Episodes Of Cytomegalovirus DNAemia Research Articles

Impact of an ultrasensitive Cytomegalovirus quantitative nucleic acid test on Cytomegalovirus detection and therapy in renal transplant recipients.

Cytomegalovirus (CMV) infection has broad implications for morbidity and mortality in renal transplant recipients (RTR). Routine surveillance for CMV replication with PCR-based quantitative nucleic acid testing (qNAT) assays is standard practice in most transplant centers, but the impact of assay sensitivity on antiviral decision-making and virologic outcomes has not been studied. We investigated the effects of an ultrasensitive CMV qNAT assay on multiple clinical outcomes, including time to detection and duration of CMV DNAemia. We conducted a single-center cohort study contrasting RTRs monitored with a qNAT with a higher lower limit of quantification (LLOQ >300IU/mL) with those monitored with a more sensitive qNAT (LLOQ >35IU/mL). Patients were stratified by donor (D)/recipient (R) CMV serostatus (D+/R-: high risk; any R+: moderate risk). CMV viral load monitoring was performed monthly post transplantation, with the primary outcomes being time to CMV DNAemia and its duration. Total 1382 patients were analyzed from 2014 to 2016 and 2019 to 2021. Moderate-risk RTRs monitored with the more sensitive assay experienced a greater hazard for the development of a first episode of CMV DNAemia (aHR: 1.95, 95% confidence interval [CI]: 1.55-2.46) and an average of 24 (95% CI: 16.40-31.98) additional days of DNAemia. There was no difference in CMV end-organ disease or 1-year all-cause mortality between moderate-risk RTRs. The more sensitive assay was associated with earlier detection and extended durations of CMV DNAemia in moderate-risk RTRs, without altering clinical outcomes. These findings inform optimal use of these assays and antiviral stewardship in RTRs. The use of ultrasensitive CMV qNAT assays in moderate-risk CMV renal transplant recipients is associated with earlier detection and longer durations of CMV DNAemia without impacting CMV end-organ disease or 1-year mortality.

Features of cytomegalovirus DNAemia and virus-specific T-cell responses in allogeneic hematopoietic stem-cell transplant recipients during prophylaxis with letermovir.

There is scarce information on the natural kinetics of cytomegalovirus (CMV) DNAemia and dynamics of CMV-specific T-cell reconstitution in allogeneic hematopoietic transplant recipients (allo-HSCT) undergoing letermovir (LMV) prophylaxis. Twelve adult CMV-seropositive high-risk recipients (median age, 53 years; 9 males/3 females) undergoing LMV prophylaxis and 13 non-LMV allo-HSCT controls (median age, 58 years; 7 males/6 females) were included. CMV DNAemia in plasma was monitored by real-time polymerase chain reaction. Preemptive antiviral therapy (PET) was administered upon detection of ≥1500IU/ml. CMV-specific interferon-gamma (IFN-γ)-producing CD8+ and CD4+ T cells were enumerated by flow cytometry around days +30, +60, and +90 after allo-HSCT. Ex vivo experiments assessing of the potential effect of LMV on CMV-specific T-cell expansion in a single CMV-seropositive donor were also conducted. Five LMV patients (41.6%) developed CMV DNAemia that cleared spontaneously. Four patients (33.3%) developed CMV DNAemia after LMV cessation, of which two required PET. Nine non-LMV patients (69.2%) developed CMV DNAemia (five required PET). The percentage of LMV and non-LMV patients exhibiting detectable CMV-specific T-cell responses was comparable (7/10 vs. 10/13; p=.71). Nevertheless, median CMV-specific CD4+ and CD8+ T-cell counts were lower in LMV patients by days +60 (p=.006 and .02, respectively) and +90 (p=.08 and .02). Ex vivo, CMV-specific CD8+ T cells expanded to the same level either in the presence (19.8%) or in the absence of LMV (20.6%). In our series, episodes of CMV DNAemia in LMV patients cleared spontaneously. A diminished degree of CMV-specific T-cell reconstitution in LMV patients compared to non-LMV patients was observed.

