Isolation and identification of Coxsackievirus A16 strains in Kunming,Yunnan Province in 2019 and analysis of genetic characteristics of VP1 gene / 中国生物制品学杂志

@#Objective To analyze the genetic characteristics of the entire VP1 gene of Coxsackievirus A16(CVA16) strains isolated from the feces of patients with hand,foot and mouth disease(HFMD) in Yunnan Province in 2019.Methods The virus was isolated from human embryonic lung diploid fibroblast(KMB-17) cells and African green monkey kidney(Vero)cells,and the primers for the complete VP1 gene sequence of CVA16 were designed.The target fragment was amplified by RT-PCR and sequenced;the complete VP1 sequence was analyzed by softwares such as MEGA 7.0 and Geneious 9.0.2.Results A total of 26 CVA16 strains were isolated,including eight KMB-17 isolates and 18 Vero isolates.Twenty CVA16isolates were randomly selected for analysis,and three isolates were found to have Bla and 17 B1b genotypes;the nucleotide and amino acid homology of 17 CVA16 B1b isolates were 93.8%—100% and 98.3%—100%,and the nucleotide and amino acid homology with other domestic isolates was 91.1 %—99.2% and 97.3%—99.0%,respectively;the nucleotide and amino acid homology of the three Bla isolates was 98.0%—98.1% and 99.3%,and those with other domestic Bla isolates was 88.7%—98.1% and 98.3%—99.7%,respectively;17 B1b isolates and other three Bla isolates showed the nucleotide and amino acid homology of 87.4%—88.4% and 97.3%—98.7%.Conclusion The CVA16 prevalent in Kunming in 2019 belonged to Bla and B1b genotypes,with B1b as the main strain,and all of them were prevalent strains in the mainland of China.

Gene characteristics of coxsackievirus A16 in hand, foot and mouth disease cases in Baoshan City, Yunnan Province, 2022 / 中国热带医学

@#Abstract: Objective　To analyze the results of the surveillance of hand, foot and mouth disease (HFMD) pathogens in Baoshan City, Yunnan Province in 2022, and to analyze the genetic characteristics of the main epidemic strain coxsackievirus A16 (CVA16), in order to provide scientific basis for the prevention and control of HFMD. Methods　Samples of hand, foot and mouth disease cases in Baoshan City submitted to Yunnan Province in 2022 were selected, the samples were processed, and the viral nucleic acid was extracted, and the Enterovirus (EV) VP4/VP2 binding region gene was amplified with primers (MD91/OL68-1) and sequenced. The sequence was BLAST searched in GenBank to determine the virus type, and then the full sequence of VP1 region was amplified with relevant primers of each type of virus and sequenced. Viruses were identified by using Enterovirus Genotyping Tool Version 1.0. according to the reference documents, the reference sequences were downloaded for making the phylogenetic tree, and the test results were statistically analyzed. Results　A total of 307 clinical samples of hand-foot-mouth disease in Baoshan City in 2022 were detected by real-time RT-PCR. There were 280 laboratory-confirmed cases (91.21%), among which the detection rate of CVA16 was 55.05% (169/307), EV-A71 was 3.58% (11/307), and other enteroviruses were 32.57% (100/307). Of the 206 HFMD test specimens, the VP4/VP2 junction and VP1 gene sequences of enterovirus were amplified and identified, and 29 enterovirus strains were obtained, with a positive virus rate of 14.08% (29/206). All viruses belonged to group A and were divided into 3 serotypes, which included 27 strains of CVA16 (93.10%, 27/29), 1 strain of EV-A71 (3.45%, 1/29), and 1 strain of CVA10 (3.45%, 1/29). Group B, C, and D viruses were not detected. The positive rate of VP4/VP2 binding region and VP1 region of coxsackievirus A16 gene in Baoshan City in 2022 was 13.11% (27/206). The genetic evolution analysis showed that the 27 strains of CVA16 belong to B1a subtype and can be divided into two evolutionary branches. Conclusions　In 2022, the main epidemic strain of HFMD in Baoshan City, Yunnan Province is CVA16 gene, belonging to B1a subtype, which can be divided into two branches, indicating 2 transmission chains prevalent in Baoshan City. Future efforts should focus on strengthening surveillance, improving the quality of monitoring, and understanding the characteristics of viral prevalence..

