新生儿巨细胞病毒感染的检测

建立了用ELISA检测巨细胞病毒(HCMV)IgA抗体的方法,並用于检测北京地区100对母婴的HCMV抗体,母血、脐带血、母乳中HCMV-IgG抗体的阳性率分别为83％,75％和38％,HCMV-IgA抗体的阳性率分别为19％,15％和58％,对其中的16名婴儿半年后追踪观察,5名出生时母、脐血全为阴性的,有2名抗体阳转。8名出生时母、脐血均阳性的,有1名IgA仍阳性並检查发现肝大肋下二指。另1名IgG持续阳性,其他6名婴儿抗体转阴。3名出生时母血HCMV-IgG阳性者中,1名婴儿IgA和‘gG转阳,此时母亲IgA也阳转。随访的16名婴儿中有3名可能是生后半年内受HCMV感染。     

肿瘤患者化疗后人巨细胞病毒活动性感染检测

探讨肿瘤患者化疗后人巨细胞病毒感染检测方法的应用价值。使用免疫组化法、酶联免疫吸附试验检测IgG/M抗体,以及实时荧光定量(FQ-PCR)检测HCMV DNA。47份全血标本中抗原阳性率为48.9%,平均抗原阳性细胞数7.9±8.1(1-65)/5×104WBC,HCMV DNA阳性率19.1%(10/47),HCMV DNA含量均值为6.320×105copies,白细胞HCMV-DNA阳性率51%(25/47),HCMV DNA含量均值为3.830×107 copies,HCMV pp65抗原阳性率为48.9%(23/47),IgG抗体均阳性,IgM抗体阳性率为23.4%(12/47),以PP65抗原阳性为对照,IgM抗体检测的敏感率仅为49.3%。在连续动态检测HCMV多种指标时,结合DNA及抗原动态检测具有更高临床应用价值。     

HCMV-DNA定量检测和HCMV-IgG抗体AI检测在儿童HCMV感染诊断中的临床价值

摘要 目的：探讨人巨细胞病毒（HCMV）-DNA定量检测和HCMV-免疫球蛋白G（IgG）抗体亲和力指数（AI）检测在儿童HCMV感染诊断中的临床价值。方法：收集高度疑似HCMV活动性感染患儿血清样本103例作为研究组,健康体检儿童血清样本94例作为对照组。分析HCMV-DNA定量检测结果和HCMV-IgG抗体AI检测结果,并比较不同年龄、不同性别患儿HCMV-DNA阳性结果检出率和低HCMV-IgG抗体AI检出情况。结果：研究组血清HCMV-DNA阳性率为33.01%（34/103）,对照组血清HCMV-DNA均为阴性,研究组血清HCMV-DNA阳性率明显高于对照组,差异有统计学意义（P<0.05）。研究组血清低HCMV-IgG抗体AI检出率为13.59%（14/103）,对照组未检出低HCMV-IgG抗体AI,研究组血清低HCMV-IgG抗体AI检出率高于对照组,差异有统计学意义（P<0.05）。研究组不同性别之间患儿血清的HCMV-DNA阳性率、低HCMV-IgG抗体AI检测结果均无统计学差异（P>0.05）。研究组年龄1～5岁患儿血清HCMV-DNA阳性率明显低于年龄1 d～<6个月和年龄6个月～<1岁患儿（P<0.05）。三个年龄段患儿的血清低HCMV-IgG抗体AI检测结果均无统计学差异（P>0.05）。结论：1岁以下儿童更易受到HCMV感染,HCMV-DNA定量检测和HCMV-IgG抗体AI检测结果可以为临床早期诊断和治疗HCMV感染提供有效依据。     

献血员巨细胞病毒感染的研究

用PCR法和DNA杂交法检测同一献血员的白细胞及血清中的HCMV-DNA,并用ELISA法检测血清中的HCMV-IgM、IgG(测四个滴度),连续两年共检测白细胞和血清样本各200人份.PCR法检测白细胞中的HCMV-DNA阳性率分别为63%和70%,DNA杂交法检测的阳性率为42%和50%.PCR法检测血清中的HCMV-DNA的阳性率为49%和53%,DNA杂交法检测的阳性率为33%和39%.HCMV-IgM阳性率两年均为5%.HCMV-IgG阳性率分别为54%和58%.     

