北京地区成人呼吸道感染病人中人偏肺病毒感染情况的初步分析

目的:了解北京地区成人呼吸道感染患者中人偏肺病毒(hMPV)的感染情况、流行分布和临床表现特点。方法:采集2010年5月至2011年4月北京地区成人呼吸道感染患者鼻咽拭子标本413份,利用巢式PCR法进行hMPV筛查,对PCR阳性片段进行核酸序列测定,确定hMPV感染基因型别;同时,分析hMPV感染的流行病学特点,并对感染阳性患者进行临床表现的初步分析。结果:413份鼻咽拭子标本中检出hMPV阳性4份(0.97%),分别存在于2010年8月、10月和2011年2月、4月,其中1例合并鼻病毒感染;感染患者临床表现主要是发热、鼻塞、流涕、头痛、咳嗽,有1例出现呕吐和腹泻;hMPV阳性片段系统进化分析发现这4例感染中的3例为B2型,1例为A2b型。结论:北京地区成人呼吸道感染患者中存在hMPV感染,其基因型为B2和A2b型。     

Evidence of Recombination and Genetic Diversity in Human Rhinoviruses in Children with Acute Respiratory Infection

Background

Human rhinoviruses (HRVs) are a highly prevalent cause of acute respiratory infection in children. They are classified into at least three species, HRV-A, HRV-B and HRV-C, which are characterized by sequencing the 5′ untranslated region (UTR) or the VP4/VP2 region of the genome. Given the increased interest for novel HRV strain identification and their worldwide distribution, we have carried out clinical and molecular diagnosis of HRV strains in a 2-year study of children with acute respiratory infection visiting one district hospital in Shanghai.

Methodology/Findings

We cloned and sequenced a 924-nt fragment that covered part of the 5′UTR and the VP4/VP2 capsid genes. Sixty-four HRV-infected outpatients were diagnosed amongst 827 children with acute low respiratory tract infection. Two samples were co-infected with HRV-A and HRV-B or HRV-C. By comparative analysis of the VP4/VP2 sequences of the 66 HRVs, we showed a high diversity of strains in HRV-A and HRV-B species, and a prevalence of 51.5% of strains that belonged to the recently identified HRV-C species. When analyzing a fragment of the 5′ UTR, we characterized at least two subspecies of HRV-C: HRV-Cc, which clustered differently from HRV-A and HRV-B, and HRV-Ca, which resulted from previous recombination in this region with sequences related to HRV-A. The full-length sequence of one strain of each HRV-Ca and HRV-Cc subspecies was obtained for comparative analysis. We confirmed the close relationship of their structural proteins but showed apparent additional recombination events in the 2A gene and 3′UTR of the HRV-Ca strain. Double or triple infections with HRV-C and respiratory syncytial virus and/or bocavirus were diagnosed in 33.3% of the HRV-infected patients, but no correlation with severity of clinical outcome was observed.

Conclusion

Our study showed a high diversity of HRV strains that cause bronchitis and pneumonia in children. A predominance of HRV-C over HRV-A and HRV-B was observed, and two subspecies of HRV-C were identified, the diversity of which seemed to be related to recombination with former HRV-A strains. None of the HRV-C strains appeared to have a higher clinical impact than HRV-A or HRV-B on respiratory compromise.     

Glycomic Analysis of Human Respiratory Tract Tissues and Correlation with Influenza Virus Infection

The first step in influenza infection of the human respiratory tract is binding of the virus to sialic (Sia) acid terminated receptors. The binding of different strains of virus for the receptor is determined by the α linkage of the sialic acid to galactose and the adjacent glycan structure. In this study the N- and O-glycan composition of the human lung, bronchus and nasopharynx was characterized by mass spectrometry. Analysis showed that there was a wide spectrum of both Sia α2-3 and α2-6 glycans in the lung and bronchus. This glycan structural data was then utilized in combination with binding data from 4 of the published glycan arrays to assess whether these current glycan arrays were able to predict replication of human, avian and swine viruses in human ex vivo respiratory tract tissues. The most comprehensive array from the Consortium for Functional Glycomics contained the greatest diversity of sialylated glycans, but was not predictive of productive replication in the bronchus and lung. Our findings indicate that more comprehensive but focused arrays need to be developed to investigate influenza virus binding in an assessment of newly emerging influenza viruses.     

