Prokaryotic expression and polyclonal antibody preparation of human adenovirus type 7 DNA binding protein / 中华预防医学杂志

Objective: To express DNA-binding protein (DBP) of human adenovirus (HAdV) type 7 using the prokaryotic expression system, and product anti-HAdV-7 DBP rabbit polyclonal antibody. Methods: The HAdV-7 DBP gene was synthesized and cloned into prokaryotic expressing vector pET30a, and the recombinant plasmid was transformed into E. coli BL21 (DE3) competent cell. The recombinant protein DBP was expressed by induced Isopropyl-beta-D-thiogalactopyranoside (IPTG) and purified with Ni-NTA affinity column. The titer of anti-DBP polyclonal antibody produced in immunized rabbit was measured by indirect ELISA, and the specificity of the antibody was identified by Western blotting and indirect immunofluorescence assay (IFA). In addition, purified rDBP was used as coating antigen for indirect ELISA assay to detect specific IgM and IgG antibodies against DBP in the serum of children infected with HAdV. Results: The HAdV-7 DBP plasmid was constructed successfully. The purified recombinant DBP was more than 95% after purification. The titer of polyclonal antibody was 1∶1 024 000. The polyclonal antibody showed high specificity in vitro using Western blotting and IFA. The positive rate of specific anti-DBP IgM and IgG antibody in acute-phase serum samples collected from children infected with HAdV were 50.0% (19/38) and 63.2% (24/38), respectively, using indirect ELISA. Conclusion: In summary, the HAdV-7 rDBP is expressed using prokaryotic expression system, and the recombinant HAdV-7 DBP protein and the anti-DBP rabbit polyclonal antibody with high titer are prepared.

Clinical characteristics of community-acquired influenza virus pneumonia in hospitalized children / 中华实用儿科临床杂志

Objective To analyze the clinical characteristics of community-acquired influenza virus pneumonia in hospitalized children and improve the clinicians' understanding level of this disease.Methods Data of 70 cases with community-acquired influenza virus pneumonia admitted to the Respiratory Department and Infectious Disease,Beijing Children's Hospital,Capital Medical University,from November 2009 to April 2018 were collected and the clinical characteristics were analyzed.Results Of the 70 cases,61 cases(89.7％) were discharged after improvement.The median age was 3.5 years old,and 50 cases(71.4％) were 0 to 5 years old.There were 29 cases with severe influenza pneumonia,41 cases with mild influenza pneumonia,3 cases died,and 19 cases (27.1％) had underlying diseases.Sixty-four cases (91.4％) were hospitalized in winter and spring.The first symptoms were mainly fever in 64 cases (91.4％) and cough in 65 cases (92.9％),and temperatures were mostly from 39.1 ℃ to 41.0 ℃.Lung auscultation was dominated by moist rales (30 cases,58.8％) and wheezing (8 cases,15.7％).There were many complications of influenza virus pneumonia,including 19 cases with myocardial injury,11 cases with liver function injury,4 cases with toxic encephalopathy,3 cases with electrolyte disturbance,2 cases with multiple organ failure,2 cases with hemophagocytic syndrome,and 1 case with septic shock.Chest radiographic results reveal bilateral inflammation in 40 children (57.1％),prodominatly in lower lobe lesions (39 cases).The common changes were patchy shadow,interstitial parenchymal lesion,ground glass shadow,and pleural effusion.Forty-seven children (67.1％) were infected by influenza A,and 23 children(32.9％) were co-infected.The percentage of severe cases with underlying diseases (68.4％) was significantly higher than that in children without chronic diseases (31.4％),the difference was statistically significant (x2 =7.830,P =0.005).The increase rate of C reaction protein (CRP) in severe cases (54.3％) was significantly higher than that in mild cases (28.6％),the difference was statistically significant (x2 =4.769,P =0.029).Conclusions Community-acquired influenza virus pneumonia in children mainly occurs in winter and spring.It is more common seen in children under 5 years of age.The main clinical manifestations of community-acquired influenza virus pneumonia are high fever and cough,extrapulmonary complications are more common.Most children have moist rales and showed bilateral inflammation and lower lobe lesions in chest radiography.Children with underlying diseases are more likely to develop severe influenza virus pneumonia.Elevated CRP is associated with severe influenza virus pneumonia.Most patients have a good prognosis,but there are still cases of death.

