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1.
将50例肾移植术后发热病人的尿标本接种细胞,用聚合酶链反应(PCR)技术检测第1代细胞培养上清中的人巨细胞病毒(HCMV)DNA。结果19例为HCMVDNA阳性,阳性率为38.00%,用固相放射免疫法(RIA)检测HCMV抗原,阳性的有15例,阳性率为27.77%。将54倒尿标本接种人胎肺二倍体单层细胞,分离到6株HCMV。用ELISA检测65例病人单份血清的HCMV-IgM抗体,阳性的有17例,检测45例病人双份血清IgG抗体,11例的第2份血清的HCMV-IgG抗体滴度比第1份有4倍以上升高。上述结果表明,PCR检测结果与RIA的检测结果基本符合;病人血清抗体的阳性率与PCR检测的阳性率基本符合。 相似文献
2.
采用多聚酶链式反应(PCR)技术,随机抽样检测了3418名孕产妇外周血乙型肝炎病毒DNA(HBV-DNA),结果发现阳性标本89例,阳性率为2.604%(89/3418),随后,随机对其中35名母血IBV-DNA阳性的新生儿脐血进行了检测,发现阳性标本5例,阳性率为14.29%(5/35)。 相似文献
3.
急性白血病患儿脑脊液PCR扩增HCMV-DNA的研究 总被引:1,自引:0,他引:1
应用多聚酶链式反应(PCR)扩增和洋地黄探针杂交的方法,对53例急性白血病(AL)患儿脑脊液(CSF)样本进行了检测。经过PCR扩增,可见明显的人巨细胞病毒DNA(HCMV-DNA)400bp扩增带;经洋地黄探针特异性杂交鉴定,AL患儿CSF标本PCR扩增产物的HCMV-DNA阳性检出率为83.02%。本实验为AL病人CSF的HCMV检测提供了一个简便方法。 相似文献
4.
急性白血病患儿脑脊液PCR扩增HCMV—DNA的研究 总被引:1,自引:0,他引:1
应用多聚酶链式反应(PCR)扩增和洋地黄深针杂交的方法,对53例急性白血病(AL)患儿脑脊液(CSF)样本进行了检测,经过PCR扩增,可见明显的人巨细胞病毒DNA(HCMV-DNA)400bp扩增带,经洋地黄探针特异性杂交鉴定,AL患儿CSF标本PCR扩增产生的HCMV-DNA阳性检出率为83.02%。本实验为AL病人CSF的HCMV检测提供了一个简便方法。 相似文献
5.
婴儿持续性黄疸与巨细胞病毒PCR检测研究 总被引:1,自引:0,他引:1
本文应用PCR技术对持续性黄疸婴儿进行巨细胞病毒检测,结果68例患儿血清中巨细胞病毒(CMV)38例阳性,阳性率为55.9%;尿中CMV45例阳性,阳性率为66.2%。PCR与病毒分离及其它血清学方法相比,既快速又敏感,尿标本用于PCR检测CMV-DNA取材及处理简便,且较血标本检出率为高[1]。 相似文献
6.
3418例孕产妇外周血与活产新生儿脐血乙型肝炎病毒感染的P… 总被引:1,自引:1,他引:0
采用多聚酶链式反应(PCR)技术,随机抽样检测了3418名孕产妇外周血乙型肝炎病毒DNA(HBV-DNA),结果发现阳性标本89例,阳性率为2.604%(89/3418),随后,随机对其中35名母血HBV-DNA阳性的新生儿脐血进行了检测,发现阳性标本5例,阳性率为14.29%(5/35)。 相似文献
7.