Establishment of national reference panel for coxsackievirus A16 nucleic acid detection kit / 中国生物制品学杂志

@#ObjectiveTo establish the national reference panel for coxsackievirus A16(CA16)nucleic acid detection kit and related quality standard.MethodsThe CA16 positive and negative samples were collected and screened,and then were filled and lyophilized to establish the national reference panel for CA16 nucleic acid detection kit. According to the cooperative calibration results of various reagent manufacturers,the quality standard of reference panel was determined.Meanwhile,the homogeneity and stability of the national reference panel were well studied.ResultsThe national reference panel of CA16 nucleic acid detection kit consisted of 9 positive samples,8 negative samples,1 limit-detecting sample and1 precision sample. The quality standard was as follows:the coincidence rate of positive samples was no less than 8/9;The coincidence rate of negative samples was 8/8;The minimum detection limit required that the dilution of limit-detecting sample was no less than 1∶103;The precision required that the coefficient of variation(CV)of Ct value of 10 precision samples diluted 100 times was no higher than 5% and the results were all positive. The homogeneity of the reference panel met the requirement,and the stability was not affected by the storage at room temperature(25 ℃)for 24 hours and repeated freezing and thawing three times.ConclusionThe first national reference panel of CA16 nucleic acid detection kit and the related quality standard have been established,which provided a reference for the quality control and evaluation of the related reagents.

Pathogen surveillance and genetic characteristics of coxsackievirus A6 and A16 in children with hand, foot and mouth disease in Wenshan Prefecture of Yunnan Province, 2020 / 中华微生物学和免疫学杂志

Objective:To analyze the etiological characteristics of hand, foot and mouth disease (HFMD) cases and to investigate the genetic characteristics of VP1 region of coxsackievirus A6 (CVA6) and CVA16 strains in Wenshan prefecture of Yunnan Province in 2020.Methods:Virus RNA was extracted directly from stool samples of children with HFMD in Wenshan prefecture of Yunnan Province in 2020. Enterovirus (EV) VP4/VP2 junction region was amplified using MD91/OL68-1 primers and sequenced, and then the serotypes of EV isolates were preliminarily identified. Amplification and sequencing of the complete VP1 gene were performed. A phylogenetic tree was constructed by MEGA 5.2 software with the reference strains from GenBank. Genetic and molecular epidemiological characteristics of CVA6 and CVA16 were analyzed.Results:Thirty-two strains of EV and one strain of astrovirus MLB1 were detected in 317 specimens with an overall virus detection rate of 10.41% (33/317). Among the 32 EV strains, 31 (96.88%) were enterovirus species A (EVA) and one (3.12%) was EVB. EVC and EVD were not detected. CVA6 was the predominant EV, accounting for 62.50% (20/32), followed by CVA16 (18.75%, 6/32), CVA4 (9.37%, 3/32) and CVA10 (3.12%, 1/32). EVA71 was not detected. The phylogenetic analysis showed 20 CVA6 strains belonged to D3a sub-genotype and could be further divided into three clusters. Six CVA16 strains belonged to B1a sub-genotype, which was one of the predominant genotypes circulating in China, and could be divided into two clusters.Conclusions:The detection rate of pathogens causing HFMD in Wenshan prefecture in 2020 was 10.41% and the most common etiologic agents were CVA6 and CVA16. Based on the genetic analysis of the VP1 gene, the predominant genotype circulating in Wenshan prefecture in 2020 was CVA6 D3a sub-genotype, followed by CVA16 B1a sub-genotype. EVA71 was not detected.

