REAL-TIME MONITORING OF CYTOMEGALOVIRUS INFECTIONS AFTER STEM CELL TRANSPLANTATION USING THE TaqMan POLYMERASE CHAIN REACTION ASSAYS1

Yun, Zhibing; Lewensohn-Fuchs, Ilona; Ljungman, Per; Vahlne, Anders

doi:10.1097/00007890-200004270-00037

Cited by 59 publications

(57 citation statements)

References 9 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…[8][9][10] Pre-emptive therapy relies on the availability of a sensitive, rapid and specific diagnostic test of CMV-I. Among the available screening techniques to detect CMV replication (infection), the most commonly used are the pp65 antigenemia (pp65Ag) assay, and several qualitative and quantitative polymerase chain reaction (PCR) analyses for CMV DNA or CMV mRNA 9,[11][12][13][14][15] At our institution, since 1993 all AlloHSCT recipients at risk for developing CMV-I were monitored with the detection of pp65Ag in peripheral blood polymorphonuclear granulocytes. 16 The progressive switch to quantitative PCR (quantPCR) for the diagnosis and follow-up of numerous viral pathogens led to the implementation of a validated quantPCR for monitoring CMV in immunocompromised hosts, and, thus, in late 2003 we switched to a PCR-based pre-emptive therapy.…”

Section: Introductionmentioning

confidence: 99%

Cytomegalovirus infection and disease after reduced intensity conditioning allogeneic stem cell transplantation: single-centre experience

Piñana

Martino

Barba

et al. 2009

Bone Marrow Transplant

View full text Add to dashboard Cite

The aim of this study was to analyse the incidence and risk factors for cytomegalovirus infection (CMV-I) and disease (CMV-D) after a reduced intensity conditioning allogeneic hematopoietic stem cell transplantation (alloHSCT-RIC). We included 186 consecutive alloHSCT-RIC adult patients at risk for CMV reactivation (patient and/or donor CMV seropositivity). Conditioning regimen was based on fludarabine plus an alkylating agent. For guiding pre-emptive anti-CMV therapy, Pp65 Antigenemia (pp65Ag) (n ¼ 116) or quantitative polymerase chain reaction (quantPCR) (n ¼ 70) were used. The 2-year incidence of CMV-I and/or CMV-D was 36% (11% for CMV-D). Of note, 12/14 (86%) episodes of CMV-D in the pp65Ag group had lung involvement compared with only 3/ 15 (20%) in the quantPCR group (P ¼ 0.01). Importantly, the number of patients who developed CMV pneumonia with prior negative screening tests was unusually high (67% overall). Multivariate analysis of risk factors for CMV-D identified two risk factors: (i) steroid therapy for moderateto-severe graft-vs-host disease (GVHD) (hazard ratio 4.7, P ¼ 0.02); and (ii) alternative donors (non-HLA-identical siblings) [hazard ratio 2.7, P ¼ 0.002]. Our findings suggest that CMV is still a major concern in alloHSCT-RIC. Variables associated with poor anti-CMV T-cell recovery (that is, GVHD and donor type) are helpful in identifying patients at higher risk for CMV-D in the alloHSCT-RIC setting.

show abstract

Section: Introductionmentioning

confidence: 99%

Cytomegalovirus infection and disease after reduced intensity conditioning allogeneic stem cell transplantation: single-centre experience

Piñana

Martino

Barba

et al. 2009

Bone Marrow Transplant

View full text Add to dashboard Cite

show abstract

“…Thus, the concentration is probably correct. Furthermore, our qPCR was compared with the same PCR assays in a single PCR format for which the standard consisted of a plasmid containing the target sequences for gB and pol (23,24). Irrespective of whether DNA preparations from a complete genome or from plasmids were used and independent of the mono or duplex format, the detection limit was about five copies per reaction.…”

Section: Discussionmentioning

confidence: 99%

“…X17403) were pol, at nucleotides (nt) 78197 to 78262, and gB, at nt 81855 to 81916. Primers and probes were designed as previously described by Yun et al (23,24), except that the pol probe was labeled with a 6CR6G fluorophore (absorption maximum, 524 nm) (Scandinavian Gene Synthesis, Köping, Sweden) instead of FAM and dark quenchers replaced TAMRA at the 3Ј ends of the two probes. To avoid degradation of the fluorogenic labeled probes, we routinely divided them into aliquots at a concentration of 45 M in 30-l volumes before storage at Ϫ20°C.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Comparison of a Duplex Quantitative Real-Time PCR Assay and the COBAS Amplicor CMV Monitor Test for Detection of Cytomegalovirus

