| Objective:Viral encephalitis is a serious neurological disease with high morbidity and high mortality.It is one of the important causes that endanger children’s health,disability and even death.The pathogens causing viral encephalitis are complex and diverse,there are about 100 kinds of viruses,including herpes virus,enterovirus,EB virus,adenovirus,arbovirus and so on.Herpes virus and enterovirus are the main pathogens of sporadic viral encephalitis and epidemic viral encephalitis.Analysis of the etiology of viral encephalitis contributes to its prevention and treatment.This study collected 297 cerebrospinal fluid specimens of patients diagnosed with viral encephalitis from a children’s hospital in Yunnan from March 2009 to February 2010,we conduct etiological and molecular epidemiological analysis,and provides experimental data for the prevention and control of viral encephalitis in Yunnan Province.Methods:1.Real-time fluorescent quantitative PCR was used to detect the cerebrospinal fluid samples of children with herpes simplex virus type Ⅰ/Ⅱ,varicella zoster virus,EB virus,cytomegalovirus,human herpesvirus 6 virus and enterovirus nucleic acid.2.Nested RT-PCR was used to amplify and sequence samples that tested positive for enterovirus,and the serotype of enterovirus was determined by BLAST comparison.3.Simultaneously analyzed the patient’s general condition and clinical characteristics with SPSS software.4.Virus isolation by RD cells.5.The entire CVB5 gene sequence was analyzed using MEGA 7,simplot 3.5.1 and RDP3 software.Results:1.The major pathogens causing viral encephalitis were detected as 25 cases of enteroviruses(25/35.71.4%),5 cases of varicella zoster virus(5/35,14.3%),3 cases of herpes simplex virus(3/35,8.6%,2 cases of type 1,2 cases of type 1),Epstein-Barr virus in 2 cases(2/35.2.9%),human herpesvirus 6 in 1 case(2/35,2.9%),the total detection rate is 11.8%(35/297).2.The enterovirus serotype causing viral encephalitis is Coxsackievirus B5(10/25,40.0%),Echovirus 30(4/25,16.0%),Coxsackievirus B1(3/25,12.0%),Coxsackievirus A9(2/25,8.0%)and Ecovirus 9(2/25,8.0%).3.The most common symptoms in the clinical characteristics analysis were fever(92.9%),neck headache(81.8%),stiffness(72.4%),and vomiting(65.7%).Statistics show that there is no corresponding relationship between the clinical characteristics of enterovirus-associated encephalitis and non-enterovirus-associated encephalitis,and the P value is greater than 0.05,and there is no statistical significance.4.A positive isolate(KMA2/YN/CHN/2010,)was identified and found to be more than 85%homologous to CVB5,according to the enterovirus serotyping criteria,belonging to CVB5.5.The full-length genome of KMA2/YN/CHN/2010 strain is 7395bp,the nucleotides and their amino acids were most homologous to A210/KM/09(98.8%and 96.1%);the nucleotide and amino acid homology with other CVB5 strains were 79.6%-93.1%and 93.1%-98.1%.Phylogenetic analysis showed that KMA2/YN/CHN/2010 was closely related to the Chinese strains and had a long relationship with other foreign CVB5 isolates,belonging to different branches.The results of the recombinant analysis showed that KMA2/YN/CHN/2010 may be a recombinant strain.Conclusion:From March 2009 to February 2010,the pathogens causing viral encephalitis were HSV,VZV,Epstein-Barr virus,CMV,HHV-6,and EV,the most important of which was EV,and the serotype in EV was CVB5,E30,CVB1,CVA9 and E9,it is consistent with the genotype of the current Chinese epidemic strain;the recombination of the isolate KMA2/YN/CHN/2010 may have occurred.For the future prevention of viral encephalitis and related research provides an effective reference and laboratory information. |
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