Etiology and clinical epidemiological characteristics of bronchiolitis / 中国实用儿科杂志

Bronchiolitis is an acute lower respiratory tract infection in early childhood. Respiratory syncytial virus(RSV)is the most common etiology of bronchiolitis. Infections with Mycoplasma pneumoniae,Chlamydia pneumonia and Chlamydia trachomatis can also cause bronchiolitis. Bronchiolitis mainly occurs in infants under 2 years of age,with a peak age of 2 to 6 months. Bronchiolitis is the most common reason for hospitalization in infants,and infants under 6 months of age have the highest hospitalization rate. Since bronchiolitis is mainly caused by RSV infection,its seasonal characteristics are similar to those of local RSV infection. Bronchiolitis is a disease with a high incidence but a low mortality rate. Being less than 3 months,prematurity,congenital heart disease and so on are risk factors of severe bronchiolitis.

Laboratory diagnostic methods for pediatric respiratory virus infection and their rational application / 中国实用儿科杂志

Viruses are the main causes of respiratory tract infection in children. Early,rapid and accurate diagnosis of pathogen in respiratory tract infection can avoid antibiotic abuse.There are advantages and disadvantages of different laboratory diagnostic methods for respiratory virus infections. Thus,appropriate choice of diagnostic methods and correct interpretation of the test results can help clinicians to make accurate pathogen diagnosis.

Characterization of BZLF1 gene and its promoter Zp of EBV strains in children with EBV-associated diseases in recent 5 years in Beijing area / 病毒学报

This study aims to investigate the genetic characteristics of BZLF1 gene and its promoter Zp of the epidemic strains in children with primary Epstein-Barr virus (EBV)-associated diseases. Total DNA was extracted from the peripheral blood of 134 children with EBV-associated infectious mononucleosis (EBV-IM) and 32 children with EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH) who were admitted to Beijing Children's Hospital from 2006 to 2011. The EBNA3C, BZLF1, and Zp genes were amplified by PCR assay. Typing of EBV was performed according to the size of the amplification product of EBNA3C gene; the amplification products of BZLF1 and Zp genes were subjected to direct sequencing, and sequence analysis was performed using BioEdit 7. 0. 9. The results were as follows: (1) EBV-1 was present in 140 samples (97.2%, 140/144) and EBV-II in 4 samples (2.8%, 4/144). (2) Three BZLF1 genotypes and their 12 subtypes (including 6 newly found subtypes) were detected in this study; there were no significant differences in the frequencies of BZLF1-A and BZLF1-B between the children with EBV-IM and EBV-HLH (P = 0.083); BZLF1-A1 was the dominant genotype in children with EBV-associated diseases; t BZLF1-A mostly had three 29-bp repeats in the first intron of BZLF1 gene, and BZLF1-B mostly had 30-bp repeats (P = 0.000), with the number of repeats varying from 1 to 13. (3) Four Zp genotypes were detected in this study, including Zp-P, Zp-V3, Zp-V4, and Zp-V1; there were no significant differences in the frequencies of these Zp genotypes between children with EBV-IM and EBV-HLH (P = 0.272, 0.252, 1.0, and 1.0, respectively). (4) The linkage analysis of BZLF1 gene and its promoter Zp showed that BZLF1-A1 was highly associated with Zp-V3 (P = 0.000), while BZLF1-B4 with Zp-P (P = 0.000); EBV-I + BZLF1 A1 was highly associated with Zp-V3 (P = 0.000), while EBV-I+BZLF1-B4 with Zp-P (P = 0.000). The conclusions are as follows: (1) BZLF1-A1 is the dominant genotype in children with EBV-associated diseases; there are mostly 29-bp repeats in the first intron of BZLF1 gene for BZLF1-A genotype and 30-bp repeats for BZLF1-B genotype. (2) Zp-P and Zp-V3 are dominant Zp genotypes of EBV in children, which shared similar detection rates. (3) BZLF1-A1 is highly associated with Zp-V3, while BZLF1-B4 with Zp-P; EBV-I+BZLF1-A1 is highly associated with Zp-V3, while EBV-I+BZLF1-B4 with Zp-P.

Genetic polymorphism of cytokine genes in Han children in Beijing area / 中华实用儿科临床杂志