巨细胞病毒宫内感染的诊断及对胎儿的影响 总被引:4,自引:0,他引:4
采用地高辛探针斑点杂交技术和聚合酶链反应(PCR)技术,对51例人巨细胞病毒(HCMV)血清学抗体IgM阳性孕妇的羊水、脐血和产后两周内的新生儿尿液,进行HCMVDNA检测。同时与酶联免疫吸附法(ELISA)检测脐血、羊水中HCMV-IgM的结果相比较。结果,地高辛探针斑点杂交检测HCMV-DNA的阳性率分别为:羊水21.57%,脐血33.33%,新生儿尿液27.45%。PCR检测HCMV-DNA的阳性率分别为:羊水29.41%,脐血43.14%,新生儿尿液3S,29%。羊水HCMV-IgM阳性率为11.76%,脐血HCMV-IgM阳性率为23.53%。脐血HCMV-DNA阳性的新生儿平均出生体重明显低于脐血HCMV-DNA阴性的新生儿,而脐血HDW-DNA阳性的新生儿肝功能异常、血小板减少以及低Apgar评分的发生率明显高于脐血HCMV-DNA阴性的新生儿。结果表明:采用ER技术检测羊水、脐血或产后两周内新生儿尿液HCMVDNA有助于HCMV官内感染的早期诊断,便于临床早期干预,HCMV官内感染严重影响胎儿的生长发育,可造成胎儿肝功能异常、血小板减少和新生儿窒息的发生。 相似文献
8.
建立了检测HHV-6病毒特异DNA的PCR方法,对43份健康献血员外周血淋巴细胞DNA进行了检测,HHV-6病毒DNA的阳性率为18.6%。同时从HHV-6中国分离株CH3中克隆到其主要衣壳蛋白(MCP)基因片段,并进行了序列分析和同源性比较。 相似文献
9.
应用PCR技术对孕妇HCMV感染的前瞻性研究 总被引:6,自引:1,他引:5
应用聚合酶链反应(PCR)技术对156 例孕妇尿中HCMV 进行检测,并追踪观察114 例新生儿脐血中HCMV 垂直传播情况。结果表明:11 例孕妇尿HCMVDNA 检测阳性,孕期感染率为7-05% ,有异常妊娠史孕妇感染率(14-71 % )较正常孕妇(4-92 %) 明显升高。9 例HCMV 阳性孕妇有4 例观察新生儿脐血HCMVDNA 阳性,占44% ,而HCMVDNA阴性孕妇无1 例新生儿脐血出现阳性结果。我们认为PCR方法是检测HCMV宫内感染的可靠而灵敏的指标,对HCMVDNA阳性的孕妇应进一步检查羊水,如羊水HCMVDNA也阳性,应终止妊娠。 相似文献
10.
套式PCR和原位杂交技术检测肝病患者单个核细胞?… 总被引:1,自引:0,他引:1
目的 了解慢性乙型肝炎(中度)和原发性肝癌(HBsAg阳性)患者PBMC内TTV DNA存在情况。方法 采用套式PCR以及原位杂交技术检测外周血单个核细胞(PBMC)内TTV DNA。结果 套式PCRSWIM26例慢性乙型肝炎(中度)患者PBMC内TTV DNA阳性7例。阳性率26.9%,非常显著高于健康对照(X^12=14.3,P〈0.001);21例原发性肝癌(HBsAg阳性)虱PBMC内TT 相似文献
11.
Sonali K. Sanghavi Kareem Abu-Elmagd Maria Cristina Keightley Kirsten St. George Kathleen Lewandowski Stephanie S. Boes Arlene Bullotta Ryan Dare Maryann Lassak Shahid Husain Eun J. Kwak David L. Paterson Charles R. Rinaldo 《Journal of clinical virology》2008,42(4):335-342
BACKGROUND: Cytomegalovirus (CMV) infection in immunocompromised patients can lead to viremia associated with morbidity and mortality. Monitoring of viral loads in blood is critical for initiating and monitoring antiviral treatment. OBJECTIVES: Validate quantitative real-time PCR assay targeting the US17 and UL54 regions of the CMV genome for automated DNA and extraction and amplification. STUDY DESIGN: 3422 blood specimens from organ transplant recipients, including longitudinal specimens from 12 organ transplant recipients, were tested by CMV PCR and pp65 antigenemia. RESULTS: CMV PCR for both US17 and UL54, was more sensitive and detected CMV DNA earlier and for longer than the CMV pp65 antigenemia test. Using antigenemia results as a reference standard, an optimal cutoff of 500 normalized copies was calculated for both US17 and UL54 PCR targets based on high sensitivity, specificity, and positive and negative predictive values. CMV DNA levels tracked well with clinical symptoms, response to treatment, and antigenemia. CONCLUSIONS: Detection of persistent increases in CMV DNA levels above 500 normalized copies by this real-time PCR assay is indicative of symptomatic CMV disease in organ transplant recipients. Quantitative real-time PCR for CMV DNA can be used in lieu of antigenemia for monitoring CMV infection and determining when to initiate preemptive treatment. 相似文献
12.