小儿呼吸道感染常见病原菌及药敏结果分析

目的:了解小儿下呼吸道感染的常见病原菌及其对抗菌药物的敏感性.方法:对北京市和平里医院儿科两年来确诊治疗的156例小儿呼吸道感染患儿的痰标本进行培养及药敏试验.结果:156例标本检出致病菌98株,其中G-杆菌71株(72.4％),G+球菌23株(23.5％),真菌4株(4.1％),主要致病菌依次为肺炎克雷伯菌26株,大肠埃希菌20株.流感嗜血杆菌13株,金黄色葡萄球菌7株,肺炎链球菌5株.药敏结果显示,G-杆菌对亚胺培南均敏感,其次为头孢西丁、庆大霉素,G+球菌对万古霉素均敏感,对头孢西丁、庆大霉素、环丙沙星、阿米卡星有较高的敏感性.结论:革兰阴性杆菌为本地区小儿呼吸道感染的主要致病菌,真菌的感染率也呈上升趋势,临床上应及时检测,合理用药.     

儿童下呼吸道感染的金黄色葡萄球菌耐药性分析

对 4 2 38例下呼吸道感染患儿深部痰液标本中的金黄色葡萄球菌进行分离培养 ,并用细菌自动分析系统Vitek 6 0进行菌种鉴定和药物敏感试验。金黄色葡萄球菌分离阳性率为 2 .1% (89/ 4 2 38)。所分离的89株金黄色葡萄球菌中 ,95 .5 % (85 / 89)菌株产 β 内酰胺酶 ;但仅 4 .5 % (4 / 89)菌株耐苯唑西林 (MRSA) ,其中 3株显示多重耐药。 96 .6 %和 4 8.3%的菌株分别对青霉素和红霉素耐药 ,14 .6 %～ 2 5 .9%分别对四环素、林可霉素、复方新诺明耐药 ;对其余临床常用抗生素 ,如庆大霉素、苯唑西林、氨苄西林 /舒巴坦、阿莫西林 /克拉维酸、头孢唑啉、环丙沙星、左氧氟沙星等的耐药率均低于 10 %。上述实验结果提示 ,本地区引起患儿下呼吸道感染的金黄色葡萄球菌对多数临床常用抗生素仍敏感 ,β 内酰胺酶阳性菌株极为普遍 ,但MRSA所占比例不高。     

Surveillance and Genome Analysis of Human Bocavirus in Patients with Respiratory Infection in Guangzhou, China

Human bocavirus (HBoV) is a novel parvovirus associated with respiratory tract diseases and gastrointestinal illness in adult and pediatric patients throughout the world. To investigate the epidemiological and genetic variation of HBoV in Guangzhou, South China, we screened 3460 throat swab samples from 1686 children and 1774 adults with acute respiratory infection symptoms for HBoV between March 2010 and February 2011, and analyzed the complete genome sequence of 2 HBoV strains. Specimens were screened for HBoV by real-time PCR and other 6 common respiratory viruses by RT-PCR or PCR. HBoV was detected in 58 (1.68%) out of 3460 samples, mostly from pediatric patients (52/58) and inpatient children (47/58). Six adult patients were detected as HBoV positive and 5 were emergency cases. Of these HBoV positive cases, 19 (32.76%) had co-pathogens including influenza virus (n = 5), RSV (n = 5), parainfluenza (n = 4), adenovirus (n = 1), coronavirus (n = 7). The complete genome sequences of 2 HBoVs strains (Genbank no. {"type":"entrez-nucleotide","attrs":{"text":"JN794565","term_id":"361064618","term_text":"JN794565"}}JN794565 and {"type":"entrez-nucleotide","attrs":{"text":"JN794566","term_id":"361064623","term_text":"JN794566"}}JN794566) were analyzed. Phylogenetic analysis showed that the 2 HBoV strains were HBoV1, and were most genetically close to ST2 (GenBank accession number DQ0000496). Recombination analysis confirmed that HBoV strain GZ9081 was an intra–genotype recombinant strain among HBoV1 variants.     

重症急性呼吸道感染患儿中人鼻病毒分型检测及流行病学分析

目的:调查重症急性呼吸道感染患儿中人鼻病毒(HRV)的感染情况,初步了解不同型别HRV流行病学和临床特点。方法:收集2008年5月至2010年3月北京儿童医院重症急性呼吸道感染住院患儿鼻咽抽吸物样本259份,采用巢式PCR法,先用鼻病毒5′UTR引物筛查样本中该病毒总的感染情况,再用针对VP4-VP2区域的引物对阳性样本进行分型检测;同时对阳性样本进行其他常见呼吸道病毒共感染检测与流行病学及临床特点分析。结果:259例样本中,5′UTR初筛HRV阳性89例(34.4%),与其他常见呼吸道病毒存在共感染54例(60.7%);可通过VP4-VP2分型测序确定39例HRV-A(48.1%)、4例HRV-B(4.9%)、38例HRV-C(46.9%);HRV-A感染率最高的季节分布是秋季;HRV-A和HRV-C型感染者在临床特点上无明显差别。结论:HRV是儿童重症急性呼吸道感染的重要病原之一,新发现的HRV-C基因型感染率与HRV-A相似,但HRV在儿童重症急性呼吸道感染中的病原学意义还须进一步确认。     