新生儿高胆红素血症与先天性巨细胞病毒感染的相关性研究

目的:探讨新生儿高胆红素血症与巨细胞病毒(HCMV)感染的相关性及临床意义。方法:对170例疑为高胆红素血症新生儿血清标本分别检测HCMV的IgG和IgM及血清总胆红素(TBIL)。排除高胆红素血症新生儿血清标本100例为正常对照。结果:高胆红素血症患者血清HCMV-IgG、IgM阳性率高于正常对照组,两者比较有统计学意义。结论:巨细胞病毒感染与新生儿高胆红素血症相关性大,是引起高胆红素血症的主要病原之一。     

孕妇尿液人巨细胞病毒DNA筛查及血液HCMV抗体测定的对照研究

目的对于孕妇尿液进行HCMV—DNA筛查,减少和有效地避免胎儿及新生儿的HCMV感染。HCMV．DNA筛查同时进行HCMV抗体检测,明确诊断HCMV感染的灵敏、准确方法。方法应用荧光定量PCR（FQ．PCR）方法进行尿液HCMV-DNA测定。血液HCMV IgM、IgG抗体检测应用酶联免疫（ELISA）方法。结果筛查6568例孕妇尿HCMV．DNA,阳性273例,阳性率为4．2％。孕妇在12—20周者,阳性率为10．3％;孕妇在21～30周者,阳性率为33．3％;孕妇在31～39周者,阳性率为56．4％。在273例尿液HCMV—DNA阳性者中,56例同时进行血液HCMV IgM、IgG抗体检测,Igi抗体阳性者2例,IgG抗体阳性者22例。结论在孕妇HCMV感染的筛查与诊断中FQ．PCR是灵敏准确的方法。孕妇中HCMV—DNA筛查是保证母婴健康,提高人口素质的保障。     

胶体金免疫层析法检测新型冠状病毒IgM/IgG抗体的临床评价与应用

新型冠状病毒(SARS-CoV-2)感染暴发流行已成为全球公共卫生事件,急需快速有效的现场诊断试剂。为探讨SARS-CoV-2病毒IgM/IgG抗体胶体金免疫层析法检测效果及临床应用价值,本研究选取304例新冠肺炎临床诊断病例、114例SARS-CoV-2核酸检测阴性的正常人(138)及其他发热伴呼吸道症状病例(64),采用胶体金法对采自上述病例的血浆或血清标本进行IgM和IgG抗体检测,并选取部分病例进行了全血和血浆或血清标本的检测同源性比较,进一步对304例临床诊断病例的病毒核酸、IgM/IgG抗体的时间分布进行了分析。结果显示,304临床诊断病例中,SARS-CoV-2核酸检测阳性病例为105例,胶体金法检测SARS-CoV-2 IgM和IgG抗体的敏感性分别为76.2%(80/105)和86.6%(91/105),IgM/IgG抗体阳性总体符合率为96.1%(101/105);核酸、抗体均为阴性者为73例;其余126例临床诊断病例中,IgM阳性率为69.2%(87/126),IgG阳性率为98.3%(125/126),IgM/IgG总体符合率为100%(126/126)。健康人和其他发热病人中,IgM和IgG检测特异性分别为99%(200/202)和98%(198/202)。同源全血与血浆或血清标本抗体检测结果的总符合率为99%,显示两者具有高度一致性。本研究提示,胶体金法检测SARS-CoV-2抗体有较好的敏感性及特异性,可用于临床辅助诊断和流行病学调查等,具有较广泛的应用场景,在新冠肺炎疫情防控中具有一定的应用价值。     

