The environmental water samples assayed by total culturable virus assay (TCVA) were evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and integrated cell culture-PCR (ICC-PCR) method for enteroviruses and reovirus since they are the usually detected virus groups by culture assays. The detection sensitivities of the TCVA, RT-PCR, and ICC-PCR were compared and the overall reliability of the detection was analyzed to confirm environmental samples for enteric viruses. A total of eight samples from different areas was analyzed by performing TCVA, RT-PCR, and ICC-PCR. Virus concentrations in surface water samples ranged from 1.03 to 47.3 most probable numbers of infectious units (MPN) per 100 liters. When primers specific for both enteroviruses and reoviruses were used in both RT-PCR and ICC-PCR, all the samples (100%) were positive for the viruses. Reoviruses were the most frequently detected ones among the samples. According to the sequence results of enteroviruses, five of the samples were contaminated by coxsackievirus type B3, and the rest by coxsackievirus type B6, echovirus type 30, or vaccine strain poliovirus type 3. It was observed that both enteroviruses and reoviruses were detected concurrently in all CPE-positive environmental water samples.
Enterovirus B, Human ; Enterovirus* ; Poliovirus ; Water*

The environmental water samples assayed by total culturable virus assay (TCVA) were evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and integrated cell culture-PCR (ICC-PCR) method for enteroviruses and reovirus since they are the usually detected virus groups by culture assays. The detection sensitivities of the TCVA, RT-PCR, and ICC-PCR were compared and the overall reliability of the detection was analyzed to confirm environmental samples for enteric viruses. A total of eight samples from different areas was analyzed by performing TCVA, RT-PCR, and ICC-PCR. Virus concentrations in surface water samples ranged from 1.03 to 47.3 most probable numbers of infectious units (MPN) per 100 liters. When primers specific for both enteroviruses and reoviruses were used in both RT-PCR and ICC-PCR, all the samples (100%) were positive for the viruses. Reoviruses were the most frequently detected ones among the samples. According to the sequence results of enteroviruses, five of the samples were contaminated by coxsackievirus type B3, and the rest by coxsackievirus type B6, echovirus type 30, or vaccine strain poliovirus type 3. It was observed that both enteroviruses and reoviruses were detected concurrently in all CPE-positive environmental water samples.
Enterovirus B, Human ; Enterovirus* ; Poliovirus ; Water*

BACKGROUND: Aseptic meningitis is a common illness of children. It seems that viruses are the usual etiologic agents. The distribution of these agents mainly depends on the isolated time and region area. This study was performed to isolate the causative viruses from patients with aseptic meningitis in Gwangju area during recent one year. METHODS:A total of 130 patients with aseptic meningitis were evaluated. Stool and/or cerebrospinal fluid specimens from patients were inoculated into rhabdomyosarcoma (RD), HEp2 and Vero cell lines. The virus propagation was examined by the presence of cytopathic effects. Neutralizing tests using enterovirus serum pool were done on each viral isolates. RESULTS: The isolation rate of enterovirus was 24.6% (32/130). The enterovirus isolates were obtained mostly from stool specimens (29/32). Twenty-two isolates were identified by neutralizing test. Ten isolates disclosed 'untyping' by neutralizing test. The distribution of isolates was coxsackievirus group B2 (11 stains, 34.4%), echovirus 30 (4 strains, 12.5%), echovirus 6 (3 strains, 9.4%), echovirus 9, 11, 25 and coxsackievirus group A16 (1 strain, respectively). These strains were predominantly isolated during summer season (June to July). CONCLUSIONS: The causative viruses from patients with aseptic meningitis in Gwangju area during recent 1 year were coxsackievirus group B2, echovirus 30, 6, 9, 11, 25 and coxsackievirus group A16 which were mostly isolated from stool specimens in summer season.
Cerebrospinal Fluid ; Child ; Coloring Agents ; Echovirus 6, Human ; Echovirus 9 ; Enterovirus ; Enterovirus B, Human ; Gwangju* ; Humans ; Meningitis, Aseptic* ; Rhabdomyosarcoma ; Seasons ; Vero Cells