住院肺炎患儿偏肺病毒和呼吸道合胞病毒感染研究

为探讨沈阳地区肺炎患儿中偏肺病毒(hMPV)和呼吸道合胞病毒(RSV)感染情况,用反转录聚合酶链反应(RT-PCR)法对部分因肺部感染住院患儿进行了病原学研究。结果显示,hMPV感染率为9%;而RSV感染率为46%,其中A型占40%,B型占60%。研究结果表明,在婴幼儿肺炎患儿中RSV感染率高于hMPV感染率,两者所致肺炎在发病年龄、性别及临床症状上无显著区别。     

Molecular Typing and Epidemiology Profiles of Human Adenovirus Infection among Paediatric Patients with Severe Acute Respiratory Infection in China

Background

Human adenoviruses (HAdVs) have been recognised as pathogens that cause a broad spectrum of diseases. The studies on HAdV infection among children with severe acute respiratory infection (SARI) are limited.

Objective

To investigate the prevalence, epidemiology, and genotype of HAdV among children with SARI in China.

Study Design

Nasopharyngeal aspirates (NPAs) or induced sputum (IS) was collected from hospitalised children with SARIs in Beijing (representing Northern China; n = 259) and Zhejiang Province (representing Eastern China; n = 293) from 2007 to 2010. The prevalence of HAdV was screened by polymerase chain reaction (PCR), followed by sequence typing of PCR fragments that targeted the second half of the hexon gene. In addition, co-infection with other human respiratory viruses, related epidemiological profiles and clinical presentations were investigated.

Results and Conclusions

In total, 76 (13.8%) of 552 SARI patients were positive for HAdV, and the infection rates of HAdV in Northern and Eastern China were 20.1% (n = 52) and 8.2% (n = 24), respectively. HAdV co-infection with other respiratory viruses was frequent (infection rates: Northern China, 90.4%; Eastern China, 70.8%). The peak seasons for HAdV-B infection was winter and spring. Additionally, members of multiple species (Human mastadenovirus B, C, D and E) were circulating among paediatric patients with SARI, of which HAdV-B (34/52; 65.4%) and HAdV-C (20/24, 83.3%) were the most predominant in Northern and Eastern China, respectively. These findings provide a benchmark for future epidemiology and prevention strategies for HAdV.     

2010-2019年北京地区腹泻患儿中人腺病毒41型Fiber基因分子特征和进化分析

人腺病毒41型(Human adenovirus 41,HAdV-41)是引起儿童腹泻的重要病原之一.为了解北京地区腹泻儿童中HAdV-41 Fiber基因的分子变异和遗传进化特征,选取2010-2019年首都儿科研究所附属儿童医院门诊腹泻患儿HAdV-41阳性的粪便标本,扩增Fiber全基因,使用多种生物信息学软件进行系统发生、遗传进化、种群演变和选择压力等分析.结果显示HAdV-41本地毒株存在1644碱基(547氨基酸)和1689碱基(562氨基酸)两种长度的Fiber基因,截短的15个氨基酸位于Fiber蛋白纤维轴区.Mann-Whitney U秩和检验提示检测到感染短Fiber基因的HAdV-41的患儿年龄更小(P=0.036).同源性分析结果显示Fiber基因的核苷酸和氨基酸序列同源性均大于97％,主要分为四个进化分支(Ⅰ、Ⅱ、Ⅲ和Ⅳ),其中Ⅰ和Ⅲ是主要的流行分支,分支Ⅰ主要在国内流行,而分支Ⅲ全球均有流行.HAdV-41共祖起源可追溯到1856年,平均进化速率每年为6.81×105碱基替换/位点,10年间理论有效种群一直比较平稳呈缓慢下降趋势.压力选择分析表明,一个正向和三个负向选择位点均位于Fiber蛋白的纤维轴区.上述结果表明2010-2019年间不同进化分支的HAdV-41在北京地区流行,进化速率稳定.Fiber蛋白轴区存在两种不同类型的氨基酸缺失.     

