ABSTRACT
Interpretation and consideration of “Substitution of in vivo method(s) by in vitro method(s) for the quality control of vaccines” in General Texts of European Pharmacopoeia@#Potency is a critical quality attribute for controlling relevant biological properties and batch consistency of vaccines.The methods can be divided into in vivo and in vitro methods according to whether animals are used.The in vivo methods are large consuming of animals and time,as well as have large variant detection results.In contrast,the in vitro alternative methods have been the hotspot of research due to their simple operations,in line with 3Rs principles,and more stable results.However,owing to the complexity of experimental design and the lack of corresponding guidance,the research progress of alternative methods is slow.Recently,“Substitution of in vivo method(s) by in vitro method(s) for the quality control of vaccines” was adopted in the European Pharmacopoeia(10th Edition),which clarifies the critical points of consideration for substitution.This paper interprets the chapter and puts forward some thoughts on that in China,which is expected to speed up the alternative methods research and improve the ability of vaccine quality control and supervision in China.
ABSTRACT
<p><b>BACKGROUND</b>Enterovirus 71 (EV71) and coxsackievirus A16 (Cox A16) are major causative agents for hand, foot and mouth disease (HFMD). Studies indicate that the frequent HFMD outbreaks result in a few hundreds children's death in China in recent years. The vaccine and other research for HFMD need to be developed urgently.</p><p><b>THE AIMS OF OUR STUDY WERE</b>to explore dynamic development of mother-source neutralizing antibodies against EV71 and Cox A16 in infants from Jiangsu Province, China, and to provide the fundamental data for further establishing of corresponding immunization course.</p><p><b>METHODS</b>Peripheral blood samples were collected from 133 of parturient women once immediately before delivery and their infants at two and seven months of age. Method of micro-dose cytopathogenic effect was used to measure neutralizing antibodies against EV71 and Cox A16, respectively.</p><p><b>RESULTS</b>Seropositive rates of anti-EV71 and anti-Cox A16 in prenatal women were 79.7% (106/133) and 92.5% (123/133), respectively; geometric mean titers (GMTs) were 29.0 and 61.9; 75.9% (101/133) prenatal women were both positive in anti-EV71 and anti-Cox A16; seropositive rates of anti-EV71 and anti-Cox A16 were 25.6% (34/133) and 38.3% (51/133) in infants at two months of age; GMTs were 12.3 and 18.0, respectively. GMTs of anti-EV71 were significantly higher for infants at seven months (82.6) compared with that at two months (P < 0.05), showing infants had inapparently infected by EV71 during two to seven months. Although only one offspring (0.75%) at seven months was found having anti-Cox A16 transfered from maternal, this observation suggested no maternal antibody may remain in infants at seven months.</p><p><b>CONCLUSIONS</b>The prevalence of EV71 and Cox A16 were relatively high in Jiangsu Province. Bivalent vaccine against both EV71 and Cox A16 should be developed, and the ideal time point for prime immunization for infants is around 2-5 months of age.</p>
Subject(s)
Female , Humans , Infant , Infant, Newborn , Antibodies, Neutralizing , Blood , Allergy and Immunology , Cells, Cultured , Enterovirus , Allergy and Immunology , Enterovirus A, Human , Allergy and Immunology , Hand, Foot and Mouth Disease , Allergy and Immunology , VirologyABSTRACT
Objective To investigate the characterization of the complete genome of EV71 in Beijing, 2008 and to provide basis for selecting appropriate virus strain to develop vaccine. Methods 12 throat swab samples were collected from children with hand-foot-mouth disease (HFMD). One sample named 08YM-3 was cultured and isolated in vero cells. Viral RNA was extracted and carried out by RT-PCR and 5' , 3' rapid amplification of eDNA ends (RACE) to obtain the sequence from 08YM-3. PCR products were cloned and analyzed. Nucleotide identity between sequences was calculated and sequence alignments were made to generate phylogenetic trees using MegAlign in DNAStar. Results 3 clones were constructed that covered EV71 complete genome. Data from sequences analysis showed that this viral strain named BJ08 shared 95.6%-96.7%, 88.3%-96.1%,78.1%-94.0%,90.8%-94.6%, 85.9%-94.1% and 90.9%-93.9% in 5' UTR, PI, P2, P3, 3' UTR region and complete genome with C4 subtype, respectively. B J08 showed low nucleotides identity (<90%) with other subtypes. Phylogenetic trees established from alignment of the complete genome and VPI region indicated that B J08 belonged to C4 subtype. BJ08 and C4 subtype strains shared the same amino acids in 6 sites in VP1 region, which were associated with EV71 subtype. There was no mutation in VP1 antigen epitope (92-107aa). Conclusion This BJ08 strain belonged to C4 subtype. Further study on EV71 complete genome would have great significance for vaccine research.