Objective: To develop RT-nPCR assays for amplifying partial and complete VP1 genes of human enteroviruses (HEVs) from clinical samples and to contribute to etiological surveillance of HEV-related diseases. Methods: A panel of RT-nPCR assays, consisting of published combined primer pairs for VP1 genes of HEV A-C and in-house designed primers for HEV-D, was established in this study. The sensitivity of each RT-nPCR assay was evaluated with serially diluted virus stocks of five serotypes expressed as CCID Results: The sensitivity of RT-nPCR assays for amplifying partial VP1 gene of HEVs was 0.1 CCID Conclusion This RT-nPCR system is capable of amplifying the partial and complete VP1 gene of HEV A-D, providing rapid, sensitive, and reliable options for molecular typing and molecular epidemiology of HEVs in clinical specimens.
Capsid Proteins/genetics* ; Enterovirus A, Human/genetics* ; Enterovirus B, Human/genetics* ; Enterovirus C, Human/genetics* ; Enterovirus D, Human/genetics* ; Humans ; Molecular Epidemiology/methods* ; Molecular Typing/methods* ; Reverse Transcriptase Polymerase Chain Reaction/methods*

PURPOSE: There was an outbreak of aseptic meningitis in Busan and Kyoungsangnamdo area from April to July, 2002. In this study, we reported the clinical manifestations, laboratory data and causative viruses. METHODS: 184 children with aseptic meningitis who had been admitted to Department of Pediatrics, Inje Paik Hospital in Kaegum and Dongrae between April and July 2002 were assessed. Virus isolation and serotype identification were performed by cell culture and reverse transcription-polymerase chain reaction(RT-PCR) of cerebrospinal fluid. RESULTS: The peak incidence was noted in May. Male-to-female ratio was 2:1. The age ranged from 2 years to 16 years. Mean age was 8.7+/-7.1 years. Clinical manifestations were fever(95.1%), headache(91.8%) and vomiting(89.7%). Mean duration of fever was 2.6 days. The numbers of peripheral blood leukocytes were 1,400-20,000/mm3 and mean value was 9,829/mm3. On cerebrospinal fluid examinations, leukocyte count was in range of 10-2,000(mean 301)/mm3, protein level in range of 10-196(mean 36.7)mg/dL and glucose level from 17 to 155(mean 58.3)mg/dL. Viruses were isolated 13(8.4%) out of 154 patients and serotypes were 6 cases of echovirus 6, 3 cases of echovirus 9, 1 case of echovirus 13, 2 cases of echovirus 25 and 1 case of echovirus 30. CONCLUSION: Aseptic meningitis was prevalent in Busan and Kyoungsangnamdo area from April to July, 2002. We thought that causative viruses were echovirus 6, 9, 13, 25, 30.
Busan* ; Cell Culture Techniques ; Cerebrospinal Fluid ; Child ; Echovirus 6, Human ; Echovirus 9 ; Enterovirus B, Human ; Fever ; Glucose ; Gyeongsangnam-do* ; Humans ; Incidence ; Leukocyte Count ; Leukocytes ; Meningitis, Aseptic* ; Pediatrics

PURPOSE: Enterovirus infection in children can manifest various disease and enterovirus have many serotypes. This study was aimed to investigate neurologic manifestations according to serotypes of enterovirus in pediatric inpatients in Incheon. METHODS: We collected the stool samples from the admitted pediatric patients in Inha University Hospital from January 2015 to September 2016. Enterovirus detection and serotypes identification were performed by real-time reverse transcriptase polymerase chain reaction (RT-PCR) and semi-nested RT-PCR. RESULTS: A total of 527 samples were collected during study period and 170 patients (32.2%) were diagnosed with enterovirus infections. Genetic sequences of enteroviruses were identified: echovirus 18 (50, 40.5%), enterovirus 71 (12, 9.6%), coxakievirus A10 (10, 8.0%), echovirus 6 (7, 5.6%). Virus in patient with meningitis were identified: echovirus 18 (15, 75%), coxakievirus B5 (2, 10%), enterovirus 71 (2, 10%), and echovirus 6 (1, 5%). Neurologic manifestations of echovirus 18 are headache (15, 30%), vomiting (17, 34%), meningeal irritation sign (10, 20.0%). And enterovirus 71 have headache (3, 25%), vomiting (3, 25%), meningeal irritation sign (2, 16.0%), seizure (1, 8.3%), neurologic sequelae (1, 8.3%). Echovirus 18 and neurologic manifestation have a statistically significant correlation with other serotypes (r=0.701, P < 0.01) CONCLUSION: Echovirus 18 infection was more prominent in neurological symptoms than in other serotypes. The major serotype of meningitis was echovirus 18 but there was no reported neurologic sequelae. Enterovirus infection has different neurological symptoms, depending on the serotypes.
Child ; Echovirus 6, Human ; Enterovirus B, Human ; Enterovirus Infections ; Enterovirus* ; Headache ; Humans ; Incheon* ; Inpatients* ; Meningitis ; Neurologic Manifestations* ; Reverse Transcriptase Polymerase Chain Reaction ; Seizures ; Serogroup* ; Vomiting