Objective To investigate the genetic polymorphism of 13 cytokines genes in Han individual children from Beijing area.Methods The polymerase chain reaction-sequence specific primers (PCR-SSP) method was used to determine cytokine gene polymorphism including interleukin (IL)-1α (-889C/T),IL-1β (-511C/T,+3942T/C),IL-1 R(pst1 1970C/T),IL-1RA(mspa1 11100T/C),IL-4Ra(+ 1902G/A),IL-12 (-1188C/A),interferon (IFN)-γ(+ 874A/T),transforming growth factor (TGF)-β1 (codon10C/T,codon25 G/C),tumor necrosis factor)(TNF)-α (-308G/A,-238G/A),IL-2 (-330T/G,+ 166G/T),IL-4 (-1098 T/G,-590 T/C,-33 T/C),IL-6(-174G/C,nt565G/A),IL-10 (-1082G/A,-819C/T,-592A/C) 13 cytokines genes in 210 unrelated healthy Han individuals from Beijing area.Results (1) Cytokine genes with frequencies more than or equal to 95 percent included IL-1β +3962 C,TGF-β Codon 25 G,IL-6-174 G,IL-6 nt565 G,TNF-α-308 G and TNF-α-238 G alleles;while cytokine genes with frequencies less than or equal to 5 persent included IL-1β + 3962 T,TGF-β Codon 25 C,IL-6-174C,IL-6 nt 565 A,TNF-α-308 A and TNF-α-238 A alleles.(2) The frequency of the high-producer-type GCC haplotype in IL-10 gene was 8.27％,while the low-producer-type ATA was 59.11％.The frequency of the highproducer-type TG haplotype in the TGF-β gene was 49.77％,while the low-producer-type CC was 0.The frequencies of the high-producer-types AG and AA haplotypes in the TNF-α gene were 3.34％,while those of the low-producer-types GG and GA were 96.67％.The frequency of the high-producer-types T/G genotype in the IL-2-330 gene was 11.90％,while those of the low-producer-types T/T and T/G were 88.10％.The frequencies of the high-producer-types G/G and G/C genotypes in the IL-6-174 gene were 99.52％,while that of the low-producer-types C/C was 0.48％.The frequency of the high-producer-types T/T genotype in the INF-γ+ 874 gene was 1.90％,and the frequency of the medium-producer-types T/A in the INF-γ+ 874 gene was 22.86％,while of the low-producer-types A/A was 75.24％.(3) New genotypes were found:the frequency of IL-2(-330,+ 166) GG/GT/TT genotype was 1.90％,and the frequency of IL-2(-330,+ 166) TG/GG/GT/TT genotype was 0.48％,The frequency of IL-6(-174,nt565) CG/CG genotype was 0.48％,the frequency of IL-6 (-174,nt565) GA/GG genotype was 1.90％,The frequency of IL-6 (-174,nt565) GG/GA/CA genotype was 0.48％,The frequency of IL-10(-1082,-819,-592) GCC/ATC/ATA genotype was 0.95％,The frequency of IL-10(-1082,-819,-592) ACC/ACA/ATC/ATA genotype was 35.71％,The frequency of IL-10 (-1082,-819,-592) GCC/ACC/ACA/ATC/ATA genotype was 0.48％.Conclusion The genetic polymorphism of cytokine genes in Han individuals from Beijing area is different from other areas of China with higher frequencies of IL-10 (-1082,-819,-592) ACC/ACA/ATC/ATA genotype.

Viral etiology of acute respiratory tract infection among pediatric inpatients and outpatients from 2010 to 2012 in Beijing, China / 中华儿科杂志

<p><b>OBJECTIVE</b>Acute respiratory tract infections (ARI) are the leading cause of pediatric morbidity and mortality worldwide, particularly in developing countries. Viruses are the main pathogens of ARI in children. The purpose of the present study was to determine the epidemiologic features of respiratory viruses, including novel viruses, in outpatient and hospitalized children with ARI.</p><p><b>METHOD</b>From March 2010 to February 2012, 2066 children with ARI, including 1050 outpatients and 1016 inpatients, were involved in this study. One nasopharyngeal aspirate or throat swab specimen was collected from each patient. Reverse transcription (RT) PCRs were performed to detect common respiratory tract viruses including respiratory syncytial virus (RSV), human rhinovirus (HRV), influenza virus (IFV), parainfluenza virus (PIV) type 1-4, adenovirus (ADV), enterovirus (EV), human coronavirus (HCOV), human metapneumonia virus (HMPV) and human bocavirus (HBOV).</p><p><b>RESULT</b>At least one viral pathogen was identified in each of 1274 out of 2066 patients and the overall positive rate was 61.7%. The positive rate in inpatient (69.7%) was higher than that in outpatient (53.9%). The frequencies of detection of various viruses among in- and outpatients were different. RSV was the most prevalent virus detected among hospitalized children, followed by HRV and PIV, whereas IFV was the most frequently identified virus in the outpatient group, followed by ADV and PIV. Simultaneous detection of two or more viruses was found in 377 cases. Coinfection was more frequent in inpatients than in outpatients (30.1% vs. 6.8%, P < 0.001).</p><p><b>CONCLUSION</b>Respiratory viruses play an important role in children with ARI, especially in young children. RSV was the most prevalent virus detected among hospitalized children, whereas IFV was the most frequently identified virus in the outpatient group. Viral coinfections are frequently identified, particularly in hospitalized patients. Further studies are required to better understand the impact of coinfections in children with ARI.</p>