686. Elevations in TNFα and IL-18 are Associated with Increased Risk of Probable Cytomegalovirus Tissue Invasive Disease in Solid Organ Transplant Recipients

BackgroundHuman cytomegalovirus (CMV) continues to cause significant morbidity and mortality in solid organ transplant (SOT) recipients despite prophylaxis. Tissue invasive CMV disease (TI-CMV) can lead to end-organ damage and graft loss. Diagnosing TI-CMV can be challenging as CMV viral load in the blood does not always correlate with episodes of TI-CMV and therefore definitive diagnosis often requires an invasive procedure such as bronchoscopy or colonoscopy. The purpose of this study was to determine if proinflammatory cytokines, including IL-18, are elevated in SOT recipients with probably TI-CMV as a way to identify patients at risk for this severe form of CMV disease.MethodsThe electronic medical record was searched for adult SOT recipients who were tested for CMV via blood qPCR during an 11-month period.Twenty-nine SOT recipients were identified that had episodes of CMV DNAemia without other concomitant infections during this time period. Patients were divided into those that had probable TI-CMV and those with CMV DNAemia alone, by chart review. Inflammatory cytokines (IFNγ, TNFα, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, IL-18, and IL-1RA) were measured in residual plasma from these patients using a commercially available multiplex assay for at least two time points during the study period. Wilcoxon-Rank-Sum, logistic regression, and principal component analysis was performed comparing patients with and without probable TI-CMV.ResultsPatients with probable TI-CMV had significantly higher IL-18, TNFα, and IL-1β than patients with CMV DNAemia alone (p < 0.001, < 0.001, and < 0.05 respectively). When adjusting for transplant type and CMV recipient serostatus, elevations in TNFα (OR 1.43, 95% CI 1.07-1.92) and IL-18 (OR 2.00, 95% CI 1.06-3.75) were associated with increased odds of having probable TI-CMV. In principal component analysis the combination of CMV viral load, IL-18, TNFa, and IL-1β accounted for 80% of the variance in the data.ConclusionTNFα and IL-18 in combination with CMV viral load may be useful in predicting likelihood of TI-CMV. This is important in situations where tissue biopsies are not feasible, and adds to our diagnostic capability for TI-CMV in SOT recipients.Disclosures Stuart C. Ray, MD, miDiagnostics, Inc. (Board Member, Research Grant or Support) Robin K. Avery, MD, Aicuris (Scientific Research Study Investigator)Astellas (Scientific Research Study Investigator)Chimerix (Scientific Research Study Investigator)Merck (Grant/Research Support, Scientific Research Study Investigator)Oxford Immunotec (Scientific Research Study Investigator)Qiagen (Scientific Research Study Investigator)Takeda/Shire (Scientific Research Study Investigator)

1075. Absolute Lymphocyte Count as a Predictor of Cytomegalovirus (CMV) Infection and Recurrence in Hematopoietic Stem Cell Transplant (HSCT) Recipients