Evolution of VP1 gene of coxsackievirus A16 in hand, foot, and mouth disease / 中华传染病杂志

Objective To investigate the genetic evolution of VP1 gene of pathogenic coxsackievirus A16 (CV-A16) strain isolated from clinical hand,foot,and mouth disease (HFMD) patients.Methods A total of 160 HFMD cases with CV-A16-positive results were collected from hospitals in Kunming during January 2015 to June 2017.Fecal samples were collected.Real-time polymerase chain reaction (PCR) was used to detect the CV-A16 virus nucleic acid.The VP1 genes of CV-A16-positive samples were amplified by reverse transcription-PCR.The amplified positive products were sequenced and aligned.The homologies were identified and their subgenotypes were determined.The phylogenetic tree was constructed and homology modeling was conducted.Results All the 160 CV-A16 isolates were B2 subtypes.The genetic distance between detected strains of CV-A16 and the strains in Fujian,Beijing,Nanjing was 0.76.The genetic distance to the strains in Malaysia was 0.78,and to the strains in Australia was 1.86.Homologous modeling revealed that the amino acid sequence of the VP1 gene of the strain had a G227R mutation.Conclusions There is no major genetic variation in the CV-A16 strains during 3 years.CV-A16 isolates are close to those of epidemic strains in Beijing,Fujian and Malaysia,but are far fram the strains from Australia.

Genetic characteristics of VP1 region of enterovirus Coxsackievirus A16 in Xining city from 2017 to 2018 / 中华实验和临床病毒学杂志

Objective@#To investigate the genetic characteristics of VP1 coding region of enterovirus Coxsackievirus A16(CV-A16) and etiological features of hand, foot and mouth disease (HFMD) in 2017 in Xining city.@*Methods@#The pharyngeal swab specimens were collected from HFMD patients, and detected by real-time reverse transcription-polymerase chain reaction (RT-PCR). For CV-A16 positive samples, virus isolation was performed. Then RNA was extracted, and then VP1 coding region was amplified by RT-PCR. The phylogenetic tree was constructed by comparing with other genotypes and sub-genotypes strains of EV-A71.@*Results@#It was shown that 70 strains of CV-A16 were isolated from 2017 to 2018 in Xining city. In 2017, 10 strains were isolated and divided into two different lineages by phylogenetic analysis, 3 strains of B1a and 7 stains of B1b. In 2018, 60 stains were isolated, which were all belong to B1b.@*Conclusions@#B1a and B1b of CV-A16 are prevalent in Xining city from 2017 to 2018, in which B1b is the prominent isolates.

Genetic characteristics of coxsackievirus A16 isolated in Shenzhen from 2016 to 2017 / 中华微生物学和免疫学杂志

Objective To investigate the genetic characteristics of VP1 genes carried by coxsack-ievirus A16 strains isolated from cases of hand foot and mouth disease ( HFMD) in Shenzhen during 2016 to 2017. Methods Fecal and anal swab specimens were collected from patients with mild HFMD in four senti-nel hospitals and the Institute of Pathogen Biology, Shenzhen Center for Disease Control and Prevention, China during 2016 to 2017. All specimens were tested for CVA16 viral RNA using real-time RT-PCR. The VP1 genes of 51 randomly selected CVA16 strains were amplified by RT-PCR and then sequenced using TaKaRa Biomedical Technology ( Dalian). Bioinformatics software, including Mega6. 02, BioEdit and DNAStar, was used for comparison and analysis of the VP1 genes. Results CVA16 strains in Shenzhen during 2016 to 2017 mainly belonged to B1a and B1b subtypes as well as an emerging subtype B3. The epi-demic of B1b subtype was found in both 2016 (28 strains) and 2017 (19 strains), while the B1a subtype ( two strains) was only detected in 2017. Two B3 subtype strains were detected in 2017. The strains of B1b subtype were closely related to the strains isolated in Shanghai ( JQ314149 ) , Wenzhou ( KP289416 ) and Beijing (KU254598), while the B1a subtype strains were closely related to the strains isolated in Kunming (JQ316639) and Tailand (GQ184139). The B3 subtype strain was an emerging CVA16 epidemic strain in mainland China. Further comparison of the CVA16 epidemic strains in Shenzhen area during 2016 to 2017 with the CVA16 strains causing severe neurological symptoms showed that two amino acid mutations ( S14N and M23L) were found in VP1 protein. Conclusions The epidemic strains of CVA16 were B1b subtype in Shenzhen area in 2016. However, B1a, B1b and the emerging B3 subtype strains were prevalent in 2017. Compared with the CVA16 strains causing severe neurological symptoms, the CVA16 strains circulating in Shenzhen during 2016 to 2017 carried two amino acid mutations inVP1 protein.

