摘要
目的 通过病原学检测鉴别诊断严重急性呼吸综合征 (SARS)。方法 对 1例没有明确SARS接触史 ,入院时临床诊断疑为“支原体肺炎”的患儿进行病原学鉴别诊断。 (1)对患儿的咽拭子标本进行常见呼吸道病毒 (包括呼吸道合胞病毒、甲、乙型流感病毒、腺病毒和Ⅰ、Ⅱ、Ⅲ型副流感病毒 )的抗原检测。 (2 )用RT PCR进行人类偏肺病毒、肠道病毒和鼻病毒的检测。 (3)采用巢式RT PCR方法 ,进行SARS病毒基因检测。根据WHO在网上公布的SARS冠状病毒复制酶基因 1b区合成3对引物 ,其中 1对为所有冠状病毒所保守的 ,用来进行第一次PCR ,另 2对为SARS冠状病毒所特异的 ,分别用于第二次PCR ,这 2对引物可分别扩增 36 8和 348bp的基因片段。结果 患儿咽拭子标本 7种常见呼吸道病毒和人类偏肺病毒、肠道病毒和鼻病毒的检测结果均为阴性 ,而用不同引物对检测 ,SARS冠状病毒基因均为阳性。经测序显示 ,该基因片段与GenBank已公布的 17株SARS冠状病毒的序列同源性为 10 0 % ,而与人冠状病毒的标准株有很低的同源性。同时该患儿的恢复期血清SARS冠状病毒特异性IgM和IgG均为阳性。结论 从患儿标本中未检测到常见的 7种呼吸道病毒、人类偏肺病毒、肠道病毒和鼻病毒 ,而检测到了SARS冠状病毒。经病原学检测 ,确诊患儿为SARS。
A Special 'Fever and Cough' Clinic was set up at the Children′s Hospital Affiliated to Capital Institute of Pediatrics for children with symptoms of fever and cough in late April when the severe acute respiratory syndrome (SARS) epidemic was at its peak in Beijing to separate the children with fever from others during their visit to the Outpatient Department.Objective For patients with fever, normal or low count of white blood cell and with suspected pneumonia suggested by X ray, it was urgent to determine the etiological agents of the diseases before they were admitted to the hospital.Methods Throat swabs or nasopharyngeal aspirate specimens were collected from those patients and common respiratory virus antigens including influenza virus A and B, respiratory syncytial virus, adenovirus, parainfluenza virus types Ⅰ, Ⅱ, and Ⅲ were tested by indirect immunofluorescent assay. The patients with atypical pneumonia diagnosed by X ray and evidences of common respiratory virus infection were admitted to the regular ward for children with respiratory diseases. Children with pneumonia demonstrated by X ray and negative for common respiratory viruses were admitted to the isolated ward for suspected SARS patients for the first step and further viral etiological studies were requested. RT PCR was performed for those patients to detect gene fragments of human metapneumovirus (HMPV), rhinovirus (RhV) and enterovirus (EV) in their specimens. Nested RT PCR was also developed to detect SARS coronavirus gene fragment from the specimens. Primer sequences for SARS virus detection with the PCR were selected according to the primer sequences published online by WHO on April 18, 2003. All the primers derived from the sequence at the 1b frame of coronavirus replicase gene and products with a size of 368 or 348bp were expected with 2 different primer pairs. Results Amplicons with the sizes of 368 bp and 348 bp were obtained from a throat swab specimen collected from a 17 years old girl, who was admitted to the isolated ward because of high fever (39.5℃) for 7 days, cough for 2 days, low WBC count, and pneumonia shown by X ray when she visited the 'Fever and Cough' Clinic, and without known history of contact with probable SARS patient. Antigens for the common respiratory viruses were all negative, RT PCR for HMPV, RhV and EV were also negative while RT PCR with different primer pairs for SARS virus were all positive which indicated that SARS coronavirus gene fragments were amplified from the specimen from this girl. The amplified fragment with a size of 368 bp was sequenced and the sequence was compared with those in the GenBank. The sequence shared 100% homology with the sequences from 1b frame of replicase genes from all 17 of SARS coronaviruses published in the GenBank so far, and shared very low homology with 2 reference strains of human coronavirus as well as other animal coronaviruses. The serum collected before her discharge from the hospital (19 days after the onset of the disease) showed SARS specific IgM and IgG antibodies. Conclusion These data indicate that the patient was a confirmed case of SARS. It is of great importance to differentiate SARS patients from those infected with common respiratory viruses during SARS epidemic, especially for pediatric patients, because most of the patients visiting the outpatient department present with the symptoms of fever, cough and normal WBC count. The data mentioned above indicate that antigen and gene detections for those common respiratory viruses are useful methods for the differentiation to avoid the spread of SARS.
出处
《中华儿科杂志》
CAS
CSCD
北大核心
2003年第9期641-644,U001,共5页
Chinese Journal of Pediatrics
基金
国家自然科学基金资助项目 (3 0 3 40 0 19)