BackgroundCytomegalovirus (CMV) is a serious complication following Hematopoietic Stem Cell Transplant (HSCT) and can lead to serious organ disease and mortality. This study aimed to investigate the association between absolute lymphocyte count (ALC) and CMV to determine whether ALC could help to identify those at an increased risk of CMV infection and recurrenceMethodsAdults undergoing HSCT between 2011 and 2016 at Rigshospitalet, Denmark were included. Cox proportional hazards models investigated risk factors, including ALC, for CMV infection in the first year post-transplant and recurrent CMV infection 6 months after clearance and stopping CMV treatment for the first infection. For the primary outcome ALC was investigated as a time-updated risk factor lagged by 7 days, and for recurrent CMV, ALC measured at the time at the time of stopping treatment for the first CMV infection was investigated (+/- 7 days).ResultsOf the 352 HSCT recipients included, 57% were male, 40% received myeloablative conditioning, 42% had high risk (D-R+) CMV IgG serostatus at transplant and the median age was 56 (IQR 43-63). 143 (40.6%) patients had an episode of CMV DNAemia a median of 47 days after transplant (IQR 35-62). A lower current ALC (≤ 0.3 x109/L) was associated with a higher risk of CMV infection in univariate analysis compared to a high current ALC (> 1 x109/L). However, this association was attenuated after adjustment, particularly for acute graft versus host disease (Figure). 102 HSCT recipients were investigated for risk of recurrent CMV of which 41 (40.2%) had a recurrent CMV episode a median of 27 days (IQR 16-50) after stopping CMV treatment for the first infection. A lower ALC (≤ 0.3 x109/L) at the time of stopping CMV treatment was associated with a significantly higher risk of recurrent CMV after adjustment (Figure). A higher peak viral load (> 1500 IU/ml) during the first episode of CMV infection was also associated with an increased risk of recurrent CMV (aHR 2.47, 95%CI 1.00-6.10 compared to < 750 IU/ml).Association between absolute lymphocyte count (ALC) and risk of CMV infection and recurrent CMV within 6 months. **First CMV infection multivariable model also adjusted for sex, CMV serostatus, age, year of transplant, Charlson Comorbidity Index, Anti-thymocyte globulin (ATG) given, HLA donor-recipient matching, and acute graft versus host disease (time-updated) *Recurrent CMV infection multivariable model also adjusted for conditioning regimen, sex, CMV serostatus, age, year of transplant Anti-thymocyte globulin (ATG) given, HLA donor-recipient matching, and acute graft versus host disease and peak CMV viral load during the first CMV infectionConclusionA lower ALC at the time of stopping treatment for the first CMV infection was associated with an increased risk of recurrent CMV and could be used to help guide decisions for augmented CMV surveillance and clinical awareness of CMV disease symptoms in these patients. Disclosures All Authors: No reported disclosures

Early cytomegalovirus DNAemia and antiviral dose adjustment in high vs intermediate risk kidney transplant recipients.

Cytomegalovirus (CMV) infection continues to negatively affect outcomes for solid organ transplant recipients, despite the advent of strategies for preemptive surveillance and prophylaxis. The impact is especially great for CMV seronegative recipients of donor seropositive organs, who typically lack the ability to control CMV infection at the time of transplantation. We reviewed episodes of CMV DNAemia in a modern cohort of kidney transplant recipients over a 3-year period at a high-volume transplant center to investigate the frequency of DNAemia during antiviral prophylaxis. Despite receipt of antiviral prophylaxis per current guidelines, 75 cases of CMV DNAemia were observed in the first 100days after transplantation. For high risk patients, median time to DNAemia was 75days after transplantation, and the majority of patients had experienced dose-reduction of valganciclovir due to renal insufficiency. Review of CMV seropositive intermediate risk patients demonstrated DNAemia occurring earlier after transplantation compared with high risk patients with a median time of 64days (P=.029). The impact of valganciclovir dose adjustment was less notable in the intermediate risk group. Guidelines recommend beginning routine surveillance for CMV after the completion of antiviral prophylaxis. Our findings suggest that closer monitoring may be beneficial, especially for high risk patients at risk for DNAemia. Patients requiring dose adjustment of valganciclovir due to renal insufficiency may be at increased risk for CMV DNAemia. Improved methods for CMV prophylaxis and evaluation of immunologic risk for CMV DNAemia and disease are needed to improve patient outcomes after kidney transplantation.

Features of Cytomegalovirus DNAemia Blips in Allogeneic Hematopoietic Stem Cell Transplant Recipients: Implications for Optimization of Preemptive Antiviral Therapy Strategies.

Cytomegalovirus (CMV) DNAemia occurs frequently in allogeneic hematopoietic stem cell transplant recipients (allo-HSCT). There is limited information about the incidence, features, and clinical impact of CMV DNAemia blips (episodes defined by an isolated positive PCR result) in this setting. In this retrospective study, 225 consecutive adult patients undergoing any modality of allo-HSCT at our center between May 2012 and July 2019 were included. Plasma CMV DNA load was monitored using a highly sensitive real-time PCR assay. In all, 187 of 225 patients had CMV DNAemia through day 365 after allo-HSCT (total number of episodes, n=379). Eighty-three of the 187 patients had 1 or more blips (n=104). Blips occurred as a first episode of CMV DNAemia as opposed to prolonged CMV DNAemia (≥2 consecutive positive PCR results) in 47 patients; in 20 of these patients, blips represented the only documented episode throughout the study period, and in 27 patients, blips preceded a prolonged CMV DNAemia episode. In the remaining 36 patients, blips developed as recurrences. Blips presenting as initial episodes occurred more frequently (P < .001) in patients receiving an allograft from a CMV-seropositive donor. The cumulative incidence of recurrent CMV DNAemia following initial blips, self-resolving prolonged CMV DNAemia episodes, or CMV DNAemia episodes treated preemptively with antivirals was not significantly different (P=.34). Receiver operating characteristic curve analysis indicated that a CMV DNA load cutoff of 48 IU/mL yielded the highest combined sensitivity (66%) and specificity (70.2%) for predicting a prolonged CMV DNAemia episode. The practical implications of our data in the optimization of preemptive antiviral therapy strategies are discussed.