Characteristic of nuclear antigen 1 gene and latent membrane protein 1 gene of Epstein-Barr virus in primary EBV infection in children in Beijing area in 2005-2010 / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To analyze the characteristic of nuclear antigen 1 gene and latent membrane protein 1 gene of Epstein-Barr virus in primary EBV infection in children in Beijing area in 2005-2012.</p><p><b>METHODS</b>Polymerase chain reaction (PCR) was used to amplify the EBNA-3C, EBNA1 and LMP1 genes. The amplified products were sequenced directly and the sequences were analyzed by BioEdit 7. 0. 9 and MEGA 4. 0. 2.</p><p><b>RESULTS</b>Type A EBV was detected in 98% samples. Nucleotide sequence analysis of the carboxy-terminal region of EBNA1 showed that Vvvl was deteted in 98% samples. DNA sequence analysis of LMP1 C-terminus indicated that China 1 was 90% in this study. There were no significant differences in the frequency of Vvv1 and China 1 between the IM and HLH samples (P = 1.00). Linkage analysis of EBV types, EBNA1 and LMP1 variants indicated that 90% of EBV type A was associated with EBNA1-Vvv1 variant and LMP1-China 1 variant in 40 cases. Full length of LMP1 gene was successfully amplified in 35 cases. Four Chinese groups (CG1-4) were identified. The percentage of CG1-CG4 were 85%, 6%, 6% and 3%, respectively.</p><p><b>CONCLUSION</b>EBV type A is predominant in primary EBV infection in children in Beijing Area. EBNA1-Vvv1 and LMP1-China 1 variants were predominant genotypes in this area. There is a high linkage between EBNA1-Vvv1 variant and LMP1-China 1 variant. Four Chinese groups (CG1-4) were identified according to the full length of LMP1 gene and CG1 was the most prevalent.</p>

Simultaneous detection of human parainfluenza viruses 1, 2, 3 by multiplex real-time reverse transcription-polymerase chain reaction with LNA probes / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>Human parainfluenza virus (HPIV) types 1, 2 and 3 are major viral pathogens responsible for upper and lower respiratory tract infections. In this study, a real-time RT-PCR was developed using multiplex primers-probe (HPIV-1, 2, 3) for the simultaneous detection of both HPIV1, HPIV2 and HPIV3 genomes.</p><p><b>METHODS</b>Optimal primers and probes were designed using specialized software. The conditions for multiplex real-time RT-PCR had been optimized. The synthesis of RNA standards of HPIV1, 2, 3 were used a T7 RNA polymerase. Check the specificity sensitivities and stability of one step RT-PCR assay.</p><p><b>RESULTS</b>Obtained in a 10-fold dilution series assay demonstrate a high sensitivity of the assay with a lowest detection limit of 10 copies for HPIV1, 100 copies for HPIV2 and 100 copies for HPIV3.</p><p><b>CONCLUSION</b>The assays demonstrates an improved sensitivity and scope of detecting HPIV1, 2, 3 viruses relative to routine antigen detection assays while the quantitative utility may facilitate investigation of the pre-diagnosis and respiratory virus pathogenesis.</p>

Three years surveillance of viral etiology of acute lower respiratory tract infection in children from 2007 to 2010 / 中华儿科杂志