Surveillance for Seasonal Influenza Virus Prevalence in Hospitalized Children with Lower Respiratory Tract Infection in Guangzhou,China during the Post-Pandemic Era

Background

Influenza A(H1N1)pdm09, A(H3N2) and B viruses have co-circulated in the human population since the swine-origin human H1N1 pandemic in 2009. While infections of these subtypes generally cause mild illnesses, lower respiratory tract infection (LRTI) occurs in a portion of children and required hospitalization. The aim of our study was to estimate the prevalence of these three subtypes and compare the clinical manifestations in hospitalized children with LRTI in Guangzhou, China during the post-pandemic period.

Methods

Children hospitalized with LRTI from January 2010 to December 2012 were tested for influenza A/B virus infection from their throat swab specimens using real-time PCR and the clinical features of the positive cases were analyzed.

Results

Of 3637 hospitalized children, 216 (5.9%) were identified as influenza A or B positive. Infection of influenza virus peaked around March in Guangzhou each year from 2010 to 2012, and there were distinct epidemics of each subtype. Influenza A(H3N2) infection was more frequently detected than A(H1N1)pdm09 and B, overall. The mean age of children with influenza A virus (H1N1/H3N2) infection was younger than those with influenza B (34.4 months/32.5 months versus 45 months old; p<0.005). Co-infections of influenza A/ B with mycoplasma pneumoniae were found in 44/216 (20.3%) children.

Conclusions

This study contributes the understanding to the prevalence of seasonal influenza viruses in hospitalized children with LRTI in Guangzhou, China during the post pandemic period. High rate of mycoplasma pneumoniae co-infection with influenza viruses might contribute to severe disease in the hospitalized children.     

泛福舒合穴位贴敷对治疗儿童反复呼吸道感染临床疗效分析

目的:探析泛福舒联合穴位贴敷对治疗儿童反复呼吸道感染的临床疗效.方法:选取2009年9月-2012年9月就诊于我院儿科的反复呼吸道感染的患儿60例随机分为两组,对照组给予常规抗生素加穴位(肺俞、膻中、天突、足三里)贴敷(每次12小时,隔日1次,15次为一个疗程)治疗;观察组在此基础上加用泛福苏治疗(3.5 mg,每天清晨空腹顿服,连服10天,停20天为一个疗程),两组均治疗3个疗程后,随诊6个月,观察对比两组患者治疗效果、免疫指标、用药不良反应、远期疗效等.结果:观察组在发热天数、咳嗽天数和抗生素使用时间上都明显少于对照组,两组相比具有统计学意义(P＜0.05);免疫指标治疗前后比较中,对照组各免疫指标治疗前后均没有明显改变,观察组血清IgG在治疗后明显升高,具有统计学差异(P＜.05),IgA在治疗后升高,但没有显著差异,IgM治疗后基本没有改变;观察组远期疗效明显优于对照组,两组相比具有统计学意义(P＜0.05).结论:泛福舒联合穴位贴敷对治疗儿童反复呼吸道感染临床效果显著,治疗有效率高,具有积极的临床意义,值得推广.     

Studies on Parainfluenza Virus Infections among Infants and Children in Sendai

A cell line sensitive enough for the recovery of all parainfluenza viruses and free of simian virus contamination frequently occurring in monkey kidney cells was sought. The VERO cell obtained from African monkey kidney was found suitable for the initial isolation of types 1, 2 and 3 parainfluenza viruses, although the cells did not always allow the successive transfer. Mixed cultures of VERO and HEp-2 cells were also useful in the recovery of various respiratory viruses including parainfluenza viruses. The characteristics of hemagglutinins of parainfluenza viruses were examined, and type 2 parainfluenza and SV5 viruses agglutinated both guinea pig and green monkey erythrocytes at 36 C, whereas types 1 and 3 parainfluenza viruses agglutinated only guinea pig erythrocytes. Thus parainfluenza viruses were divided into two groups by the presence or absence of hemagglutinins for green monkey erythrocytes. Identification of these parainfluenza isolates, employing HI microtechnique was simple and reliable, even with the first passage harvest, when guinea pig erythrocytes were used and the test read at 36 C. Specific standard antisera for these parainfluenza viruses were prepared by immunizing chickens intravenously and bleeding within a short period. These type-specific antisera were useful for the identification of parainfluenza isolates by HI test.     