人巨细胞病毒的生物素DNA探针的制备和应用

本研究用克隆的HCMV AD169株DNA片段,制备了生物素标记的DNA探针,建立了检测临床脐带血、尿标本中HCMV DNA的核酸探针杂交方法。该探针可测出100pg同源DNA,不与人胚肺细胞、Hep-2细胞DNA以及其他疱疹病毒的DNA发生反应。用核酸杂交方法检测了30份脐带血标本,有11例阳性,阳性率为33％。10例孕妇尿标本中,3例阳性,阳性率为30％。检测结果表明：我们建立的生物素标记的HCMV DNA探针的点杂交法,具有高度的特异性、敏感性,比分离病毒法更迅速,可用于HCMV感染的临床标本的病毒核酸检测。     

人巨细胞病毒pp65特异性抗体测定在原发感染诊断中的应用

目的 通过测定患者单份血清人巨细胞病毒(HCMV)pp65特异性IgM抗体和IgG亲和指数(AI),建立HCMV原发感染的临床判断标准.方法 从临床收集40份患儿血清和尿标本,以本室自制的pp65为抗原,运用间接酶联免疫吸附试验(ELISA)检测血清标本中HCMV pp65特异性IgM抗体;同时通过尿素变性实验,以6M尿素作为温和蛋白变性剂,测定HCMVpp65 IgG AI.尿标本常规处理后接种人胚成纤维(HF)细胞进行病毒分离,观察HCMV特异性细胞病变效应(CPE),聚合酶链反应(PCR)试验检测细胞培养物UL83基因,间接免疫荧光试验检测细胞玻片HCMV抗原.并将病毒分离结果 同血清学方法 进行比较.结果 40例标本中,IgM阳性13例,IgG阳性30例,其中,仅IgM阳性4例,病毒分离结果 亦为阳性,可诊断为原发感染;30例IgG阳性标本中,2种抗体均为阳性9例(A组),其中,5例AI＜50﹪,病毒分离结果 均为阳性,判断为原发感染;1例AI在50﹪与60﹪之间,为可疑原发感染,需要进一步鉴定;3例AI＞60﹪,判断为继发感染.IgG阳性21例(B组),其中仅3例(14.29﹪)AI＜50﹪,但病毒分离结果 为阴性,提示患者不久前曾发生HCMV原发感染,特异性IgM抗体已经转阴,病毒进入潜伏状态;余18例患者AI均＞60﹪,判断为继发感染;2种抗体均为阴性的标本有6例,病毒分离结果 亦为阴性,说明患者未被感染.统计学分析,A组与B组之间差异有统计学意义(P＜0.05).血清学方法 与病毒分离比较,一致率为75.49﹪,该方法 灵敏度为100﹪,特异度为87.10﹪,准确度为90.00﹪.结论 以HCMV pp65重组蛋白为抗原检测特异性抗体以及相应IgG AI的ELISA,可快速诊断HCMV原发感染和继发感染;该方法 具有高度特异性与敏感性,操作简便,重复性好,具有良好的临床应用前景.     

肾移植术后巨细胞病毒pp65检测的临床意义

目的探讨肾移植受者术后,巨细胞病毒被膜磷蛋白pp65的检测在活动性巨细胞病毒感染中的意义。方法用间接免疫荧光法检测肾移植受者术后HCMV-pp65,同时用酶联免疫捕获法检测HCMV-IgM抗体,共采集91份血标本。结果 91份血标本中,HCMV-pp65阳性24份（26.4%）,HCMV-IgM抗体阳性4份（4.4%）,在24例HC-MV-pp65抗原血症阳性的患者中,有20例出现HCMV感染症状及HCMV病。结论 HCMV-pp65在活动性巨细胞病毒感染的检测中具有早期、准确的优点,可辅助临床对HCMV感染进行早期诊断与治疗。     

