Incorrect identification of recent Asian strains of Coxsackievirus A16 as human enterovirus 71: Improved primers

The EV71 culture supernatant was concentrated 10-fold with a Amicon 100K centrifugal filter (Millipore). After 24 and 48 h of incubation, the medium was removed and 50 μL 10% trichloroacetic acid (4°C) was added in each well for 1 h. Background of the EV71 viruses isolated and genotyped in Ho Chi Minh City, Vietnam, 2011. Proportions of complicated infections were 29.4% (5/17) (95% CI 6–52.8%) for C4a genotype and 16.9% (11/65) (95% CI: 7.6–26.3%) for B5 genotype without statistical significance. Constructs including the VP1 gene region targeted by Tan et al. Informed consent was obtained from each child’s accompanying parent or guardian. Of the 37 EV-screening PCR positive specimens, 21 (56.8%) were positive by RT-PCR amplification of the VP1 region with primers 188 and 222.

EV-A71 infected mouse tissues [21] were used as positive controls. Consider the distribution of incidences in medical institutions. 2C). This IFN-γ skewing Th1 response is interesting and unlike other finding with subunit vaccines formulated with alum [29]. A discrete phylogeographic model was used to analyze virus spread between countries [15]. Briefly, 600 µl of lysis buffer was added to 25 mg of homogenized tissue. Neutralizing antibody titers below 1∶8 were assumed to be 1∶4 and were log-transformed to calculate the geometric mean titer (GMT) and 95% confidence intervals.


The 50% effective concentrations (EC50s) were calculated according to the formula EC50 = [(Y − B)/(A − B)] × (H − L) + L], where Y represents half of the mean optical density at 570 nm (OD570) of the cell control without the compound, B represents the mean OD570 of wells with the compound dilution nearest to and below Y, A represents the mean OD570 of wells with the compound dilution nearest to and above Y, and L and H are the compound concentrations at B and A, respectively. One could almost say that there is a continuum between naked replicating transmissible RNAs, depender packaged transmissible RNAs, simple viruses and complicated viruses like the herpesviridae and poxviridae. The extracted RNA was analyzed for the viral load using the TaqMan quantitative RT-PCR for amplification of the EV71 VP1 gene as described previously [26]. Hexon-specific PCR primers HexU and SP70r, Hu1 and Hr1 confirmed incorporation of coding regions for SP70 epitope inserts at the hexon HVR1, HVR2, and HVR7 sites. Stool specimens were collected from each child hospitalized with clinical diagnosis HFMD. During this period, most patients were less than 15 years of age (95.4 %); in particular, 476 (81.4 %) cases occurred in young children less than 5 years age. The Chinese strains displayed together an overall nt difference ≤2.7% (Table 1).

Proteins bound to biotinylated RNA were fractionated by 10% SDS-PAGE, and specific proteins were detected by Western blotting analyses as described below. The CVA16 virus culture supernatant was harvested from the bioreactor culture. A whole genome BLAST search and a phylogenetic tree with available EV full-length genomes (http://www.picornaviridae.com/enterovirus/hev-a/hev-a_seqs.htm) revealed that this strain clusters with other EV71 serotypes within the human EV-A species. This strain was also used for the construction of pseudotypes and for propagation and titration on RD cells. Several proteins that modulate the EV71 IRES and function as ITAFs are known. The genus Enterovirus of the family Picornaviridae contains many important pathogens for humans and animals. The 2011 samples fell within the same clade and 2012 samples fell within a different clade.

Recent studies have explained this paradox. Competing interests: The authors have declared that no competing interests exist. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The peak incidence density of disease caused by EV71 occurred from April to June. Viral replication was determined by quantitative real-time PCR (qRT-PCR). No epigenetic array studies of KD have previously been published. National Academies of Sciences, Engineering, and Medicine (US); Health and Medicine Division (US); Division of Behavioral and Social Sciences and Education (US); Board on Global Health (US); Board on Children, Youth, and Families (US); Forum on Investing in Young Children Globally (US).

All the patients were in the age group of 3-5 years from different schools. The mean epidemic area proportion in the 7 years was 6.0% for influenza and 0.2-7.4% for pediatric diseases. Hand, foot, and mouth disease should not be confused with foot-and-mouth disease of cattle, which is rare in human beings and is not caused by Coxsackie virus. These events triggered a study on the epidemiology of EV71 in The Netherlands. Nucleoside-based inhibitors have broad-spectrum activity but often exhibit off-target effects. EV71 is known to be a causative agent of HFMD, herpangina, aseptic meningitis, paralysis, and meningoencephalitis [1]. Housakos admitted to soliciting a donation from his former BPR boss at the time, but he said he did not have any formal role in the Montreal event.

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