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[Enterovirus type 70, a pathogenic agent of acute haemorrhagic conjunctivitis (author's transl)]

Kopecká D

Cesk Epidemiol Mikrobiol Imunol, 1977 Nov;25(6):342-4.
PMID: 188556
Matched MeSH terms: Enterovirus/isolation & purification
Detection of enteroviruses from clinical specimens

Poh CL, Tan EL

Methods Mol Biol, 2011;665:65-77.
PMID: 21116796 DOI: 10.1007/978-1-60761-817-1_5

Enteroviruses are positive stranded RNA viruses belonging to the genus Enterovirus of the Picornaviridae family. Human enteroviruses are transmitted through the fecal-oral route and have been shown to cause mild to life-threatening diseases. Various diagnostic methods have been developed to detect enteroviruses from clinical specimens but many were impeded by requirements for special reagents, lengthy procedures, low sensitivity or cross-reactivity. This chapter describes rapid and highly sensitive methods of enteroviral detection directly from clinical specimens based on a conventional one-step Reverse Transcription polymerase chain reaction (RT-PCR) and a one-step real-time RT-PCR.

Matched MeSH terms: Enterovirus/isolation & purification*
Fulltext Outbreaks of enterovirus 71 infection

AbuBakar S, Chan YF, Lam SK

N Engl J Med, 2000 Feb 3;342(5):355-6.
PMID: 10660400 DOI: 10.1056/NEJM200002033420513
Matched MeSH terms: Enterovirus/isolation & purification
Neurogenic pulmonary oedema and enterovirus 71 encephalomyelitis

Lum LC, Wong KT, Lam SK, Chua KB, Goh AY

Lancet, 1998 Oct 24;352(9137):1391.
PMID: 9802304
Matched MeSH terms: Enterovirus/isolation & purification*
An outbreak of acute conjunctivitis caused by coxsackievirus A24 in Kuala-Lumpur, Malaysia, 1978

Tan DS, Yin-Murphy M, Kandiah S

Southeast Asian J. Trop. Med. Public Health, 1980 Mar;11(1):24-7.
PMID: 6250226

An investigation of an outbreak of acute conjunctivitis in Kuala Lumpur from May to August 1978 was made. A total of 2,133 cases was involved, most of whom were adult Malay males of low income status from the surrounding villages and low-cost flats. The majority of cases had bilateral conjunctivitis with clear discharge. Pain and subconjunctival haemorrhage were not common and recovery, mostly without complications, occurred within 1 week. Eye scrapings and paired sera specimens were examined and the causal agent was found to be Coxsackievirus A24 (CA24).

Matched MeSH terms: Enterovirus/isolation & purification
Incorrect identification of recent Asian strains of Coxsackievirus A16 as human enterovirus 71: improved primers for the specific detection of human enterovirus 71 by RT PCR

Perera D, Podin Y, Akin W, Tan CS, Cardosa MJ

BMC Infect Dis, 2004 May 4;4:11.
PMID: 15122971

Human enterovirus 71 has emerged as an important pathogen in the Asia Pacific region and it is important to be able to make a rapid and specific diagnosis for outbreak control. Recent Asian strains of Coxsackievirus A16 have changes in the VP1 gene which causes mispriming of widely used primers for human enterovirus 71 specific identification.

Matched MeSH terms: Enterovirus/isolation & purification*
Fulltext Enterovirus type 70: the etiologic agent of pandemic acute haemorrhagic conjunctivitis