Gimeno C Solano C Latorre JC Hernández-Boluda JC Clari MA Remigia MJ Furió S Calabuig M Tormo N Navarro D 《Journal of clinical microbiology》2008,46(10):3311-3318
The performance of a plasma real-time PCR (cytomegalovirus [CMV] PCR kit; Abbott Diagnostics) was compared with that of the antigenemia assay for the surveillance of active CMV infection in 42 allogeneic hematopoietic stem cell transplantation (Allo-SCT) recipients. A total of 1,156 samples were analyzed by the two assays. Concordance between the two assays was 82.2%. Plasma DNA levels correlated with the number of pp65-positive cells, particularly prior to the initiation of preemptive therapy. Fifty-seven episodes of active CMV infection were detected in 37 patients: 18 were defined solely by the PCR assay and four were defined on the basis of the antigenemia assay. Either a cutoff of 288 CMV DNA copies/ml or a 2.42-log10 increase of DNAemia levels between two consecutive PCR positive samples was an optimal value to discriminate between patients requiring preemptive therapy and those not requiring therapy on the basis of the antigenemia results. The real-time PCR assay allowed an earlier diagnosis of active CMV infection and was a more reliable marker of successful clearance of CMV from the blood. Analysis of the kinetics of DNAemia levels at a median of 7 days posttreatment allowed the prediction of the response to CMV therapy. Two patients developed CMV colitis. The PCR assay tested positive both before the onset of symptoms and during the disease period. The plasma real-time PCR from Abbott is more suitable than the antigenemia assay for monitoring active CMV infection in Allo-SCT recipients and may be used for guiding preemptive therapy in this clinical setting. 相似文献
13.
Comparison of PCR and pp65 antigenemia assay with quantitative shell vial culture for detection of cytomegalovirus in blood leukocytes from solid-organ transplant recipients. 总被引:6,自引:8,他引:6
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G A Storch R S Buller T C Bailey N A Ettinger T Langlois M Gaudreault-Keener P L Welby 《Journal of clinical microbiology》1994,32(4):997-1003
This study compared PCR and an assay for cytomegalovirus (CMV) pp65 antigenemia (CMV-vue; INCSTAR Corp.) with a quantitative shell vial culture (QSVC) technique for the detection of CMV in serial blood specimens from 46 solid-organ transplant recipients. In a comparison based on 535 specimens tested by PCR and QSVC, CMV was detected by PCR in 41 and by QSVC in 37 of 43 recipients at risk of CMV infection. The mean number of days after transplantation of initial detection of CMV was 29.9 for PCR and 34.0 for QSVC (P = 0.01). The antigenemia assay was performed on 395 specimens, including 304 of those also tested by PCR. In these specimens, CMV was detected by the antigenemia assay, QSVC, and PCR in 30, 32, and 35 (respectively) of 38 patients at risk, with no statistically significant difference in the time to detection. Each of the assays detected CMV in similar proportions of patients with and without clinically significant CMV infection. PCR stayed positive longer after transplantation than the other assays but frequently returned to negative when more than 6 months had elapsed after transplantation. The antigenemia assay and PCR stayed positive longer after institution of antiviral therapy than QSVC. PCR can provide highly sensitive detection of CMV viremia, but a PCR assay for CMV is not yet available in kit form. The pp65 antigenemia assay and shell vial culture are quantifiable and comparable in sensitivity. Either is recommended for rapid detection of CMV in blood specimens from solid-organ transplant recipients. 相似文献
14.