<p><b>OBJECTIVE</b>Viruses are common pathogens of acute lower respiratory tract infection (ALRTI) in children. There are few studies on consecutive monitoring of viral pathogens of ALRTI in a larger cohort during the past several years. The aim of this study was to investigate the viral pathogens of ALRTI in children of different age groups and to outline the epidemic feature of different viruses.</p><p><b>METHOD</b>(1) Totally 1914 (1281 male and 709 female) children with clinical diagnosis of ALRTI during the period of March 2007 to March 2010 were recruited into this study. These patients were hospitalized patients in department of internal medicine or outpatients in emergency department in Beijing Children's Hospital. The patients were divided into four groups, including 1072 patients < 1 year old, 326 patients 1- < 3 years old, 158 patients 3- < 6 years old, 358 patients ≥ 6 years old. One nasopharyngeal aspirate specimen was collected from each patient. Reverse transcription (RT) PCR methods were applied to detect common respiratory viruses including respiratory syncytial virus (RSV), human rhinovirus (HRV), influenza virus type A, B and C (IFA, IFB, IFC), parainfluenza virus (PIV) type 1-4, adenovirus (ADV), enterovirus (EV), human coronavirus (HCOV), human metapneumovirus (HMPV) and human bocavirus (HBOV).</p><p><b>RESULT</b>(1) The total positive rate of viruses was 70.3%. The positive rate was 83.0% (890/1072) in the group of < 1 year old, and 80.1% (261/326) in group of 1- < 3 years old, 60.8% (96/158) in group of 3- < 6 years old and 27.7% (99/358) in group of ≥ 6 years old, respectively. There was a significant difference in the positive rate among different age groups (χ² = 2213.5, P = 0.000). The top three viruses were RSV, HRV and PIV; and the positive rates were 50.9%, 36.2% and 12.0% respectively in group of < 1 year old. (2) The epidemic seasons of RSV and HRV were winter and spring, and PIV infection was epidemic in spring and summer. (3) The detection rates of 2 or more viruses were 38.2%, 36.4%, 30.2% and 15.2% in groups of < 1 year old, 1- < 3 years old, 3- < 6 years old and ≥ 6 years old, respectively. There was a significant difference in the mixed infection rate among different age groups (χ² = 1346.00, P = 0.000).</p><p><b>CONCLUSION</b>RSV, HRV and PIV were the most predominant pathogens in younger children with ALRTI. Different viral infections had different seasonal features. Mixed infections with two or more viruses were detected in substantial proportion of patients with ALRTI, but further studies are needed to explore the clinical significance of mixed infection with viruses in patients with ALRTI.</p>

Incidence and relationship of human cytomegalovirus infection and human herpesvirus 6 infection in pediatric patients after hemopoietic stem cell transplantation / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study the incidence of human cytomegalovirus (CMV) and human herpesvirus 6 (HHV-6) infection in pediatric patients with hemopoietic stem cell transplantation (HSCT), and to explore the relationship between CMV and HHV-6 infection in pediatric patients with HSCT.</p><p><b>METHODS</b>Pediatric patients with HSCT in hemotology center of Beijing Children's Hospital were enrolled into this study from June 2007 to October 2009. Peripheral blood were collected every week after HSCT, and Fluorescent quantitation PCR and conventional PCR were used to detect CMV DNA load in serum and HHV-6 DNA in peripheral blood respectively. Genetic typing was conducted on HHV-6.</p><p><b>RESULTS</b>Fifty two pediatric patients with HSCT were enrolled into this study, and six hundreds and thirty six specimens were collected totally. CMV DNA was detected in fifty two specimens from twenty cases. The median time was 56 days after HSCT. The incidence of CMV infection was 38.5% (20/52) in all HSCT patients and 47.6% (20/42) in allogene HSCT patients. The incidence of late CMV infection was 22.2% (6/27) in allogene HSCT. Three patients died of CMV infection,and two died of CMV interstitial pneumonia. HHV-6 DNA was detected in thirty three specimens from fourteen cases. The median time was 23 days after HSCT. The incidence of HHV-6 infection was 26.9% (14/52)in all HSCT patients and 31% (13/42) in allogene HSCT patients. The genotype of HHV6 was all type B. HHV-6 DNA was positive in six of twenty cases with CMV infection. The incidence of co-infection was 30% (6/20).</p><p><b>CONCLUSIONS</b>There was a substantial incidence of CMV and HHV6 infection after HSCT. The relationship between earlier HHV6 infection and later CMV infection in pediatric patients with HSCT need further study.</p>

Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis: a retrospective study of 78 pediatric cases in mainland of China / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>The clinical characteristics of Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH) are largely unreported in the pediatric patients in mainland of China. The main aim of this study was to recognize the clinical features of EBV-HLH in children and to explore its prognosis and risk factors.</p><p><b>METHODS</b>A retrospective study was performed on 78 pediatric patients with EBV-HLH who were admitted to Beijing Children's Hospital between 2003 and 2008. All patients' medical records were reviewed and analyzed. For each patient, demographic, clinical, laboratory and outcome information was collected. Statistical analysis was conducted via multivariate and univariate analysis.</p><p><b>RESULTS</b>The age of onset peaked between 1 - 2 years and boys were more likely developed EBV-HLH. EBV-HLH occurred mainly in the serological pattern with EBV nuclear antigen (EBNA) positive (70.5%). The overall fatality of the disease was 56.7%. Twelve of the 39 fatalities (30.8%) died rapidly within 2 months after diagnosis. Multivariate analysis revealed that not receiving chemotherapy (P = 0.002), > or = 4 weeks of illness prior to diagnosis (P = 0.004), and albumin levels < 20 g/L (P = 0.045) significantly predicted an increased fatality risk.</p><p><b>CONCLUSIONS</b>EBV-HLH is a severe disease with a high fatality rate that occurs mainly in the serological pattern with EBNA positive. Early initiation of chemotherapy and timely diagnosis significantly improves survival rate. Practical strategies should focus on reducing the likelihood of early death.</p>