Incidence, features, and outcomes of cytomegalovirus DNAemia in unmanipulated haploidentical allogeneic hematopoietic stem cell transplantation with post-transplantation cyclophosphamide.

Conflicting data have been published as to the risk of cytomegalovirus (CMV) DNAemia and CMV disease in patients undergoing haploidentical hematopoietic stem cell transplantation (haplo-HSCT) with post-transplantation cyclophosphamide. We conducted a multicenter retrospective study including 118 patients subjected to unmanipulated haplo-HSCT to further clarify this issue. An historic cohort comprising 165 patients undergoing other transplant modalities (HLA-matched related, matched unrelated or mismatched) was built for comparison purposes. Plasma CMV DNA monitoring was performed using two highly sensitive real-time PCR assays. Overall, the cumulative incidence of CMV DNAemia, recurrent CMV DNAemia, and CMV DNAemia requiring preemptive antiviral therapy in patients undergoing haplo-HSCT was 63.9%, 34.9%, and 50.1%, respectively. These figures were rather comparable for other transplant modalities (P=.22, P=.13 and P=.72, respectively). A trend toward longer duration of episodes and shorter CMV DNA doubling times was observed in haplo-HSCT patients in comparison with other transplant modalities. Furthermore, median CMV DNA peak load was significantly higher in haplo-HSCTs (P=.008), yet overall mortality by day 180 and 365 was the same across comparison groups. There were five cases of CMV disease, and all occurred in haplo-HSCT patients. This latter observation is worrying and merits further investigation. The incidence of initial and recurrent episodes of CMV DNAemia either requiring or not antiviral therapy in unmanipulated haplo-HSCT was comparable to other transplant modalities in our cohort.

An investigation of the utility of plasma Cytomegalovirus (CMV) microRNA detection to predict CMV DNAemia in allogeneic hematopoietic stem cell transplant recipients.

Precise identification of patients at highest risk for developing Cytomegalovirus (CMV) DNAemia may improve CMV infection management in the allogeneic hematopoietic stem cell transplantation (allo-HSCT) setting. Here, we studied the potential use of detecting free CMV micro(mi)RNAs circulating in plasma for predicting CMV DNAemia in this clinical scenario. A total of 62 adult allo-HSCT recipients were included in this prospective observational multicenter study. Plasma CMV DNA load was monitored using the CMV RealTime CMV PCR (Abbott Molecular, Des Plaines, IL, USA). Detection of mature CMV miRNAs in plasma drawn by days + 7, + 14 and + 30 after allo-HSCT was performed using the miScript PCR System (Qiagen, Hilden, Germany). Assays could be optimized for five out of the seven targeted CMV miRNAs: UL36-5p, US33-5p, UL148D, UL22A-5p and UL112-3p. Of the 62 patients included in the study, 42 developed a first episode of CMV DNAemia at a median of 35days after allo-HSCT. All targeted CMV miRNA were detected early after transplantation, with CMV miRNA US33-5p and UL112-3p the most commonly found species at any time point; nevertheless, neither the detection rate of CMV miRNAs nor their abundance allowed discrimination between patients with subsequent CMV DNAemia and those with no CMV DNAemia. The data presented herein do not support any predictive utility of these CMV miRNAs for first episodes of CMV DNAemia in a cohort consisting primarily of allo-HSCT patients receiving haploidentical allografts.