Human cytomegalovirus immediate-early mRNA detection by nucleic acid sequence-based amplification as a new parameter for preemptive therapy in bone marrow transplant recipients 总被引:2,自引:0,他引:2
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Gerna G Baldanti F Lilleri D Parea M Alessandrino E Pagani A Locatelli F Middeldorp J Revello MG 《Journal of clinical microbiology》2000,38(5):1845-1853
Human cytomegalovirus (HCMV) infection was monitored retrospectively by qualitative determination of immediate-early (IE) mRNA by nucleic acid sequence-based amplification (NASBA) in a series of 51 bone marrow transplant (BMT) recipients. The qualitative results for IE mRNA obtained by NASBA were compared with those obtained by prospective quantitation of HCMV viremia and antigenemia and retrospective quantitation of DNA in blood (DNAemia) by PCR as well as by qualitative determination of late pp67 mRNA by NASBA. On the whole, of the 39 HCMV-positive patients (all asymptomatic), HCMV was detected in 14 (35.9%) by quantitation of viremia, 15 (38.5%) by detection of pp67 mRNA by NASBA, 32 (82.1%) by quantitation of DNAemia, and 33 (84.6%) by quantitation of antigenemia, while HCMV was detected in 38 (97.4%) patients by detection of IE mRNA by NASBA. In the immunocompetent host, IE mRNA was not detected by NASBA in 100 blood donors or during reactivated infections in 30 breast-feeding mothers. Likewise, NASBA did not detect IE mRNA in 56 solid-organ transplant recipients in the first 21 days after transplantation. By using NASBA for detection of IE mRNA as the reference standard for detection of HCMV infection in blood samples, the diagnostic sensitivities were 67.7% for quantitation of DNAemia, 59.0% for quantitation of antigenemia, 18.3% for detection of pp67 mRNA by NASBA, and 16.0% for quantitation of viremia. Specificities and negative and positive predictive values were >90.0, >70.0, and >80.0%, respectively, for all four assays. The mean times to first HCMV detection after bone marrow transplantation were 37.7 +/- 15.4 days for detection of IE mRNA by NASBA, 39.6 +/- 15.6 days for quantitation of antigenemia, 40.9 +/- 15.2 days for quantitation of DNAemia, and 43.7 +/- 16.3 or 43.7 +/- 17.5 days for quantitation of viremia and detection of pp67 mRNA by NASBA, respectively. On the whole, 31 BMT recipients received preemptive therapy by using confirmed antigenemia positivity as a cutoff, while 35 patients could have been treated by using NASBA positivity as a cutoff and 31 could have been treated by using quantitation of DNAemia as a cutoff. In single patients, IE mRNA was detected in every episode of active HCMV infection, mostly preceding and sometimes accompanying antigenemia and DNAemia, whereas pp67 mRNA was detected only concomitantly with the highest peaks of infection. HCMV IE mRNA detection may represent a useful parameter for initiation of preemptive therapy in BMT recipients. 相似文献
15.
Mengoli C Cusinato R Biasolo MA Cesaro S Parolin C Palù G 《Journal of medical virology》2004,74(1):78-84
After bone marrow (BM) or solid-organ (SO) transplantation viremic Cytomegalovirus (CMV) infection is observed frequently. Quantitative assay of CMV in blood helps the management of this clinical condition. In the present report, 83 samples from 39 solid organ recipients, three CMV assays were compared simultaneously for the first time: the Nuclisens CMV pp67 assay (nucleic acid sequence-based amplification, NASBA), an "in-house" quantitative real-time PCR assay (TaqMan) for CMV DNA, and pp65 antigenemia. The relation between CMV DNA and pp65 antigenemia, the quantitative assays, was evaluated on a larger group including 251 blood samples from 118 solid organ recipients. Real-time PCR provided the best results; > or =130 CMV DNA copies/2 x 10(5) peripheral blood leukocytes (PBLs) predicted > or =1 pp65 antigen positive (Ag+) cell/2 x 10(5) PBLs. By taking pp65 antigenemia as the "gold standard," the sensitivity of CMV DNA quantitation and of the pp67 RNA assay were 0.95 and 0.20, respectively, while the corresponding specificity values were 0.50 and 0.93. When real-time PCR was considered as the "gold standard," the sensitivity and specificity of the pp65 antigenemia were 0.65 and 0.91, respectively. Among the three tests examined, the sensitivity of the pp67 RNA assay was the lowest. On the other hand, the pp67 RNA assay was highly specific and effective in pinpointing high viremia patients. The present report, by providing predictive values for all three diagnostic profiles, DNA load, antigenemia, and pp67RNA, is a contribution for validation of real-time PCR as a new standard for quantitative assessment of CMV viremia in clinical settings. 相似文献
16.