北京地区人呼吸道合胞病毒核蛋白基因的序列分析及原核表达

为了解北京地区流行的人呼吸道合胞病毒(HRSV)N蛋白基因的特征,并用大肠杆菌表达获得N蛋白,从2006年1月至3月收集的首都儿科研究所附属儿童医院急性呼吸道感染患儿标本中分离到7株HRSV毒株(3株A亚型,4株B亚型),分别经RT-PCR扩增得到HRSV N蛋白全基因,克隆至pUCm-T载体中并进行测序和分析;从重组质粒pUCm-N9968中经PCR扩增获得N基因,亚克隆入原核表达载体pET30a( )中,得到重组表达质粒pET30a-N9968,转化大肠杆菌BL21(DE3),IPTG诱导培养;SDS-PAGE和Western blot检测目的蛋白的表达和抗原性。经扩增并测序,7株HRSV N蛋白基因全长均为1 176 bp,编码一个含391个氨基酸的蛋白;北京地区7株RSV分离株的N蛋白基因之间的核苷酸和推导的氨基酸同源性分别在85.4%~99.7%之间和95.4%~100%之间,进一步证明N蛋白基因的高度保守性。经诱导培养产生大量带6个组氨酸标记的重组N蛋白,主要以包涵体形式存在。N蛋白粗提物经Ni2 亲和层析获得较理想纯化。Western blot结果显示,所表达的蛋白能与特异性单克隆抗体和人血清反应。说明本研究构建的重组pET30a-N9968原核表达质粒使N蛋白获得了高效表达,并且具有特异的抗原活性,为今后进一步研究奠定了基础。     

阿奇霉素口服治疗小儿呼吸道肺炎支原体感染的临床研究

目的:探讨阿奇霉素口服治疗小儿呼吸道肺炎支原体感染的临床效果及安全性。方法:选取2012年5月至2013年10月在我院就诊的144例呼吸道肺炎支原体感染患儿,随机分为观察组和对照组,每组各72例。对照组加用注射用乳糖酸红霉素静滴治疗,观察组加用阿奇霉素口服治疗。对比两组患者的治疗效果及不良反应的发生情况。结果:观察组的总有效率为94.45%,明显高于对照组的83.33%;观察组患儿各临床症状及体征的消失时间均明显短于对照组;观察组患儿不良反应的总发生率为6.95%,显著低于对照组的16.67%,差异均有统计学意义(P0.05)。结论:相比于注射用乳糖酸红霉素静滴治疗,阿奇霉素口服治疗小儿呼吸道肺炎支原体感染疗效更佳,且具有不良反应小,用药安全,患儿的配合度高的优势。     

NICU患儿呼吸道感染病原菌的分布及耐药性分析

目的:探讨新生儿重症监护病房(NICU)呼吸道感染常见病原菌的分布及耐药情况,为临床合理选用抗菌药物提供参考.方法:对本院2009年1月至2010年12月NICU患儿送检呼吸道标本所分离病原菌的分布及药敏结果进行统计分析.结果:共检出病原菌367株,分离率前6位的病原菌依次为肺炎克雷伯菌(46.0％)、大肠埃希菌(13.9％)、铜绿假单胞菌(10.9％)、阴沟肠杆菌(8.7％)、金黄色葡萄球菌(5.7％)、真菌(4.9％).耐药性分析显示,肺炎克雷伯菌耐药情况严重,ESBLs产生率54.4％,对头孢噻肟、头孢曲松的耐药率分别达91.7％、90.5％,对头孢吡肟、头孢哌酮/舒巴坦、阿莫西林/克拉维酸、头孢他啶、庆大霉素的耐药率分别为68.6％、66.9％、66.9％、47.3％、44.4％,对环丙沙星和阿米卡星的耐药率较低,分别为7.7％和16.6％.大肠埃希菌对青霉素、头孢菌素类的耐药率较高,对亚胺培南、环丙沙星和阿米卡星敏感或较为敏感,ESBLs产生率51.0％,未检出对亚胺培南耐药的肠杆菌科细菌.铜绿假单胞菌除对阿莫西林/克拉维酸、头孢曲松和头孢噻肟的耐药率较高外,对其余抗生素较为敏感或高度敏感.金黄色葡萄球菌对青霉素、红霉素耐药率较高,分别为95.2％、71.4％,对克林霉素、头孢他啶、头孢西丁、阿莫西林/棒酸的耐药率则较低,分别为23.8％、23.8％、14.3％、4.7％,检出MARSA 3株(14.3％),未发现万古霉素耐药的菌株;检出4株肺炎链球菌全部对红霉素的耐药,对β-内酰胺类药物耐药性不严重.结论:肺炎克雷伯菌为NICU患儿呼吸道感染的主要病原菌,且对常用抗生素耐药情况严重.根据病原菌种类及药敏结果合理应用抗菌药是有效控制危重病患儿感染和减少耐药菌株产生的重要手段.