PURPOSE: We identified the causative viruses from patients with aseptic meningitis, acute hemorrhagic conjunctivitis and other enterovirus-related diseases to understand the epidemiological patterns and prevailing strains of enterovirus infections each year. MATERIALS AND METHODS: During 1999-2003, we examined 3,260 specimens from 2,939 patients with aseptic meningitis or other clinical manifestations for the presence of enteroviruses by using both cell culture/ neutralisation test and reverse transcription-polymerse chain reaction-sequencing. To investigate the etiological agents which caused an epidemic of acute haemorrhagic conjunctivitis, conjunctival swab samples from acute haemorrhagic conjunctivitis patients showing cytopathic effects in HEp2 cells were tested by enteroviral specific PCR. RESULTS: We identified 603 isolates of enteroviruses (20.5%) among 2,939 cases and 22 serotypes of human enteroviruses were isolated during this 5 year period. Echovirus 13 and coxsackievirus A24 in 2002 and coxsackievirus A9 in 2003 were the first enterovirus to be indentified in Korea since we began the enterovirus surveillance in 1993. While an epidemic of echovirus 13 infection in Korea began in Gwangju and Jeolla province in 2002 and spread to Seoul, Gyunggi, Busan, Ulsan and other regions, echovirus 6 isolates in 2002 were mainly detected in Busan specimens and some Gwangju samples. From the nucleotide sequencing of enteroviral PCR products of conjunctival swab specimens, we found 85% nucleotide homology to coxsackievirus A24 (D90457). CONCLUSIONS: We isolated 603 enteroviral isolates among 2939 cases during 1999-2003. Echovirus 13 and coxsackievirus A24 were the first enterovirus to be identified in Korea and caused nationwide epidemics in 2002.
Busan ; Conjunctivitis ; Conjunctivitis, Acute Hemorrhagic ; Echovirus 6, Human ; Enterovirus B, Human ; Enterovirus Infections ; Enterovirus* ; Gwangju ; Humans ; Korea* ; Meningitis, Aseptic ; Polymerase Chain Reaction ; Seoul ; Ulsan

PURPOSE: We identified the causative viruses from patients with aseptic meningitis, acute hemorrhagic conjunctivitis and other enterovirus-related diseases to understand the epidemiological patterns and prevailing strains of enterovirus infections each year. MATERIALS AND METHODS: During 1999-2003, we examined 3,260 specimens from 2,939 patients with aseptic meningitis or other clinical manifestations for the presence of enteroviruses by using both cell culture/ neutralisation test and reverse transcription-polymerse chain reaction-sequencing. To investigate the etiological agents which caused an epidemic of acute haemorrhagic conjunctivitis, conjunctival swab samples from acute haemorrhagic conjunctivitis patients showing cytopathic effects in HEp2 cells were tested by enteroviral specific PCR. RESULTS: We identified 603 isolates of enteroviruses (20.5%) among 2,939 cases and 22 serotypes of human enteroviruses were isolated during this 5 year period. Echovirus 13 and coxsackievirus A24 in 2002 and coxsackievirus A9 in 2003 were the first enterovirus to be indentified in Korea since we began the enterovirus surveillance in 1993. While an epidemic of echovirus 13 infection in Korea began in Gwangju and Jeolla province in 2002 and spread to Seoul, Gyunggi, Busan, Ulsan and other regions, echovirus 6 isolates in 2002 were mainly detected in Busan specimens and some Gwangju samples. From the nucleotide sequencing of enteroviral PCR products of conjunctival swab specimens, we found 85% nucleotide homology to coxsackievirus A24 (D90457). CONCLUSIONS: We isolated 603 enteroviral isolates among 2939 cases during 1999-2003. Echovirus 13 and coxsackievirus A24 were the first enterovirus to be identified in Korea and caused nationwide epidemics in 2002.
Busan ; Conjunctivitis ; Conjunctivitis, Acute Hemorrhagic ; Echovirus 6, Human ; Enterovirus B, Human ; Enterovirus Infections ; Enterovirus* ; Gwangju ; Humans ; Korea* ; Meningitis, Aseptic ; Polymerase Chain Reaction ; Seoul ; Ulsan