Mirkovic RR, Kono R, Yin-Murphy M, Sohier R, Schmidt NJ, Melnick JL

Bull World Health Organ, 1973;49(4):341-6.
PMID: 4368683

A new enterovirus, now classified as enterovirus type 70, was isolated from the conjunctiva of patients with acute haemorrhagic conjunctivitis during the 1971 epidemics that occurred in Japan, Singapore, and Morocco. These epidemics were parts of a pandemic involving Africa (Algeria, Ghana, Morocco, Nigeria, and Tunisia), Asia (Cambodia, China (Province of Taiwan), Hong Kong, India, Indonesia, Japan, Malaysia, the Philippines, Singapore, and Thailand), and England during 1969-71. A representative strain from each of the three epidemic areas was studied cooperatively. The strains exhibited the physicochemical characteristics of enteroviruses. Cross-neutralization tests showed that these viruses were distinct from all known human enterovirus immunotypes, but that they were antigenically closely related. The human origin of the viruses was demonstrated by the appearance of homologous neutralizing antibodies during convalescence in patients with acute haemorrhagic conjunctivitis.

Matched MeSH terms: Enterovirus/isolation & purification*
Fulltext Isolation and Identification of Inter-Species Enterovirus Recombinant Genomes

Bentley K, Tee HK, Pearson A, Lowry K, Waugh S, Jones S, et al.

Viruses, 2021 11 29;13(12).
PMID: 34960659 DOI: 10.3390/v13122390

Positive-strand RNA virus evolution is partly attributed to the process of recombination. Although common between closely genetically related viruses, such as within species of the Enterovirus genus of the Picornaviridae family, inter-species recombination is rarely observed in nature. Recent studies have shown recombination is a ubiquitous process, resulting in a wide range of recombinant genomes and progeny viruses. While not all recombinant genomes yield infectious progeny virus, their existence and continued evolution during replication have critical implications for the evolution of the virus population. In this study, we utilised an in vitro recombination assay to demonstrate inter-species recombination events between viruses from four enterovirus species, A-D. We show that inter-species recombinant genomes are generated in vitro with polymerase template-switching events occurring within the virus polyprotein coding region. However, these genomes did not yield infectious progeny virus. Analysis and attempted recovery of a constructed recombinant cDNA revealed a restriction in positive-strand but not negative-strand RNA synthesis, indicating a significant block in replication. This study demonstrates the propensity for inter-species recombination at the genome level but suggests that significant sequence plasticity would be required in order to overcome blocks in the virus life cycle and allow for the production of infectious viruses.

Matched MeSH terms: Enterovirus/isolation & purification
Fulltext Cytokine and Chemokine Profiling in Patients with Hand, Foot and Mouth Disease in Singapore and Malaysia

Teo FMS, Nyo M, Wong AA, Tan NWH, Koh MT, Chan YF, et al.

Sci Rep, 2018 03 06;8(1):4087.
PMID: 29511232 DOI: 10.1038/s41598-018-22379-6

Hand, foot and mouth disease (HFMD) is a prevalent contagious childhood disease typically associated with fever, oral lesions and limb exanthema. While HFMD is caused by a plethora of serotypes of viruses under the genus Enterovirus within the Picornaviridae family, Coxsackievirus A16 (CV-A16) and Enterovirus 71 (EV-A71) are considered the main etiological agents. In recent years however, other viruses have also been isolated in considerable numbers from infected individuals in many regions, joining the legion commonly associated with HFMD. The present study investigated the cytokine and chemokine profiles of HFMD patients from Singapore and Malaysia for the first time. Comparative cohort studies of EV-A71-associated HFMD cases revealed that the Malaysia cohort had a distinct profile from the Singapore cohort, and this could be partly attributed by different EV-A71 genotypes. As the isolation of CV-A6, instead of CV-A16, had become prevalent in the Singapore cohort, it was also of particular interest to study the differential cytokine and chemokine profiles. Our data revealed that overlapping as well as unique profiles exist between the two major causative clinical isolates in the Singapore cohort. Having a better understanding of the respective immunological profiles could be useful for more accurate HFMD diagnosis, which is imperative for disease transmission control until multi-valent vaccines and/or broad-spectrum anti-viral drugs become available.