Genetic characteristics of coxsackievirus A16 isolated in Shenzhen from 2016 to 2017 / 中华微生物学和免疫学杂志

Objective@#To investigate the genetic characteristics of VP1 genes carried by coxsackievirus A16 strains isolated from cases of hand foot and mouth disease (HFMD) in Shenzhen during 2016 to 2017.@*Methods@#Fecal and anal swab specimens were collected from patients with mild HFMD in four sentinel hospitals and the Institute of Pathogen Biology, Shenzhen Center for Disease Control and Prevention, China during 2016 to 2017. All specimens were tested for CVA16 viral RNA using real-time RT-PCR. The VP1 genes of 51 randomly selected CVA16 strains were amplified by RT-PCR and then sequenced using TaKaRa Biomedical Technology (Dalian). Bioinformatics software, including Mega6.02, BioEdit and DNAStar, was used for comparison and analysis of the VP1 genes.@*Results@#CVA16 strains in Shenzhen during 2016 to 2017 mainly belonged to B1a and B1b subtypes as well as an emerging subtype B3. The epidemic of B1b subtype was found in both 2016 (28 strains) and 2017 (19 strains), while the B1a subtype (two strains) was only detected in 2017. Two B3 subtype strains were detected in 2017. The strains of B1b subtype were closely related to the strains isolated in Shanghai (JQ314149), Wenzhou (KP289416) and Beijing (KU254598), while the B1a subtype strains were closely related to the strains isolated in Kunming (JQ316639) and Tailand (GQ184139). The B3 subtype strain was an emerging CVA16 epidemic strain in mainland China. Further comparison of the CVA16 epidemic strains in Shenzhen area during 2016 to 2017 with the CVA16 strains causing severe neurological symptoms showed that two amino acid mutations (S14N and M23L) were found in VP1 protein.@*Conclusions@#The epidemic strains of CVA16 were B1b subtype in Shenzhen area in 2016. However, B1a, B1b and the emerging B3 subtype strains were prevalent in 2017. Compared with the CVA16 strains causing severe neurological symptoms, the CVA16 strains circulating in Shenzhen during 2016 to 2017 carried two amino acid mutations inVP1 protein.

Hand, Foot and Mouth Disease in the Andaman Islands, India

Objectives: To investigate an outbreak of Hand, foot and mouth disease (HFMD) inAndaman Islands during 2013. Methods: Epidemiological, clinical data and samples werecollected from HFMD patients who attended selected hospitals. Data were analyzed andsamples were processed for detection of Enterovirus and further confirmed by sequencing.Serotype-specific molecular typing was also done to identify the etiological agent. Results:Of the 246 suspected patients, most were affected in August 2013 (92/246, 37.4%). Fever(71.2%) associated with typical HFMD rashes (100%) were the most common presentingsymptoms and rashes were mostly distributed on hands (100%), legs (92%), mouth (77%),and buttocks (52.8%). All cases were reported as mild and recovered completely without anycomplications. Enterovirus was detected in 63 cases (50.4%). Conclusion: HFMD was mild,mostly reported in children <60 months of age, and in boys. Coxsackie virus A16 was found tobe the only etiological agent for this specific outbreak.