Clinical analysis and follow-up study of Epstein-Barr virus associated-hemophagocytic lymphohistiocytosis in childhood / 中华儿科杂志

<p><b>OBJECTIVE</b>To identify the clinical characteristics of and to explore the prognostic factors influencing mortality in children with Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH).</p><p><b>METHOD</b>A retrospective study was conducted on 62 pediatric patients with EBV-HLH who were admitted to our hospital between 2003 and 2008. All their medical records were reviewed and analyzed. For each patient, demographic, clinical and laboratory data, genetic findings and outcome information were collected. The patients were divided into two groups: deceased or survived based on the follow-up results. Comparative analysis of the data was done by using independent-samples t test and Logistic multiple and univariate regression.</p><p><b>RESULT</b>(1) Among the 62 EBV-HLH patients, 36 were male and 26 were female. The age of onset ranged from 2 months to 14 years and most of the patients were between 1 and 3 years of age. EBV-HLH occurred mainly in the setting of reactivation (61.3%). (2) All patients exhibited persistent or intermittent fever and cytopenia >/= 2 cell lines. Most of the patients presented with hepatomegaly (83.9%), splenomegaly (72.6%) and lymphadenopathy (69.4%). The main laboratory features showed an elevation of serum ferritin and aminotransferase levels. A reduction in serum albumin was observed and exhibited coagulopathy with hypofibrinogenemia and hypertriglyceridemia in most of the patients. Forty-eight of patients had hemophagocytosis in bone marrow at diagnosis of EBV-HLH. The serum EBV DNA level in 14 of 31 patients with EBV-HLH was in the range of 5.12 x 10(2) - 7.69 x 10(7) copies/ml with a mean value of 10(3.9) copies/ml. (3) Three heterozygous mutations in coding region were found, which resulted in amino acid change (C102F, S108N and T450M) in 3 patients. One patient had compound heterozygous mutations (S108N and T450M) in the PRF1 gene as the background defect and documented familial HLH type 2 (FHL2). (4) During the observational period, 35 of 57 patients (61.4%) died 3 months to 3 years after the onset, while 21 of whom died despite aggressive polychemotherapy, 15 of whom died within 2 months after hospitalization. The deceased patients were more likely to have lower albumin level and more prolonged activated partial thromboplastin time than the survived patients (P < 0.05 for all comparisons). Multivariate Logistic regression analysis revealed that duration of illness >/= 1 month, non-chemotherapy, albumin level < / = 25 microg/L and internal organs hemorrhage were related with the prognosis significantly (P < 0.05 for all comparisons).</p><p><b>CONCLUSION</b>This study revealed that EBV-HLH infection in pediatric patients had severe clinical courses and prognosis was poor and the majority of cases underwent EBV reactivation. The early diagnosis, prompt and proper chemotherapy can improve the survival rate. The duration of illness >/= 1 month, non-chemotherapy, decreases in albumin and internal organs hemorrhage were the risk factors related to mortality in children with EBV-HLH.</p>

Clinical analysis and follow-up study of chronic active Epstein-Barr virus infection in 53 pediatric cases / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Chronic active Epstein-Barr virus infection (CAEBV) has been previously reported to be sometimes associated with an aggressive clinical course. The characteristics of CAEBV in Mainland Chinese pediatric patients are largely unreported. The main aims of this survey were to recognize the clinical features of CAEBV in children and to explore its diagnostic criteria and risk factors.</p><p><b>METHODS</b>A retrospective study was performed on 53 pediatric patients (36 boys and 17 girls) with CAEBV who were admitted to Beijing Children's Hospital between 2003 and 2007. All their medical records were reviewed and analyzed. For each patient, demographic, clinical, laboratory data and outcome were collected. Independent-samples t test was used for statistical analysis.</p><p><b>RESULTS</b>The age at onset of CAEBV was from 2 months to 14.6 years (mean (5.3+/-3.3) years). At the time of onset, 43.4% patients had an infectious mononucleosis-like symptom. Most patients exhibited intermittent fever (92.5%, 49/53), hepatomegaly (81.1%, 43/53) and splenomegaly (77.4%, 41/53). Life-threatening complications including hemophagocytic syndrome (24.5%, 13/53), interstitial pneumonia (24.5%, 13/53), hepatic failure (15.1%, 8/53) and malignant lymphoma (11.3%, 6/53) were also observed. The serum EBV DNA level in 23 patients with CAEBV was in the range of 5.05 x 10(2)-4.60 x 10(6) copies/ml with a mean value of 10(3.7) copies/ml. Many patients with CAEBV generally had continuous symptoms during the observational period. Eleven out of 42 patients (26.2%) died 7 months to 3 years after onset. Deceased patients were more likely to have had lower platelet counts and albumin levels than the living patients (P<0.05 for all comparisons).</p><p><b>CONCLUSIONS</b>The study reveals that CAEBV in Chinese pediatric patients has a severe clinical course and prognosis is poor. Thrombocytopenia and decreases in albumin might potentially be risk factors for a poor prognosis. EBV loads should be measured and tissue should be stained on hybridization probes for EBV-encoded small RNA (EBER) if a patient presents with the known symptoms of CAEBV.</p>