Matched MeSH terms: Enterovirus/isolation & purification*
Development of a sandwich ELISA to detect virus-like-particles in enterovirus A71 vaccines

Lim PY, Cardosa MJ

J Virol Methods, 2019 08;270:113-119.
PMID: 31100287 DOI: 10.1016/j.jviromet.2019.05.005

The goal of this paper was to develop a sandwich ELISA that can detect intact human enterovirus A71 (EV-A71) virus-like particles (VLPs) in vaccines. This assay specifically detected EV-A71 viruses from different sub-genogroups as well as EV-A71 VLPs, and treatment of VLPs with high heat and low pH reduced or completely abolished detection of the VLPs suggesting that the ELISA detected assembled particles. Using a purified VLP as a reference standard, a quantitative sandwich ELISA (Q-ELISA) was established which was used to monitor the yield and purity of the VLPs during manufacturing. Coupled with immunogenicity studies, the Q-ELISA was used to evaluate the performance of the VLPs and formalin-inactivated EV-A71 vaccine. This assay has the potential to play an important role in the development of an efficient process to produce and purify the VLPs and in examining the quality of EV-A71 vaccines.

Matched MeSH terms: Enterovirus/isolation & purification*
Molecular epidemiology of human enterovirus 71 strains and recent outbreaks in the Asia-Pacific region: comparative analysis of the VP1 and VP4 genes

Cardosa MJ, Perera D, Brown BA, Cheon D, Chan HM, Chan KP, et al.

Emerg Infect Dis, 2003 Apr;9(4):461-8.
PMID: 12702227

This study provides a comprehensive overview of the molecular epidemiology of human enterovirus 71 (HEV71) in the Asia-Pacific region from 1997 through 2002. Phylogenetic analysis of the VP4 and VP1 genes of recent HEV71 strains indicates that several genogroups of the virus have been circulating in the Asia-Pacific region since 1997. The first of these recent outbreaks, described in Sarawak (Malaysian Borneo) in 1997, was caused by genogroup B3. This outbreak was followed by large outbreaks in Taiwan in 1998, caused by genogroup C2, and in Perth (Western Australia) in 1999, where viruses belonging to genogroups B3 and C2 cocirculated. Singapore, Taiwan, and Sarawak had HEV71 epidemics in 2000, caused predominantly by viruses belonging to genogroup B4; however, large numbers of fatalities were observed only in Taiwan. HEV71 was identified during an epidemic of hand, foot and mouth disease in Korea; that epidemic was found to be due to viruses constituting a new genogroup, C3.

Matched MeSH terms: Enterovirus/isolation & purification
Fulltext Broad reactive monoclonal antibodies for rapid identification of enteroviruses show cross-reactivity with chikungunya virus infected cells

Khairul AH, Chem YK, Keniscope C, Rosli J, Hassan S, Mat J, et al.

Malays J Pathol, 2010 Jun;32(1):49-52.
PMID: 20614726 MyJurnal

In the past decade, enterovirus 71 (EV71) and chikungunya (CHIK) virus have re-emerged periodically causing serious public health problems in Malaysia, since their first emergence in 1997 and 1998 respectively. This study demonstrates that CHIK virus causes similar patterns of cytopathic effect in cultured Vero cells as some enteroviruses. They also show positive cross-reaction on direct immunofluorescence staining using monoclonal antibodies meant for typing enteroviruses. Without adequate clinical and epidemiological information for correlation, CHIK virus isolated from patients with acute febrile rash can be wrongly reported as untypeable enterovirus due to its cross-reactivity with commercial pan-enterovirus monoclonal antibodies. This is due to the diagnostic laboratory being unaware of such cross-reactions as it has not been reported previously. Final identification of the virus could be determined with specific antibodies or molecular typing using specific oligonucleotide primers for the CHIK virus.