S1640 POLYMORPHISMS IN CYTOKINES, CHEMOKINES AND THEIR RECEPTORS GENES CONTRIBUTED TO CYTOMEGALOVIRUS REACTIVATION AFTER ALLOGENEIC STEM CELL TRANSPLANTATION

Background: Cytomegalovirus (CMV) is the most common viral infection after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Without prophylaxis, 80% of CMV-seropositive patients experience CMV infection after allo-HSCT. Cytokines and chemokines are the first line of defence against viral infections and recent studies have shown that gene polymorphisms result in inter-individual differences in cytokine production. Aims: To determine the genotype of Cytokines and chemokines in the donor (D) and recipient (R) and their association with the CMV reactivation of patients receiving an allo-HSCT. Methods: Eighty-five patients who received allo-HSCT from an HLA-identical sibling donor from 2000 to 2015 were included. CMV DNAemia was evaluated until approximately day 100 after allo-HSCT. CMV reactivation was defined as the detection of CMV DNAemia ≥100 copies/ml in plasma. All of patients were classified according to epidemiological risk factors of clinical interest. Fifty genes were selected for their potential role in the pathogenesis of CMV: C-C Motif Chemokine Ligand (CCL) and Receptor (CCR), C-X-C Motif Chemokine Ligand (CXCL) and Receptor (CXCR), Forkhead Box P3 (FOXP3), CD48 Molecule (CD48), Interferon (IFN), Interleukin (IL), Killer Cell Immunoglobulin Like Receptor (KIR), Transforming Growth Factor Beta (TGFB), tumor necrosis factor alpha (TNF) and Lymphotoxin Alpha (LTA). Genomic DNA was purified from 170 PB (D and R) using Maxwell® RSC Blood DNA Kit (Promega, USA). Libraries were performed using an enrichment-capture gene panels according to the manufacturer's protocol. Paired-end 2x101 bp was performed using the Illumina HiSeq platform (Illumina, USA). Variants located in coding region, splicing sites and intronic polymorphisms were analyzed. Synonymous variants were excluded. During the variant calling process, the values of Depth, variant allele frequency (VAF) and the minor allele frequency (MAF) in European population are obtained and used to variants filtration (Depth ≥ 30, VAF ≥ 0.4 and MAF≥ 10%). Differences among groups were evaluated by Chi-Squared and Fischer Exact Test. Results: CMV reactivation was observed in 51/85 (60%) patients. Initial episodes of CMV DNAemia occurred at a median of 48 days (2–151) after transplant. Clinical variables are not associated with CMV reactivation: age, gender, stem cells, hematological disease, conditioning regimen, serology status and prior autologous transplant. Twenty-four patients (47.1%) had only one episode of CMV reactivation, 12 (23.5%) had two episodes and 15 (29.3%) had more than two episodes after allo-HSCT. Using filters discussing previously, 221 polymorphisms were detected in D and R. Although 209 variants studied had no apparent impact on the features of CMV reactivation, but we found that 12 variants in 7 different genes were associated significantly with the development or protection of CMV reactivation (Table 1). Our data showed that determined polymorphism in TNF-α, IL12A, TGBF2, IL1RN and CD48 play an important role to protection against CMV. On other hand, KIR3DL1 and CXCL12 represent examples of how receptor polymorphism can influence CMV development.Summary/Conclusion: The data presented suggest that screening of patients and donor pre-transplantation could help to predict the individual risk of the development of CMV infection. These results might also enable the identification of patients at high risk of CMV reactivation, enabling pre-emptive therapy or attempts to cure the infection by administrating antiviral therapy or CMV-specific T lymphocytes.

Kinetics of inflammatory biomarkers in plasma predict the occurrence and features of cytomegalovirus DNAemia episodes in allogeneic hematopoietic stem cell transplant recipients.