Effect of Coxsackievirus A16 on primary gerbil muscle cells and its mechanism / 预防医学

Objective To explore the killing mechanism induced by Coxsackievirus A16 (CV-A16) in primary muscle cells of gerbils, and to lay the foundations for elucidation the pathogenesis of CV-A16 and the further application of gerbil model. Methods The primary muscle cell model was established by digestion of trypsase/collagenase double enzyme hydrolysis. Primary muscle cells were infected by different dose of CV-A16 and the cell viability was detected by CCK-8 assays. Chromatin condensation and break were measured by Hoechst 33258 staining. The early and last stage of apoptosis cells were measured by AnnexinV/PI double staining. Expression changes of Caspase-3, Caspase-8, JNK and NF-κB pathway proteins were detected by Western Blot. Results The cell viability were 88.95％ and 64.05％ at groups of different multiplicity of infection (MOI=0.50 and 1.00), which was significantly different from those of the negative control group. The cell viability and multiplicity of infection were negative correlation (rs=-0.857, P=0.014) . The apoptosis rates were 7.2％, 21.8％ and 50.7％ at MOI=0.01,0.10 and 1.00 groups, respectively. The apoptosis rate and MOI were positive correlation (rs=1.000, P<0.001) . When the primary cells were infected by CV-A16, cleavage of Caspase-3 and Caspase-8 were detected. Western Blot assays showed that the expression of NF-κB pathway proteins IκBα, p65 and p-p65 were reduced, which was different in enterovirus 71-infected cells. The JNK kinase was actived. Conclusion CV-A16 could induce apoptosis in primary muscle cells from gerbils.

Effect of Siji Kangbingdu Mixture on Inhibition of the Proliferation of Coxsackievirus A16 / 医药导报

Objective To verify the antiviral effects of Siji Kangbingdu mixture (SJKBDM) against coxsackievirus A16 (CoxA16).Methods Vero cells and 5-day-old suckling mice, injected with 75/50 and 1×106 TCID50 CoxA16, were used as evaluation models.The preventive influences of SJKBDM against CoxA16 in Vero cells were assessed in the models.The effects of SJKBDM on the mortality, survival time, change rate of body weight, and clinical symptom scores of suckling mice were observed.Results ①The half maximal inhibitory concentration of SJKBDM on Vero cells was 9.59 mg·mL-1.②Toxic effects were not observed from 32.3 g·kg-1 single dose or continuous intraperitoneal injectin of SJKBDM in suckling BALB/c mice.③The SJKBDM had significant inhibitory effect against CoxA16 virus.Doses higher than 1.22 mg·mL-1 could significantly improve the Vero cell survival rate, and the SJKBDM inhibition of 75/50 TCID50 CoxA16 induced pathological changes in Vero cells.④The SJKBDM significantly improved clinical symptoms of mice with CoxA16 viral infection, especially with crude drug doses of higher than 1.62 g·kg-1.The survival rate and other indicators were comparable or slightly higher compared with ribavirin, and the clinical score was higher than that of ribavirin.Conclusion The SJKBDM has significant inhibitory effect on CoxA16 cell proliferation, significantly decreases death rate, and improves clinical symptoms of mice infected with CoxA16 virus.

Establishment of an animal model for coxsackievirus A16 infection related immunological evaluation / 中国比较医学杂志

