| [Background]Human enterovirus (EV) consists of many serotypes. Infection of enterovirus is usually asymptomatic, however, it can also cause a variety of clinical disorders, such as acute flaccid paralysis (AFP), meningitis or encephalitis, hand-foot-and-mouth disease (HFMD), acute hemorrhagic conjunctivitis, myocarditis, flu-like symptoms, etc. EV is an important human pathogen.The genome of EV consists of a single-stranded positive-sense RNA of approximately7500nucleotides. It has a single open reading frame (ORF) flanked by5’and3’nontranslated regions (NTR). The ORF consists of a structural proteins coding region (PI) and two structural proteins coding regions (P2and P3). In1999, the molecular typing method based on VP1sequence was introduced by Oberste et al, and then more than40newer EVs were identified which were untypeable by serological method. Most complete genome sequences of newer EVs prototype had been available by the biological typing method. However, complete genome sequences of other newer EVs strains were insufficient. The evolution of EVs is active, and point mutation and recombination were found to took place in all of the newer EVs discovered to date, such as EV-A76, EV-B74, EV-C96, etc. Analysises of the complete genome sequence of EVs can investigate characters of recombination and evolution, which fulfills the Genbank and provides information for the analysis of the virus transmission in the district and the whole world.Although discovered lately, the newer EVs had been associated with outbreaks of multiple diseases. HFMD pandemics caused by EV-A71in East and Southeast Asia had become a major global public health problem. The outbreak of meningitis in India and Spain in2006was reported to be associated with EV-B75, EV-A76and EV-A89. Some symptoms caused by newer EVs infection were unable to diagnose, such as fever, gastrointestinal/respiratory symptoms. A variety of syptoms could be caused by EVs, and it leads to difficulties to take prevention and early treatment measures. The antibody status and its dynamic changes can be investigated by the seroepidemiology studies of EVs. To date, the epidemiology study of EVs was scarely.[Objectives]1. On the basis of prophase researches, to further study the complete genome sequence characterizations of the newer EVs strains isolated in Shandong (EV-A71C2subtype, EV-B74, EV-B80, and EV-B87).2. To explore levels of neutralizing antibody of newer EVs (EV-A71C4subtype, EV-B80, EV-A90) in healthy population.[Methods]1. Complete genome sequencing was performed on EV-A71C2subtype, EV-B74, EV-B80, and EV-B87Shandong isolates.2. Homologous comparison on the complete genome and amino acid sequences between Shandong isolates and prototypes was performed using BioEdit7.1.7.3. Phylogenetic trees on P1, P2, and P3sequences were constructed using neighbor-joining method in Mega4.0.4. Recombination in the genome of Shandong newer EV strains was analyzed using Simplot3.5.1.5. The level of neutralizing antibodies of newer EVs Shandong isolates (EV-A71C4subtype, EV-B80, and EV-A90) was detected and analyzed.[Results]1. The complete genome characterization analysis of Shangdong newer EVsResults of complete genome characterization analysis showed that the recombination event of EV-B74, EV-B80, and EV-B87Shandogn isolates occured in the non-structural protein coding region (P2and P3). The sequence similarities between EV-B74Shandong isolate and E13,E19prototypes, in the non-structural protein coding region were relatively higher. Similar results were found on EV-B80and EV-B87Shandong isolates (with E19, E27, and E13prototypes/CVB2, EV-B85, and EV-B98prototypes, respectively). Comparied with the prototype strain, an insertion of36nucleotides was observed in the VP1coding region of EV-B80Shandong isolate. The nucleotide insertion point is just the region varing among different gene types.EV-A71C2subtype Shandong isolate96200formed in one lineage with EV-A71C2subtype Australia isolate7F/AUS/6/99, Taiwan isolates NCKU9822, Tainan/5746/98/TW/1998, and pinf7-54A in the tree on P1, P2, and P3coding regions, respectively. However,96200clustered with EV71C2-like subtype Taiwan isolate2008-00643in the tree only on P1coding region. The similarity plot and bootscanning analysis with other HEV-A prototype and EV-A71strains suggested that sequence similarities between96200and Australia isolate7F/AUS/6/99, Taiwan isolates NCKU9822, Tainan/5746/98/TW/1998, and pinf7-54A were more than90%in the P1, P2, and P3coding regions. However, the similarities between96200and EV71C2-like subtype Taiwan isolate2008-00643, EV-A71prototype strain BrCr, EV-A71Shandong local strain JN01/10were much high in the P1coding region than P2and P3coding regions.2. Seroepidemilogy analysisThe serum samples used for the epidemiology study come from samples collected for AFP surveillance in2010. The sample size was390, which were devided into10age groups. The neutralizing antibody positive rates of EV-A71, EV-B87, and EV-A90in the population were46.0%,47.0%, and8.8%. Close relationship appeared between the neutralizing antibody positive rates of EV-A71, EV-B87, EV-A90and the age. Antibody positive rate of them displayed low value in the age group of7months to2years, and increased gradually after then.The Geometric Mean Titer (GMT) of EV-A71, EV-B87, and EV-A90in the population were1:5.20,1:4.02, and1:1.49. The lowest GMT value of them appears in the group on7months to2years, and the highest GMT value appeared in5to14years old. Antibody titers≥1:256showed in the3.5%of the population for EV-A71, which fitted the aboratory diagnostic criteria of HFMD cases. The rate of antibody titers≥1:256displayed in0.3%for EV-A90, and0for EV-B87in the population. [Conclusions and suggestions]1. In this study, the complete genome characterization analysises of EV-A71C2subtype, EV-B74, EV-B80, EV-B87Shandong isolates were conducted. All the complete genome sequences were first reported in China except EV-B74.2. EV-A71C2subtype isolates were more conserved in1996-1999, while no significant recombination evidence was found according to the Genbank data available. Taiwan EV-A71C2-like subtype2008-00643isolated in2008was recombined while complared with EV-A71C2subtype isolated in1996-1999.3. Recombination occured in the P2and P3non-structural protein coding regions of EV-B74, EV-B80and EV-B87. The rusult of genome comparation for EV-B80showed that insertions and deletions were also the evolution mechanisms on EV coding regions.4. Antibody positive rates of EV-A71, EV-B87and EV-A90were relatively lower among the age group7months-2years, which may be the susceptible population.5. Cross-antibodies react may occure on EV-B87with some other enterovirus popular among the population. Large-scale epidemic would not be caused by EV-B87because of the natural immune barrier.6. The antibody positive rates of EV-A90in the population was low. However, strains isolate rates in the global and domestic was relatively higher, involving a wide geographical scope. Surveillance on it should be strengthened in the future.7. GMT of EV-A71, EV-B87and EV-A90were relatively higher among the school age group of5～14years old, which implied that they were the susceptible population. Prevention and immunization measures should be taken among the school age group. |
PDF Full Text Request