Jordi Reina Victoria Fernandez-Baca Juan Saurina Isabel Blanco Maria Munar 《Journal of medical virology》1998,55(3):240-242
The aim of the study was to compare the efficacy of pp65 antigenemia assay and the shell-vial culture (SVC; viremia) for the diagnosis of cytomegalovirus (CMV) infection in renal transplant recipients, comparing the results obtained in the first blood sample and the total number of blood samples analyzed in this group of patients. During the study period, 70 renal transplant recipients were studied: 44 (62.8%) with CMV infection. The method of sedimentation in a dextran solution for leukocyte extraction was used in the pp65 antigenemia assay. The MRC-5 shell-vial assay was used for CMV isolation from leukocytes (viremia). Eighty blood samples were examined from 70 renal transplant recipients: Of the 44 positive samples studied, in 77.5% of cases, both the antigenemia assay and the SVC were positive. In 16.2%, only the antigenemia assay was positive, and, in 6.2%, only the SVC was positive. In all blood samples studied, the antigenemia was present in 93.7% of cases, and the SVC was present in 83.7% (P = 0.04). If the results obtained in only the first blood sample taken for the diagnosis are studied, then we observe that the antigenemia assay was positive in 39 patients (88.6%), whereas the SVC was positive in 41 patients (93.1%), although the difference was not statistically significant (P = 0.39). It is concluded that the inoculation of all of the leukocytes extracted from blood samples in the SVC seems to produce a slight increase in the sensitivity of the cell culture and that the SVC becomes positive before the antigenemia for the detection of CMV in peripheral blood, especially in the first blood sample. J. Med. Virol. 55:240–242, 1998. © 1998 Wiley-Liss, Inc. 相似文献
17.
Monitoring of leukocyte cytomegalovirus DNA in bone marrow transplant recipients by nested PCR.
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K Y Yuen S K Lo E K Chiu S S Wong Y L Lau R Liang T K Chan M H Ng 《Journal of clinical microbiology》1995,33(10):2530-2534
A nested PCR assay for the detection of human cytomegalovirus (CMV) DNA was evaluated by weekly monitoring of blood samples taken from 101 bone marrow transplant (BMT) recipients. When peripheral blood leukocytes were used as the source of CMV DNA, even a modified assay with stringent temperature-cycling conditions was as sensitive as the standard assay. The sensitivity, specificity, and positive predictive value of two consecutively positive leukocytic PCR results with this modified assay in predicting CMV disease of 101 patients submitting 1,441 peripheral blood leukocyte samples were found to be 92.1, 63.5, and 60.3%, respectively. The positive predictive value of patients' seropositivity for CMV was 40%, while that of viremia was 72%. However, viremia followed CMV disease by a median of 1.5 days, while the first leukocytic positive PCR assay preceded disease by a median of 14 days. By use of the criteria of two consecutively positive PCR results instead of recipient CMV seropositivity for starting preemptive ganciclovir treatment, 38 of the 43 recipients with isolated single positive or negative assays (groups I and II) would be spared unnecessary ganciclovir treatment. Moreover, two other findings support the use of antiviral prophylaxis before engraftment in high-risk cases and subsequent preemptive treatment of patients with two consecutively positive PCR assays. First, for 7.9% of 76 patients with positive assays (groups II and III), the first positive PCR assay occurred before engraftment, which implied the presence of viral DNA in the blood (DNAemia) soon after transplantation.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
18.