Objective To establish a simple and reliable experimental rodent model sensitive to coxsackievirus A16 ( CVA16) .Methods Mongolian gerbils with different age were selected and inoculated intraperitoneally with live CVA16, and the gerbils were observed daily until 14 days postinoculation to screen for the most optimal ages sensitive to the virus.The dose-dependent symptoms were evaluated and the 50% lethal dose (LD50) was determined.The virus titers were measured in blood and various tissues of CVA 16-infected Mongolian gerbils 3 days post-infecton.Finally, the gerbils were immunized twice with inactivated CVA 16 vaccine at day 1 and day 11, respectively, followed by challenge with the virus with a dose of LD50 at day 14.The gerbils were then observed for another 2 weeks to record their body weight , symptom and mortality rate .Their blood samples were collected from the eyes , and CVA16-specific neutralizing antibodytiters and total antibody titers was checked by microneutralization test and ELISA , respectively .Results Various clinical symptoms, such as inactivity, hind limb weakness, paralysis and even death occurred in gerbils following CAV 16 infection. 7-day-old and 14-day-old gerbils are susceptible to CVA 16 infection whereas 28-day-old gerbils are resistant .The most sensitive and appropriate age is 14-day-old.The 50%lethal dose was determined to be 1×104.5 CCID50.High titers of the virus were confirmed in blood and various tissues of Mongolian gerbils contracted CAV 163 days post-infecton.The survival rate is 87.5%for 14-day-old gerbils preimmunized with two doses of inactivated CVA 16 vaccine and challenged with the virus.The geometric mean titers ( GMTs) of neutralizing antibody was 28.14, and the seroprevalence was 87.5%.Conclusions Mongolian gerbils is sensitive to CVA16 and the virus reproduces actively in Vivo.Thus, it can be used as a reliable small animal model for studies of CVA 16 pathogenesis , vaccine development and drug evaluation .

Epidemic characteristics of hand, foot and mouth disease related virus in Suzhou from 2010 to 2015 / 中华实验和临床病毒学杂志

Objective@#To investigate the epidemical features, etiological and clinical characteristics of HFMD in Suzhou city, from 2011 to 2015, providing the scientific supports for HFMD prevention and control.@*Methods@#In each district of Suzhou city, at least five specimens of mild cases were collected per month, while all of the severe cases were sampled. The RNA from each sample was examined using a commercially available real-time PCR kit. Statistical analysis was performed using SPSS version 11.5 software.@*Results@#We retrospectively analyzed HFMD epidemiological data in Suzhou from 2011 to 2015, a total of 4 552 outpatients in Suzhou city were diagnosed with HFMD, including 2 818 positive specimen, the total positive rate was 61.90%, and there was a significant difference in the positive rates between the adjacent years (χ2=186.09, P<0.0001). From 2011 to 2015 in Suzhou, HFMD mainly affected children aged 1 to 5 years old, 66.17% of them were 1to 3 years old. Enterovirus 71(EV71) and coxsackievirus A16(CVA16) were the predominant viral genotypes in Suzhou from 2011 to 2012, and 2014. In 2013, other EVs were dominant, the severe cases mainly correlated with EV71 subtypes, and the proportion of patients with severe disease was significantly decreased in 2013.@*Conclusions@#EV71 and CVA16 are still the important pathogens of HFMD, other EVs also occupy a certain proportion, it is the time to select one or two advantage pathogens from the other EVs into the HFMD monitoring.

Epidemic and etiological characteristics of hand, foot and mouth disease in Gansu province during 2013—2015 / 中华实验和临床病毒学杂志

Objective@#To analyze the epidemiological and etiological characters of hand, foot and mouth disease ( HFMD ) in Gansu during 2013 to 2015 and provide evidence for the development of effective prevention and control measures.@*Methods@#The descriptive epidemiological analysis was used to analyze the data of HFMD, and the specimens were collected from hospitals to detect the pathogens by RT- PCR or Real-time RT-PCR. The nucleotide sequences of VP1 encoding region of virus strains were amplified by RT-PCR method, and determined and analyzed.@*Results@#Total of 29 934 HFMD cases were reported in Gansu from 2013 to 2015, including 81 severe cases and 2 deaths; the largest number occurred in Lanzhou was 7 053, accounting for 23.56% of the cases in the province. HFMD cases were mostly reported during May to July, accounted for 61.69% in total cases; the male to female ratio was 1.5: 1, and most cases were under the age of five and accounted for 83.02%. A total of 5 251 laboratory confirmed cases were reported, of which the number of cases caused by human enterovirus (HEV) were 2 972, the positive rate was 56.60%. Among the severe 81 cases, the positive rate of HEV was 67.90%, and both of the 2 death cases were infected by EV71. 341 strains of viruses were isolated, the genotyping of VP1 encoding region showed that all the 133 EV71 were C4a; among the 134 CVA16 isolates, 6 were B1a and 128 were B1b.@*Conclusions@#In Gansu province, there is a high infection rate of HFMD in children under the age of five, the proportion of other HEV is more and more, and the incidence is related to the difference of pathogens that showed alternant epidemic characteristics. CVA16 and EV71 viruses have the phenomenon of alternating.