Genetic characterization of the N protein of subgroups A and B human respiratory syncytial viruses / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To clarify the genetic characteristics of N protein coding region of HRSV isolates from Beijing and GenBank downloaded sequences.</p><p><b>METHODS</b>Reverse transciption polymerase chain reaction (RT-PCR) was performed to amplify the N protein gene of 2 A and 2 B subgroups HRSV isolates from Beijing in the year 2004. The RT-PCR products were sequenced for N protein coding region. The sequences of N protein coding region of 4 Beijing isolates and those downloaded from GenBank were compared and analyzed.</p><p><b>RESULTS</b>The differences in number of nucleotide and deduced amino acid between 2 A Beijing 2004 isolates and prototype strain Long were 36-40 (3.1%-3.4%) and 4 (1.0%). The differences in number of nucleotide and deduced amino acid between 2 B Beijing 2004 isolates and prototype strain CH18537 were 17 (1.4%) and 1 (0.3%). The differences in number of nucleotide and deduced amino acid were 3-172 (0.25%-14.63%) and 0-18 (0-4.6%) respectively between 4 Beijing 2004 isolates and GenBank sequences.</p><p><b>CONCLUSION</b>N gene is the highly conservative gene in the HRSV genome. The variation between A and B subgroups were widely distributed in the entire gene of N protein, while the variation within the A or B subgroups HRSV was considerably lower.</p>

Determination of plasma EBV DNA in 91 children with EBV-associated diseases / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To determine the plasma level of Epstein Barr virus (EBV) DNA in children with EBV associated diseases, and to investigate the dynamic changes of EBV DNA level after initial infection as well as the relationship between EBV DNA level and the diseases severity.</p><p><b>METHODS</b>The subjects consisted of 73 children with primary EBV infection (infectious mononucleosis, pneumonia,etc.) and 18 children with severe EBV-associated diseases (chronic active EBV infection, hemophagocytic lymphohistiocytosis, etc.). The plasma EBV DNA level was detected by a real-time PCR assay.</p><p><b>RESULTS</b>The plasma EBV DNA level decreased with the infection time in children with primary EBV infection. Two weeks after infection, plasma EBV DNA was almost undetectable. The positive rate of plasma EBV DNA in children with severe EBV associated diseases increased significantly when compared with that in children with primary EBV infection (89% vs 16%; p<0.05).</p><p><b>CONCLUSIONS</b>The level of EBV replication may be reduced with the infection time. Dynamic determination of blood EBV DNA is useful for the evaluation of disease severity in children with EBV infection.</p>

Mutations in the perforin gene in children with hemophagocytic lymphohistiocytosis / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Recent studies have reported germline mutations in the perforin gene (PRF1) in some types of hemophagocytic lymphohistiocytosis (HLH). However, the prevalence of PRF1 mutations in HLH in Chinese pediatric patients has not been extensively studied. The aim of this study was to investigate the prevalence of mutations and sequence variations in the PRF1 gene in Chinese pediatric patients with HLH.</p><p><b>METHODS</b>Polymerase chain reaction (PCR) was performed with five pairs of primers for the coding exons and the flanking intron sequences of PRF1. Sequencing of PCR products was subsequently applied in 30 pediatric patients with HLH and in 50 controls.</p><p><b>RESULTS</b>Three heterozygous mutations in a coding region were found, which resulted in amino acid changes (C102F, S108N and T450M) in three patients. These mutations were not detected in control subjects. One patient had compound heterozygous mutations (S108N and T450M) in PRF1 as the background defect, and documented familial HLH type 2 (FHL2). One synonymous sequence variant (Q540Q) was observed in one patient but not in the controls. Two SNPs (A274A, H300H) in the coding region were detected in HLH patients and controls, but without differences in the heterozygosity rate between the two groups (P > 0.05 for all comparisons).</p><p><b>CONCLUSIONS</b>We have identified three patients with three heterozygous missense mutations in PRF1; two of those three mutations (C102F and S108N) have so far been found only from Chinese patients. These findings are useful in evaluating the prevalence of PRF1 mutations in Chinese pediatric patients with HLH, and to correlate their genotype with phenotype. Some patients without familial history probably have primary HLH, which should be suspected even beyond the usual age range.</p>