PURPOSE: There occurred an explosive outbreak of aseptic meningitis in Ulsan, in the year of 2002. In this study, we report clinical manifestations, laboratory data and causative viruses of the outbreak. METHODS: Were assessed 371 children with aseptic meningitis who were admitted to the Department of Pediatrics, Ulsan University Hospital in Ulsan between May and July, 2002. We reviewed the sex, age, clinical manifestations, peripheral blood examinations and CSF examinations of the patients. Virus isolation and serotype identification were performed by cell cultures of cerebrospinal fluid and stool. RESULTS: The ratio of males to females was 1.9:1. The majority of aseptic meningitis occurred in children between 1 and 5 years of age with a mean age of 6.8+/-5.1 years. The clinical manifestations was fever, headache, vomiting, and abdominal pain and 16.2% of the patients developed neck stiffness. The mean duration of fever was 3.2 days. The mean value of peripheral blood leukocytes was 10,355+/-4,800/mm3 and the mean value of CRP was 0.96+/-1.67 mg/dL. Initial CSF findings revealed leukocytes 212+/-385/mm3, protein 37.0+/-18.9 mg/dL and glucose 68.3+/-17.6 mg/dL. Viruses were isolated 67(18.1%) out of 371 patients and the serotypes were echovirus 6 in 10 cases, echovirus 9 in 15 cases, echovirus 11 in 4 cases, echovirus 13 in 18 cases, and unclassified enterovirus in 20 cases. CONCLUSION: Aseptic meningitis was prevalent in Ulsan, in the year of 2002. The clinical manifestations and the laboratory findings of the patients were compatible with those of previous reports. The causative viruses were echovirus 6, echovirus 9, echovirus 11, echovirus 13 and unclassified enterovirus.
Abdominal Pain ; Cell Culture Techniques ; Cerebrospinal Fluid ; Child ; Echovirus 6, Human ; Echovirus 9 ; Enterovirus ; Enterovirus B, Human ; Female ; Fever ; Glucose ; Headache ; Humans ; Leukocytes ; Male ; Meningitis, Aseptic* ; Neck ; Pediatrics ; Ulsan* ; Vomiting

BACKGROUND: The determination of serotype of enteroviruses is useful for the discrimination between sporadic and epidemic infections. The conventional serotyping method is time-consuming and labor-intensive. Recently, molecular method was introduced for the serotyping of enteroviruses. The aim of this study was to establish a method to isolate and analyze enteroviruses from various specimens utilizing molecular biological techniques and to determine which strains were phylogenetically related to clinical samples. METHODS: Clinical samples in this study included 164 cerebrospinal fluid (CSF), 136 stool, 15 sera, 6 throat swab, 5 urine, and 4 sputa, which were obtained from hospitalized patients, primarily infants or children presenting symptoms of aseptic meningitis in 1998. RD cells were used for enterovirus isolation. RT-PCR was performed with RD cell lysate showing CPE. The primers 011 and 012 were used for the VP1 region, and the primers EN1 and EN2 for 5'-UTR. The nucleotide sequences of VP1 region were determined and analyzed with BLAST program. RESULTS: Among 333 samples, only 23 samples produced CPE: 17 samples at first and six samples at the second blind passage. Fifteen isolates were related to coxsackievirus B2 two to echovirus 4, three to echovirus 6, and three to echovirus 18. All 23 viral isolates displayed a nucleotide sequence identity of 80-95%, compared with the reference serotypes. However, the identity was increased up to 93-100% when the VP1 region was translated into amino acids CONCLUSIONS: Since CB2 type was 55% among enteroviral isolates, the CB2 was determined as the major causative serotype of enteroviral meningitis in 1998. CB2 type was emerged between June and July, EC4 and EC6 was limited to July, and EC18 was in August.
Amino Acids ; Base Sequence ; Cerebrospinal Fluid ; Child ; Discrimination (Psychology) ; Echovirus 6, Human ; Enterovirus B, Human ; Enterovirus* ; Humans ; Infant ; Meningitis* ; Meningitis, Aseptic ; Pharynx ; Serotyping*