Acute pancreatitis in hand, foot and mouth disease caused by Coxsackievirus A16: case report / 소아과

Coxsackievirus A16 (CA16), which primarily causes hand, foot, and mouth disease (HFMD), is associated with complications, such as encephalitis, acute flaccid paralysis, myocarditis, pericarditis, and shock. However, no case of pancreatitis associated with CA16 has been reported in children. We report a case of CA16-associated acute pancreatitis in a 3-year-old girl with HFMD. She was admitted because of poor oral intake and high fever for 1 day. Maculopapular rashes on both hands and feet and multiple vesicles on the soft palate were observed on physical examination. She was treated conservatively with intravenous fluids. On the fourth hospital day, she had severe abdominal pain and vomiting. The serum levels of amylase and lipase were remarkably elevated (amylase, 1,902 IU/L; reference range, 28–100 IU/L; lipase, >1,500 IU/L; reference range, 13–60 IU/L), and ultrasonography showed diffuse swelling of the pancreas with a small amount of ascites. The real-time reverse transcription polymerase chain reaction result from a stool sample was positive for CA16. CA16 can cause acute pancreatitis, and should be considered in the differential diagnosis of abdominal pain in children with HFMD.

Research advances in the etiology and detection method of hand,foot and mouth disease / 中华实用儿科临床杂志

Hand,foot and mouth disease was caused by many kinds of enterovirus,while enterovirus 71 (EV71)and coxsackievirus A16(CA16)were prevalent causative agents of this disease,and each various subtypes of EV71 and CA16 were appear alternatively. Early diagnosis is significant for timely treatment and prognosis of children with hand,foot and mouth disease. Now,the laboratory detection methods of the etiology including virus isolation cul-ture,reverse transcription polymerase chain reaction,real - time polymerase chain reaction,reverse transcription loop -mediated isothermal amplification,enzyme - linked immunosorbent assay,neutralization tests. This review will summa-rized the advancement in the research of the etiology and detection method of hand,foot and mouth disease.

Comparative pathogenicity of Coxsackievirus A16 circulating and noncirculating strains in vitro and in a neonatal mouse model

An enterovirus 71 (EV71) vaccine for the prevention of hand, foot, and mouth disease (HMFD) is available, but it is not known whether the EV71 vaccine cross-protects against Coxsackievirus (CV) infection. Furthermore, although an inactivated circulating CVA16 Changchun 024 (CC024) strain vaccine candidate is effective in newborn mice, the CC024 strain causes severe lesions in muscle and lung tissues. Therefore, an effective CV vaccine with improved pathogenic safety is needed. The aim of this study was to evaluate the in vivo safety and in vitro replication capability of a noncirculating CVA16 SHZH05 strain. The replication capacity of circulating CVA16 strains CC024, CC045, CC090 and CC163 and the noncirculating SHZH05 strain was evaluated by cytopathic effect in different cell lines. The replication capacity and pathogenicity of the CC024 and SHZH05 strains were also evaluated in a neonatal mouse model. Histopathological and viral load analyses demonstrated that the SHZH05 strain had an in vitro replication capacity comparable to the four CC strains. The CC024, but not the SHZH05 strain, became distributed in a variety of tissues and caused severe lesions and mortality in neonatal mice. The differences in replication capacity and in vivo pathogenicity of the CC024 and SHZH05 strains may result from differences in the nucleotide and amino acid sequences of viral functional polyproteins P1, P2 and P3. Our findings suggest that the noncirculating SHZH05 strain may be a safer CV vaccine candidate than the CC024 strain.