糖尿病患者人巨细胞病毒感染的抗体检测分析

目的 :了解人巨细胞病毒 (HCMV)感染与糖尿病的关系。方法 :运用捕获 EL ISA、间接 EL ISA对 516例糖尿病患者及 14 8例健康正常人 HCMV-Ig M、HCMV-Ig G进行检测 ,并对比分析 HCMV感染状态 ,根据糖尿病患者的初发年龄和病程的不同进行了分组探讨。结果 :糖尿病患者 HCMV-Ig M、HCMV-Ig G抗体阳性率明显高于对照组 (P<0 .0 1) ,始发病年龄越小及病程越短的糖尿病病人 HCMV-Ig M抗体阳性率明显升高 ;年龄较大及病程较长的糖尿病人 HCMV-Ig G检出率较高。结论 :年龄小或病程短的糖尿病人以原发性 HCMV感染为主 ,HCMV感染在糖尿病的发病机制中起一定的作用     

1253例孕妇和新生儿TORCH检测结果分析

目的 了解TORCH感染情况,为妇幼保健提供参考依据.方法 采用捕获ELISA方法对1 253例孕妇和新生儿的血清进行TORCH-IgM抗体检测.结果 689例孕妇CMV、TOX、RV及HSVⅡ特异性IgM抗体阳性率分别为1.89％、0.87％、0.44％和0.73％;564例新生儿CMV-IgM、TOX-IgM、RV-IgM和HSVⅡ-IgM阳性率分别为6.91％、0.18％、0和0.35％.结论 检测TORCH特异性抗体对优生优育有重要关系.     

Preparation of P52 recombinant antigenic protein from human cytomegalovirus (HCMV)

Analysis of published reports helped us single out the most potent antigens among HCMV proteins: phosphoproteins pp150(UL32) and p52(UL44). Theoretical computer analysis of p52 epitopes showed the main antigenic determinants not cross-reacting with antigens of other viruses. Virus-containing (strain AD169) material was obtained and genome DNA was isolated. Amplification of a site of gene UL44 coding for unique determinants detected a PCR fragment of required electrophoretic mobility. The fragment was cloned in vector pLBE. The specificity of cloning was confirmed by restriction analysis of theoretical sites. Nucleotide sequence of cloned fragment of UL44 gene was studied by Maxam-Gilbert's method. Cloning in expressing bacterial vectors helped obtain HCMV recombinant protein p52 in the pure form and fused with beta-galactosidase. Enzyme immunoassay with HCMV-positive and negative donor sera and ABBOTT HCMV sera showed that recombinant p52 increased the sensitivity and specificity of a previously obtained recombinant pp150 as an antigen to HCMV-IgG and HCMV-IgM. The sensitivity and specificity is 100% with 98-99% reliability.     

兰州地区孕妇和新生儿血清中风疹病毒特异性IgM抗体测定研究

本文采用ＥＬＩＳＡ抗ｕ抗体捕捉法检测了兰州地区７１２例孕妇和６２４例新生比血清中风疹病毒特异性ＩｇＭ抗体（ＲＶ－ＩｇＭ）。实验结果为：７１２例孕妇中,ＲＶ－ＩｇＭ阳性者有８例,阳性率为１．１２％：６２４例新生儿脐带血清标本中,ＲＶ－ＩｇＭ阳性者６例,阳性率为０．９６％。结果表明,兰州地区孕妇中有一定的风疹病毒原发感染病例,新生儿也存在一定的风疹病毒先天性感染问题。     

Preterm and term neonates transplacentally acquire IgG antibodies specific to LPS from Klebsiella pneumoniae, Escherichia coli and Pseudomonas aeruginosa

