China during 1993-2009 showed that the H gene sequences were clustered into 2 groups, cluster 1 and cluster 2. Cluster1 strains were the most frequently detected cluster and had a widespread distribution in China after 2000. The predicted amino acid sequences of the H protein were relatively conserved at most of the functionally significant amino acid positions. However, most of the genotype H1 cluster1 viruses had an amino acid substitution (Ser240Asn), which removed a predicted N-linked glycosylation site. In addition, the substitution of Pro397Leu in the hemagglutinin noose epitope (HNE) was identified in 23 of 56 strains. The evolutionary rate of the H gene of the genotype H1 viruses was estimated to be approximately 0.76×10(-3) substitutions per site per year, and the ratio of dN to dS (dN/dS) was

Coxsackievirus A16 (CVA16) is responsible for nearly 50% of all the confirmed hand, foot, and mouth disease (HFMD) cases in mainland China, sometimes it could also cause severe complications, and even death. To clarify the genetic characteristics and the epidemic patterns of CVA16 in mainland China, comprehensive bioinfomatics analyses were performed by using 35 CVA16 whole genome sequences from 1998 to 2011, 593 complete CVA16 VP1 sequences from 1981 to 2011, and prototype strains of human enterovirus species A (EV-A). Analysis on complete VP1 sequences revealed that subgenotypes B1a and B1b were prevalent strains and have been co-circulating in many Asian countries since 2000, especially in mainland China for at least 13 years. While the prevalence of subgenotype B1c (totally 20 strains) was much limited, only found in Malaysia from 2005 to 2007 and in France in 2010. Genotype B2 only caused epidemic in Japan and Malaysia from 1981 to 2000. Both subgenotypes B1a and B1b were potential recombinant viruses containing sequences from other EV-A donors in the 5'-untranslated region and P2, P3 non-structural protein encoding regions.

Genetic recombination is a well-known phenomenon for enteroviruses. To investigate the genetic characterization and the potential recombination of enterovirus 71 (EV71) circulating in China, we determined the 16 complete genome sequences of EV71 isolated from Hand Foot Mouth Disease (HFMD) patients during the large scale outbreak and non-outbreak years since 1998 in China. The full length genome sequences of 16 Chinese EV71 in present study were aligned with 186 genome sequences of EV71 available from GenBank, including 104 China mainland and 82 international sequences, covering the time period of 1970-2011. The oldest strains of each subgenotype of EV71 and prototype strains of HEV-A were included to do the phylogenetic and Simplot analysis. Phylogenetic analysis indicated that all Chinese strains were clustered into C4 subgenotype of EV71, except for HuB/CHN/2009 clustered into A and Xiamen/CHN/2009 clustered into B5 subgenotype. Most of C4 EV71 were clustered into 2 predominant evolutionary branches: C4b and C4a evolutionary brunches. Our comprehensive recombination analysis showed the evidence of genome recombination of subgenotype C4 (including C4a and C4b) sequences between structural genes from genotype C EV71 and non-structural genes from the prototype strains of CAV16, 14 and 4, but the evidence of intratypic recombination between C4 strains and B subgenotype was not enough strong. This intertypic recombination C4 viruses were first seen in 1998 and became the predominant endemic viruses circulating in China mainland for at least 14 years. A shift between C4a and C4b evolutionary brunches of C4 recombination viruses were observed, and C4a viruses have been associated with large scale nationwide HFMD outbreak with higher morbidity and mortality since 2007.

Large-scale Hand, Foot, and Mouth Disease (HFMD) outbreaks have frequently occurred in China since 2008, affecting more than one million children and causing several hundred children deaths every year. The pathogens of HFMD are mainly human enteroviruses (HEVs). Among them, human enterovirus 71 (HEV71) and coxsackievirus A16 (CVA16) are the most common pathogens of HFMD. However, other HEVs could also cause HFMD. To rapidly detect HEV71 and CVA16, and ensure detection of all HEVs causing HFMD, two real-time hybridization probe-based RT-PCR assays were developed in this study. One is a multiplex real-time RT-PCR assay, which was developed to detect and differentiate HEV71 specifically from CVA16 directly from clinical specimens within 1-2 h, and the other is a broad-spectrum real-time RT-PCR assay, which targeted almost all HEVs. The experiments confirmed that the two assays have high sensitivity and specificity, and the sensitivity was up to 0.1 TCID50/ml for detection of HEVs, HEV71, and CVA16, respectively. A total of 213 clinical specimens were simultaneously detected by three kinds of assays, including the two real-time RT-PCR assays, direct conventional RT-PCR assay, and virus isolation assay on human rhabdomyosarcoma cells (RD cells). The total positive rate of both HEV71 and CVA16 was 69.48% with real-time RT-PCR assay, 47.42% with RT-PCR assay, and 34.58% with virus isolation assay. One HFMD clinical specimen was positive for HEV, but negative for HEV71 or CVA16, which was identified as Echovirus 11 (Echo11) by virus isolation, RT-PCR, and sequencing for the VP1 gene. The two real-time RT-PCR assays had been applied in 31 provincial HFMD labs to detect the pathogens of HFMD, which has contributed to the rapid identification of the pathogens in the early stages of HFMD outbreaks, and helped to clarify the etiologic agents of HFMD in China.

The relevance of human parainfluenza viruses (HPIVs) to the epidemiology of acute respiratory infections (ARI) in China is unclear. From May 2008 to September 2010, 443 nasopharyngeal aspirates (NPAs) from hospitalized pediatric patients (age from 1 to 93 months) in Beijing were collected and screened for HPIVs and other common respiratory viruses by real-time RT-PCR. Sixty-two of 443 samples were positive for HPIVs with 4 positive for HPIV-2 and 58 positive for HPIV-3, indicating that HPIV-3 was the predominant virus present during the study period. A phylogenetic tree based on all the available HN (hemagglutinin-neuraminidase) sequences of HPIV-3 indicated that three distinct clusters (A,B, and C) were circulating with some temporal and regional clustering. Cluster C was further divided into sub-clusters, C1, C2, C3 and C4. HPIV-3 from Beijing isolates belonged to sub-cluster C3, and were grouped with the isolates from two Provinces of China and the neighboring country of Japan. Genetic analysis based on entire HN gene revealed that the HPIV-3 isolates from Beijing were highly similar with 97.2%-100% identity at the nucleotide level and these could be divided into two closely related lineages, C3a and C3b. These findings suggested that there was co-circulation of multiple lineages of HPIV-3 in the Beijing region during the study period. This is the first study to describe the epidemiology and molecular characterization of HPIVs in China.