The antiviral and antimicrobial activities of licorice, a widely-used Chinese herb

Licorice is a common herb which has been used in traditional Chinese medicine for centuries. More than 20 triterpenoids and nearly 300 flavonoids have been isolated from licorice. Recent studies have shown that these metabolites possess many pharmacological activities, such as antiviral, antimicrobial, anti-inflammatory, antitumor and other activities. This paper provides a summary of the antiviral and antimicrobial activities of licorice. The active components and the possible mechanisms for these activities are summarized in detail. This review will be helpful for the further studies of licorice for its potential therapeutic effects as an antiviral or an antimicrobial agent.

Enterovirus types and clinical characteristics of 5 070 cases with hand,foot and mouth disease in Shen-yang area,2014 / 中国小儿急救医学

Objective To study the epidemic characteristics,distribution of pathogen and clinical characteristics of hand,foot and mouth disease in Shenyang area,2014.Methods Swab specimens was collected from 5 070 cases of hand,foot and mouth disease caces,the F-PCR method was adopted for entero-virus 71 (EV71 ),coxsackievirus A 16(CA16)and universal enteroviruses(EU)detection,and combining the relevant clinical data for comparative analysis .Results June to August was the peak epidemic period.5 070 nasopharyngeal swab specimens were detected positive 3 715 intestinal virus nucleic acid,and the detection rate was 73.27%,including CA16 positive 1 481 (39.87%),EU positive 1 148 (30.90%),EV71 positive 1 086(29.23%).The proportion of EU positive was highest in June(29.71 %).The proportion of EV71 pos-itive(24.78%)and CA16 positive(33.27%)were highest in July,respectively .The proportion of nervous system symptoms in EV71 infection group(88 /1 12,78.57%)was higher than those in EU infection group (97 /147,65.99%)and CA16 infection group (44 /78,56.41 %).The proportion of abnormal myocardial enyzme in CA16 infection group (37 /78,47.44%)was higher than those in EU infection group (48 /147, 32.65%)and EV71 infection group(34 /1 12,30.36%).Conclusion CA16 is the major pathogen causing hand,foot and mouth disease in Shenyang area,2014.Severe hand,foot and mouth is still dominated by EV71 .

Genetic characteristics of coxsackievirus A16 strains isolated in Liaocheng city in Shandong province ;during 2013 / 中华微生物学和免疫学杂志

Objective To study the etiological agent of hand, foot and mouth disease ( HFMD) and the genetic characteristics of coxsackievirus A16 ( CVA16 ) strains isolated from clinical specimens of patients with HFMD in Liaocheng city in 2013.Methods Throat swab and stool specimens were collected from patients with HFMD in the disease surveillance hospitals in Liaocheng city from January to December 2013.Samples pos-itive for CVA16 strains were screened out for the isolation of virus strains with rhabdomyosarcoma ( RD) cells and Vero cells.The entire VP1 coding regions of 9 randomly selected CVA16 isolates were amplified and se-quenced.BioEdit and MEGA4 softwares were used for homology analysis.A phylogenetic tree among the 9 CVA16 isolates and 56 CVA16 representative strains of known genotypes and subgenotypes was constructed.Re-sults The results of PCR analysis showed that 747(77.73%) out of 961 specimens were positive for HFMD and among them, 74 samples (9.91%) were positive for EV71 strains, 130(17.40%) were CVA16 strains and 543(72.69%) were other enterovirus strains.The 9 CVA16 strains clustered into the B2b evolution branch of B genotype with the representative strains, sharing 97.7%to 100%homologies in nucleotide sequences and 99.3%to 100%in amino acid sequences.Conclusion Although EV71 and CVA16 strains were identified, other enteric viruses were the predominant pathogens causing HFMD in Liaocheng city in 2013.The CVA16 iso-lates belonged to B2b subgenotype.The pathogen spectrum of HFMD had already changed.It is necessary to strengthen the surveillance for EV71, CVA16 and other enteric viruses and understand their genetic characteriza-tions, which would be of great significance for the prevention and control of HFMD.