Detection of C-terminal 30 bp-deletion mutation of latent membrane protein 1 of EBV in childhood lymphoma / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study C-terminal 30 bp-deletion mutation of latent membrane protein 1 of the virus from childhood lymphoma.</p><p><b>METHODS</b>Nested-PCR was used to amplify C-terminal of EBV-LMP1 from childhood lymphoma and non-lymphoma associated to EBV, including Hodgkin lymphoma (HL), non-Hodgkin lymphoma (NHL)and reactive hyperplasia of lymph node (RL). Sequence analysis was performed on the positive PCR product.</p><p><b>RESULTS</b>LMP1 with 30 bp deletion in C-terminal was detected in 11/25 HL,3/8 NHL and 5/15 RL cases respectively. There were no significant differences among HL, NHL and RL (P = 0.793). Sequence analysis showed that LMP1 detected in this study belongs to the following three subgroups: B95.8, China1 and China2.</p><p><b>CONCLUSION</b>LMP1 with 30 bp deletion in C-terminal widely existed in childhood HL,NHL and RL. There was no correlation between special types of LMP1 and the diseases. There were 3 LMP1 subgroups of EBV in children's lymphoma in the cases studied, including B95.8, China 1 and China 2.</p>

Correlation of Epstein-Barr virus infection and childhood lymphoma / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the correlation of Epstein-Barr virus (EBV) infection and childhood lymphoma.</p><p><b>METHODS</b>Paraffin-embedded specimens of lymphoma collected between 1996 and 2005, including 36 Hodgkin lymphomas (HL) and 51 non-Hodgkin lymphomas (NHL), were included in this study. Paraffin-embedded specimens of reactive hyperplasia of lymph nodes (RL) collected during the same period were used as controls. Immunohistochemical (IHC) assay was used to detect EBV-LMP1 and in situ hybridization (ISH) to detect EBV-EBERs.</p><p><b>RESULTS</b>EBV was detected in 72.2% (26/36) of the Hodgkin lymphomas, 15.7% (8/51) of the non-Hodgkin lymphomas and 33.3% (15/45) of the reactive hyperplasia of lymph nodes. There was a significant difference among Hodgkin lymphoma, non-Hodgkin lymphoma and RL (P = 0. 000).</p><p><b>CONCLUSION</b>Childhood Hodgkin lymphoma is closely correlated with Epstein-Barr virus infection. However, the low rate of EBV infection detected in childhood non-Hodgkin lymphoma might be due to heterogeneous distribution of pathological types in this study.</p>

Human bocavirus in children suffering from acute lower respiratory tract infection in Beijing Children's Hospital / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Human bocavirus (HBoV) is a parvovirus recently found to possibly cause respiratory tract disease in children and adults. This study investigated HBoV infection and its clinical characteristics in children younger than five years of age suffering from acute lower respiratory tract infection in Beijing Children's Hospital.</p><p><b>METHODS</b>Nasopharyngeal aspirates were collected from children suffering from acute lower respiratory tract infection during the winters of 2004 to 2006 (from November through the following February). HBoV was detected by polymerase chain reaction amplification and virus isolation and the amplification products were sequenced for identification.</p><p><b>RESULTS</b>HBoV infection was detected in 16 of 333 study subjects. Coinfections with respiratory syncytial virus were detected in 3 of 16 HBoV positive patients with acute lower respiratory tract infection. The median age for HBoV positive children was 8 months (mean age, 17 months; range, 3 to 57 months). Among the HBoV positive children, 14 were younger than 3 years old, 9 were younger than 1 year old and 7 were younger than 6 months. These 16 positive HBoV children exhibited coughing and abnormal chest radiography findings and more than 60% of these children had wheezing and fever. Ten children were clinically diagnosed with pneumonia, 2 bronchiolitis, 2 acute bronchitis and 2 asthma. One child died.</p><p><b>CONCLUSIONS</b>HBoV was detected in about 5% of children with acute lower respiratory infection seen in Beijing Children's Hospital. Further investigations regarding clinical and epidemiologic characteristics of HBoV infection are needed.</p>