Clinical relevance of direct quantification of pp65 antigenemia using flow cytometry in solid organ and stem cell transplant recipients 总被引:9,自引:0,他引:9
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Poirier-Toulemonde AS Milpied N Cantarovich D Morcet JF Billaudel S Imbert-Marcille BM 《Journal of clinical microbiology》2000,38(9):3143-3149
A total of 1,305 blood samples from 85 solid organ transplant (SOT) recipients and 25 stem cell transplant (SCT) recipients at risk for cytomegalovirus (CMV) infection were prospectively collected and tested using the shell vial assay (SVA) and a leukocytic qualitative PCR (q-PCR). Of these, 462 specimens were further tested by direct quantification of CMV antigenemia by flow cytometry (FC-Ag), 125 were tested with a quantitative competitive PCR, and 200 were tested for pp65 antigenemia using the slide method (S-Ag). Laboratory data were statistically analyzed according to the presence of CMV-related symptoms. In SOT and SCT recipients, active CMV infection occurred in 63.5 and 36%, respectively, and CMV disease occurred in 53 and 24%, respectively. FC-Ag results correlated better with q-PCR and S-Ag than with SVA. The first test found to be positive during follow-up was FC-Ag in 73% of cases. In SOT recipients, FC-Ag showed the highest sensitivity and negative predictive value for the diagnosis of any grade of CMV disease. For FC-Ag, the threshold beyond which CMV disease was highly probable seemed to lie at 0.20% positive polymorphonuclear leukocytes. FC-Ag appears to be a useful test for the early detection of CMV infection and the prediction of CMV disease. 相似文献
19.
Comparison of cytomegalovirus antigenemia and culture assays in patients on and off antiviral therapy. 总被引:3,自引:0,他引:3
We examined 1,869 consecutive blood specimens from 529 patients (>80% organ transplant recipients) for detection of CMV by antigenemia and culture assays, and compared results between patients on and off antiviral therapy. All 1,869 specimens were tested by the shell vial assay and antigenemia, and 503 were also tested by standard tube culture. The overall positivity rate for each test was 17.0% for antigenemia, 1.8% for shell vial culture assay, and 0.7% by tube culture. No specimens were positive by either shell vial or tube culture, while negative by antigenemia. These findings were consistent across all organ transplant and other patient types. Shell vial positivity was associated with higher antigenemia levels in patients either on or off anti-CMV drug therapy. Among the shell vial positive specimens, the antigenemia counts were higher in patients on antiviral drug therapy as compared to those not on therapy. We conclude that the pp65 antigenemia assay is superior to culture methods for detection of CMV in blood, particularly for patients on anti-CMV drug treatment. Additionally, its quantitative nature renders the antigenemia assay an excellent tracking tool for both resolution of asymptomatic, low level CMV reactivations and response of CMV infection to antiviral treatment. 相似文献
20.
Lilleri D Baldanti F Gatti M Rovida F Dossena L De Grazia S Torsellini M Gerna G 《Journal of medical virology》2004,73(3):412-418
Transplantation Centers using human cytomegalovirus (HCMV) antigenemia-based preemptive therapy will need to replace in the near future the antigenemia assay with a more standardized and automatable assay, such as a molecular assay quantifying HCMV DNA in blood (DNAemia). Thus, in view of replacing antigenemia with clinically safe cutoff values, DNAemia levels corresponding to antigenemia cutoffs guiding HCMV preemptive therapy were determined retrospectively in solid organ and hematopoietic stem cell transplant recipients (HSCTR) using an "in-house" quantitative PCR (QPCR) method. Since preemptive therapy had prevented appearance of HCMV disease in all patients tested, DNA cutoffs determined retrospectively had to be considered as safe clinically as antigenemia cutoffs used prospectively. However, in solid organ transplant recipients (SOTR), initiating preemptive therapy upon an antigenemia cutoff of 100 pp65-positive leukocytes, a DNAemia cutoff of 300,000 copies/ml blood had positive and negative predictive values of >90%, indicating that a DNAemia cutoff could achieve, in terms of prevention of HCMV disease, the same clinical results as the antigenemia cutoff. In HSCTR, initiating preemptive therapy upon first antigenemia positivity, a DNAemia cutoff of 10,000 copies/ml blood had a positive predictive value of >90%, indicating that the great majority of patients treated under the antigenemia guidance would have been treated also using this DNA cutoff. On the other hand, the negative predictive value of 28.6% indicated that two out of three HSCTR had been treated under the antigenemia guidance having the same levels of viral DNA as the untreated patients. The data suggest that a quantitative cutoff could be adopted as a guiding criterion for preemptive therapy also in HSCTR. Regression analysis allowed to determine the DNAemia (corresponding to QPCR) cutoff values for two commercial assays tested both in solid organ and HSCTR. Retrospective DNAemia cutoff values will be verified for safety in prospective trials. 相似文献