Matched MeSH terms: Enterovirus/isolation & purification*
Fulltext Laboratory acute flaccid paralysis surveillance in Malaysia: a decade of commitment to the WHO global polio eradication initiative

Saraswathy TS, Khairullah NS, Sinniah M, Fauziah MK, Apandi MY, Shamsuddin M

Southeast Asian J. Trop. Med. Public Health, 2004 Jun;35(2):421-4.
PMID: 15691149

The Institute for Medical Research, Malaysia, was designated the National Reference Laboratory for Poliomyelitis Eradication (NRLPE) in 1992. Since then, our Polio Laboratory has collaborated actively with the Disease Control Division, Ministry of Health (MOH), Malaysia and WHO towards achieving polio eradication. Since 1992, the NRLPE has investigated 1,063 stool specimens from 641 acute flaccidparalysis (AFP) cases. One hundred and one enteroviruses were isolated from these specimens. Positive cell cultures were confirmed by microneutralization assay using standard WHO antisera. All enterovirus isolates were sent to the Victorian Infectious Disease Reference Laboratory in Melbourne, Australia, for further identification and poliovirus intratypic differentiation. Thirty-one out of these 101 virus isolates (30%) were polioviruses (PV) and the remaining 70 (70%) were non-polio enteroviruses (NPEV) which included coxsackie B viruses, echoviruses and enterovirus 71. Three of the poliovirus isolates were wild-type polioviruses isolated in 1992 which were the last wild-type polioviruses isolated in Malaysia. The rest were vaccine-related Sabin-like strains. Monthly reports of the virological investigation of AFP cases are sent to WHO and to the MOH, AFP control committee. The NRLPE continues to play an integral role in AFP surveillance and is committed to the WHO's goal of global polio eradication by the year 2005.

Matched MeSH terms: Enterovirus/isolation & purification
Isolation of subgenus B adenovirus during a fatal outbreak of enterovirus 71-associated hand, foot, and mouth disease in Sibu, Sarawak

Cardosa MJ, Krishnan S, Tio PH, Perera D, Wong SC

Lancet, 1999 Sep 18;354(9183):987-91.
PMID: 10501361

In mid-1997, several children died in Sarawak, Malaysia, during an epidemic of enterovirus-71 (EV71) hand, foot, and mouth disease. The children who died had a febrile illness that rapidly progressed to cardiopulmonary failure and the cause was not satisfactorily resolved. We describe the isolation and identification of a subgenus B adenovirus from the children who died.

Matched MeSH terms: Enterovirus/isolation & purification
Molecular detection of enteroviruses from an outbreak of hand, foot and mouth disease in Malaysia in 1997

Abubakar S, Chee HY, Shafee N, Chua KB, Lam SK

Scand. J. Infect. Dis., 1999;31(4):331-5.
PMID: 10528868

Enterovirus 5'UTR sequences were detected by RT-PCR in 22 out of 47 suspected hand, foot and mouth disease (HFMD) patients during an outbreak of the disease with incidences of fatal brainstem encephalomyelitis in Malaysia in 1997. Genetic and phylogenetic analyses of the isolates 5'UTR sequences suggest the presence of predominantly enteroviruses with high sequence similarities to Echovirus 1 and Coxsackievirus A9 in the Malaysian peninsula. No fatal cases, however, were associated with these isolates. The remaining isolates, including all (4/4) isolates of the fatal cases from the Malaysian peninsula and Sarawak shared very high sequence identity with enterovirus 71MS (EV71). These findings suggest that several enteroviruses were circulating in Malaysia during the outbreak period, with only EV71 causing fatal infections.

Matched MeSH terms: Enterovirus/isolation & purification*
Genetic diversity of enterovirus 71 isolated from cases of hand, foot and mouth disease in Yokohama City between 1982 and 2000

Munemura T, Saikusa M, Kawakami C, Shimizu H, Oseto M, Hagiwara A, et al.