Herrmann¹,

Larsson²,

Rubin³

et al. 2004

J Clin Microbiol

Self Cite

View full text Add to dashboard Cite

A duplex quantitative real-time PCR (qPCR) assay was designed to detect both the polymerase gene (pol) and the glycoprotein gene (gB) of cytomegalovirus (CMV). The detection limit of the qPCR was determined to be 1 to 3 copies/reaction and the linear measure interval was 10 3 to 10 8 copies/ml. The qPCR system was compared to the COBAS Amplicor CMV Monitor test (COBAS) by an analysis of 138 plasma samples. Both systems detected CMV in 71 cases and had negative results for 33 samples. In addition, 34 samples were positive by qPCR and negative by the COBAS assay, but in no case was the COBAS result positive and the qPCR result negative. Thus, qPCR detected 48% more positive cases than the COBAS method. For samples with >10 5 copies/ml by qPCR, a saturation effect was seen in the COBAS assay and quantification required dilution. Copy numbers for pol and gB by qPCR generally agreed. However, the reproducibility of qPCR assays and the need for an international standard are discussed. Discrepant copy numbers for pol and gB by qPCR were found for samples from two patients, and sequence analysis revealed that the corresponding CMV strains were mismatched at four nucleotide positions compared with the gB fragment primer sequences. In conclusion, a duplex qPCR assay in a real-time format facilitates quantitative measurements and minimizes the risk of false-negative results.Human cytomegalovirus (CMV) is a common opportunistic pathogen found in immunocompromised patients, e.g., patients in a posttransplantation state or those infected with human immunodeficiency virus. Early detection and close monitoring of CMV infections are important for preventing the reactivation of endogenous CMV and for the surveillance of recipients of seropositive donor transplants. Appropriate antiviral therapy is based on a knowledge of the viral load as well as a quantification of the initial amount of CMV and the rate of increase between the last negative and the first positive PCR sample. These are independent risk factors for CMV disease

show abstract

“…The real-time PCR assay for CMV is based on the protocol of Yun et al [4] The target sequence is the viral glycoprotein B. The linear range of the assay extends from 8 9 10E2 to 8 9 10E7 IU/ml, and the 95 % confidence interval of the precision within this range is 3/4 0.5 log10.…”

Section: Preemptive CMV Protocolmentioning

confidence: 99%

A stringent preemptive protocol reduces cytomegalovirus disease in the first 6 months after kidney transplantation

et al. 2012

View full text Add to dashboard Cite

Background The optimal strategy to prevent cytomegalovirus (CMV) disease after kidney transplantation continues to be open to debate. The preemptive approach requires regular determination of CMV viremia and prompt initiation of therapy. Methods We retrospectively compared the incidence of CMV disease during two periods at our center: A first phase (P1, n = 84 kidney recipients), during which time the intensity of surveillance was determined by the responsible physician, was compared to a second phase (P2, n = 74), when a stringent protocol of CMV surveillance was required for all patients. The preemptive approach was applied for all CMV risk groups; prophylaxis was optional in the case of treatment for rejection or delayed graft function in the intermediate-and high-risk group. Follow-up was truncated at 6 months after transplant surgery. CMV syndrome was differentiated from asymptomatic replication by the presence of at least one systemic symptom, while diagnosis of CMV end-organ disease required histological confirmation.Results Immunosuppression was similar in the two periods. CMV prophylaxis was used equally (26 %) in both periods. The probability for asymptomatic viremia episodes was not different for patients in P1 and P2 regardless of the prevention strategy. For patients following the preemptive strategy, the probability for CMV disease was increased during P1 (p = 0.016), despite fewer PCR assays being performed in phase 2. Protocol violations were only observed during P1. Conclusions The probability of CMV disease episodes (CMV syndrome and CMV end-organ disease) was substantially reduced using a very stringent protocol. This study highlights the crucial importance of a stringent protocol with optimal adherence by all caregivers if the preemptive strategy is to be successful.

show abstract

REAL-TIME MONITORING OF CYTOMEGALOVIRUS INFECTIONS AFTER STEM CELL TRANSPLANTATION USING THE TaqMan POLYMERASE CHAIN REACTION ASSAYS1

Abstract: The TaqMan assays may potentially serve as a useful tool for rapid quantification of CMV infections in stem cell transplant patients.

Cited by 59 publications

References 9 publications

Cytomegalovirus infection and disease after reduced intensity conditioning allogeneic stem cell transplantation: single-centre experience

Cytomegalovirus infection and disease after reduced intensity conditioning allogeneic stem cell transplantation: single-centre experience

Comparison of a Duplex Quantitative Real-Time PCR Assay and the COBAS Amplicor CMV Monitor Test for Detection of Cytomegalovirus

A stringent preemptive protocol reduces cytomegalovirus disease in the first 6 months after kidney transplantation

Contact Info

Product

Resources

About