High incidences of Gram-negative bacteria are found in neonatal nosocomial infections. Our aim was to investigate placental transmission of immunoglobulin G (IgG) reactive with lipopolysaccharide from Klebsiella pneumoniae, Pseudomonas aeruginosa and Escherichia coli O111, O6 and O26. The total and lipopolysaccharide-specific IgM and IgG were determined in 11 maternal/umbilical-cord sera aged ≤33 weeks (GI); 21 aged >33 and <37 weeks (GII); and 32 term newborns (GIII). The total and lipopolysaccharide-specific IgM concentrations were equivalent in maternal sera. The total IgG concentrations were equivalent in maternal and newborn sera, with the exception of GIII newborns as compared with their mothers (P<0.0001) and with neonates from GI and GII (P<0.05). Lipopolysaccharide-specific IgG concentrations were lower in GI neonates than in their mothers (P<0.01) and lower in GII (P<0.05). Lower lipopolysaccharide-specific IgG levels were observed among neonates only for O111 in GI (P<0.05) and for O26 and Pseudomonas in GII, both as compared with GIII (P<0.05). The anti-lipopolysaccharide IgG transfer ratios were lower in GI (except for O26) and in GII (except for Klebsiella and O111) as compared with GIII (P<0.05). Our results suggest that the greater susceptibility to infections in preterm infants is influenced (besides the humoral response) by factors intrinsic and extrinsic to the condition of prematurity.     

Human Rotavirus-Specific IgM Memory B Cells Have Differential Cloning Efficiencies and Switch Capacities and Play a Role in Antiviral Immunity In Vivo

Protective immunity to rotavirus (RV) is primarily mediated by antibodies produced by RV-specific memory B cells (RV-mBc). Of note, most of these cells express IgM, but the function of this subset is poorly understood. Here, using limiting dilution assays of highly sort-purified human IgM(+) mBc, we found that 62% and 21% of total (non-antigen-specific) IgM(+) and RV-IgM(+) mBc, respectively, switched in vitro to IgG production after polyclonal stimulation. Moreover, in these assays, the median cloning efficiencies of total IgM(+) (17%) and RV-IgM(+) (7%) mBc were lower than those of the corresponding switched (IgG(+) IgA(+)) total (34%) and RV-mBc (17%), leading to an underestimate of their actual frequency. In order to evaluate the in vivo role of IgM(+) RV-mBc in antiviral immunity, NOD/Shi-scid interleukin-2 receptor-deficient (IL-2Rγ(null)) immunodeficient mice were adoptively transferred highly purified human IgM(+) mBc and infected with virulent murine rotavirus. These mice developed high titers of serum human RV-IgM and IgG and had significantly lower levels than control mice of both antigenemia and viremia. Finally, we determined that human RV-IgM(+) mBc are phenotypically diverse and significantly enriched in the IgM(hi) IgD(low) subset. Thus, RV-IgM(+) mBc are heterogeneous, occur more frequently than estimated by traditional limiting dilution analysis, have the capacity to switch Ig class in vitro as well as in vivo, and can mediate systemic antiviral immunity.     

Significance of circulating toxin and antitoxin in unimmunized tetanus cases of neonates and infants

The level of circulating tetanus toxin, antitoxin and their individual influence on the outcome of tetanus cases were determined in unimmunized 125 neonatal and 39 infant cases of tetanus. PHA (passive haemagglutination) test showed 40% positive cases for toxin while its absence in the remaining cases indicated of either toxin fixation to the central nervous system (CNS) or it got neutralized by antitoxin. TN (toxin neutralization) and PHA test carried out in 46 sera samples revealed a strong positive correlation (r = 0.9) showing that 35/46 (76%) and 38/46 (82.6%) samples were positive for antitoxin, respectively. 25.4% of the neonate and infant cases and 34% of the control group had a protective serum tetanus antitoxin level. 42.5% of the paired sera from unimmunized mothers and their neonates showing nonprotective antitoxin levels suggested that a high level of antitoxin is needed for transplacental transfer, although transfer may not play a decisive role in the resistance against the disease. The presence of toxin or antitoxin in the clinical cases did not affect the outcome of the disease, although in neonates, presence of toxin was found to be a bad prognostic sign. This study explicitly advocates for the need to improve the vaccination coverage strategy.