Arch Virol, 2003 Feb;148(2):253-63.
PMID: 12556991

Enterovirus 71 (EV71) is known as one of the major causative agents of hand, foot and mouse disease (HFMD) and is also associated with neurological manifestations such as aseptic meningitis, polio-like paralysis and encephalitis. Recently, large HFMD outbreaks, involving severe neurological complications, have been experienced in Malaysia, Taiwan and some other countries in the Western-Pacific region. To investigate the genetic diversity of EV71 isolates in a single community in Japan, nucleotide sequences of the VP4 region of 52 EV71 isolates in Yokohama City from 1982 to 2000 were determined and the phylogenetic relationship was compared with other referential EV71 strains in Japan and in the world. There were two major genotypes of EV71 in Yokohama City through the 1980's and 1990's. Six EV71 isolates in the early 1980's in Yokohama City were closely related to those from HFMD outbreaks in Japan and from outbreaks of polio-like paralysis in Europe in the 1970's. During recent HFMD outbreaks in 1997 and 2000, two distinct genotypes of EV71 were co-circulating in Yokohama City as in HFMD outbreaks in Malaysia and Taiwan. However, the genetic diversity of EV71 in Yokohama City was not directly correlated with the severity of HFMD. The results confirmed the circulation of two distinct genotypes of EV71 over the past 20 years in Japan.

Matched MeSH terms: Enterovirus/isolation & purification*
Enterovirus 71 from fatal and nonfatal cases of hand, foot and mouth disease epidemics in Malaysia, Japan and Taiwan in 1997-1998

Shimizu H, Utama A, Yoshii K, Yoshida H, Yoneyama T, Sinniah M, et al.

Jpn J Infect Dis, 1999 Feb;52(1):12-5.
PMID: 10808253

Enterovirus 71 (EV71), one of the major causative agents for hand, foot and mouth disease (HFMD), is sometimes associated with severe central nervous system diseases. In 1997, in Malaysia and Japan, and in 1998 in Taiwan, there were HFMD epidemics involving sudden deaths among young children, and EV71 was isolated from the HFMD patients, including the fatal cases. The nucleotide sequences of each EV71 isolate were determined and compared by phylogenetical analysis. EV71 strains from previously reported epidemics belonged to genotype A-1, while those from recent epidemics could be divided into two genotypes, A-2 and B. In Malaysia, genotype A-2 was more prevalent, while in Japan and Taiwan, B genotype was more prevalent. Two isolates from fatal cases in Malaysia and one isolate from a fatal case in Japan were genotype A-2. However, all isolates from three fatal cases in Taiwan belonged to genotype B. The severity of the HFMD did not link directly to certain genotypes of EV71.

Matched MeSH terms: Enterovirus/isolation & purification*
Enterovirus survey before and after poliomyelitis vaccination in Kuala Lumpur, Malaysia

Tan DS, Lam SK

Southeast Asian J. Trop. Med. Public Health, 1978 Sep;9(3):301-7.
PMID: 219550

Stool samples from healthy children mainly of the low income group aged 0 to 7 years of age from five Maternal and Child Health Centres in Kuala Lumpur were obtained for isolation of enteroviruses. The specimens were collected before and after the mass vaccination given in the face of polio type 1 epidemic which started in October, 1971. The prevelance rate of enteroviruses was 11.9% (3.0% polioviruses, 8.9% non-polio enteroviruses) before the vaccination and essentially the same after. Coxsackie A viruses predominated over the other enteroviruses in the pre- and post-vaccination phases. The highest isolation rate of enteroviruses was observed in children 0 to 2 years age. No significant differences in distribution by sex, race and month were noted. A sharp fall in the prevalence rates of total enteroviruses and polioviruses was noted shortly after the mass vaccination campaign However, the rates reverted to the pre-vaccination state during the next successive years.

Matched MeSH terms: Enterovirus/isolation & purification*
Human enterovirus 71 disease in Sarawak, Malaysia: a prospective clinical, virological, and molecular epidemiological study

Ooi MH, Wong SC, Podin Y, Akin W, del Sel S, Mohan A, et al.

Clin Infect Dis, 2007 Mar 01;44(5):646-56.
PMID: 17278054

BACKGROUND: Human enterovirus (HEV)-71 causes large outbreaks of hand-foot-and-mouth disease with central nervous system (CNS) complications, but the role of HEV-71 genogroups or dual infection with other viruses in causing severe disease is unclear.
METHODS: We prospectively studied children with suspected HEV-71 (i.e., hand-foot-and-mouth disease, CNS disease, or both) over 3.5 years, using detailed virological investigation and genogroup analysis of all isolates.
RESULTS: Seven hundred seventy-three children were recruited, 277 of whom were infected with HEV-71, including 28 who were coinfected with other viruses. Risk factors for CNS disease in HEV-71 included young age, fever, vomiting, mouth ulcers, breathlessness, cold limbs, and poor urine output. Genogroup analysis for the HEV-71-infected patients revealed that 168 were infected with genogroup B4, 68 with C1, and 41 with a newly emerged genogroup, B5. Children with HEV-71 genogroup B4 were less likely to have CNS complications than those with other genogroups (26 [15%] of 168 vs. 30 [28%] of 109; odds ratio [OR], 0.48; 95% confidence interval [CI], 0.26-0.91; P=.0223) and less likely to be part of a family cluster (12 [7%] of 168 vs. 29 [27%] of 109; OR, 0.21; 95% CI, 0.10-0.46; P

Matched MeSH terms: Enterovirus/isolation & purification
Fulltext Validation and utilization of an internally controlled multiplex Real-time RT-PCR assay for simultaneous detection of enteroviruses and enterovirus A71 associated with hand foot and mouth disease

Thanh TT, Anh NT, Tham NT, Van HM, Sabanathan S, Qui PT, et al.

Virol J, 2015 Jun 09;12:85.
PMID: 26050791 DOI: 10.1186/s12985-015-0316-2

BACKGROUND: Hand foot and mouth disease (HFMD) is a disease of public health importance across the Asia-Pacific region. The disease is caused by enteroviruses (EVs), in particular enterovirus A71 (EV-A71). In EV-A71-associated HFMD, the infection is sometimes associated with severe manifestations including neurological involvement and fatal outcome. The availability of a robust diagnostic assay to distinguish EV-A71 from other EVs is important for patient management and outbreak response.
METHODS: We developed and validated an internally controlled one-step single-tube real-time RT-PCR in terms of sensitivity, linearity, precision, and specificity for simultaneous detection of EVs and EV-A71. Subsequently, the assay was then applied on throat and rectal swabs sampled from 434 HFMD patients.
RESULTS: The assay was evaluated using both plasmid DNA and viral RNA and has shown to be reproducible with a maximum assay variation of 4.41 % and sensitive with a limit of detection less than 10 copies of target template per reaction, while cross-reactivity with other EV serotypes was not observed. When compared against a published VP1 nested RT-PCR using 112 diagnostic throat and rectal swabs from 112 children with a clinical diagnosis of HFMD during 2014, the multiplex assay had a higher sensitivity and 100 % concordance with sequencing results which showed EVs in 77/112 (68.8 %) and EV-A71 in 7/112 (6.3 %). When applied to clinical diagnostics for 322 children, the assay detected EVs in throat swabs of 257/322 (79.8 %) of which EV-A71 was detected in 36/322 (11.2 %) children. The detection rate increased to 93.5 % (301/322) and 13.4 % (43/322) for EVs and EV-A71, respectively, when rectal swabs from 65 throat-negative children were further analyzed.
CONCLUSION: We have successfully developed and validated a sensitive internally controlled multiplex assay for rapid detection of EVs and EV-A71, which is useful for clinical management and outbreak control of HFMD.

Matched MeSH terms: Enterovirus/isolation & purification*

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