Cytomegalovirus (CMV) DNAemia occurs frequently in CMV-seropositive allogeneic hematopoietic stem cell transplant (allo-HSCT) recipients, and usually results from reactivation of latent infection established in the recipient. Predicting the occurrence of CMV DNAemia may be helpful in managing CMV infection in allo-HSCT recipients. Here, the kinetics of several inflammatory biomarkers in plasma were characterized and assessed for their potential value in anticipating the development and features of active CMV infection in allo-HSCT recipients, as documented using real-time PCR assays. The cohort consisted of 46 non-consecutive adult patients who underwent T-cell replete allo-HSCT at our center. Plasma levels of C-reactive protein (CRP), soluble tumor necrosis factor receptor type 2 (sTNF-R2), transforming growth factor-β1 (TGF-β1), and interferon-inducible protein 10 (IP-10/CXCL10) were measured in consecutive specimens obtained from conditioning either by nephelometry (CRP) or by specific immunoassays (the rest). Of the 46 patients, 22 had a first episode of CMV DNAemia at a median of 34 days after allo-HSCT (range, day 19-day 50). We found that both the TGF-β1 area under a curve (AUC) and peak levels were significantly lower in patients who subsequently developed CMV DNAemia than in patients with no CMV DNAemia. Interestingly, CRP but not TGF-β1 AUC and peak levels predicted the occurrence of CMV DNAemia episodes requiring preemptive antiviral therapy. The data presented herein suggest that kinetics of inflammatory biomarkers in plasma might be useful to anticipate post-engraftment CMV DNAemia episodes and predict the need for preemptive antiviral therapy in allo-HSCT recipients.

Spontaneously-resolving episodes of cytomegalovirus DNAemia in allogeneic hematopoietic stem cell transplant recipients: Virological features and clinical outcomes.

It has been reported that low-plasma cytomegalovirus (CMV) DNA loads are associated with an increased risk of overall mortality in allogeneic hematopoietic stem cell transplantation (allo-HSCT). Utilizing a conservative strategy for initiation of preemptive antiviral therapy (>1500 IU/mL), we characterized the virological features of spontaneously-resolving episodes of CMV DNAemia and assessed their impact on mortality through the first year after transplantation. We reviewed the CMV DNA polymerase chain reaction results and clinical charts of 230 consecutive adult patients who underwent T-cell replete allo-HSCT at our center. A total of 280 episodes of CMV DNAemia were registered in 164 patients, of which 144 episodes cleared spontaneously. Clearance of CMV DNAemia was significantly delayed in initial and recurrent self-resolving episodes featuring CMV DNA peak loads > 250 IU/mL compared with those displaying lower values. All-cause mortality in patients with self-resolving episodes of CMV DNAemia was comparable (P = 0.7) to that in patients with no CMV DNAemia and was not related to the CMV DNA peak load (≥250 IU/mL vs <250 IU/mL) (P = 0.6). In summary, in our setting, the magnitude of the CMV DNA peak load reached during self-resolving episodes of CMV DNAemia correlated directly with duration of episodes, but had no apparent impact on all-cause mortality taking patients with no documented CMV DNAemia as a reference.

Factors influencing cytomegalovirus DNA load measurements in whole blood and plasma specimens from allogeneic hematopoietic stem cell transplant recipients

We assessed the impact of several parameters, including the nature of the episode of Cytomegalovirus (CMV) DNAemia, the use of preemptive antiviral therapy, and the blood cell content in CMV DNA loads measured in whole blood (WB) and plasma (PL). CMV DNA load was quantified in 245 paired specimens collected within 43 postengraftment episodes of CMV DNAemia by using the CMV RealTime CMV PCR (Abbott Molecular). Concordant categorical results were obtained for 78.4% of paired specimens (Kappa index, 0.385; P = <0.001). Overall, CMV DNA loads in PL were higher than those in WB (mean bias, +0.115 log IU/mL) in both initial and recurrent episodes; this was so in post-antiviral treatment but not in pretreatment paired specimens. Median CMV DNA doubling time values in both compartments were not significantly different. Leukocyte counts had a significant impact on the comparability of CMV DNA loads measured in both matrices.

Impact of cytomegalovirus DNAemia on overall and non-relapse mortality in allogeneic stem cell transplant recipients.

We conducted a retrospective single-center study to investigate the potential impact of cytomegalovirus (CMV) DNAemia on mortality in allogeneic stem cell transplant (allo-SCT) recipients. A total of 151 consecutive patients who underwent T-cell replete allo-SCT were included in the study. Patients with CMV DNAemia were treated preemptively with antivirals upon detection of plasma CMV DNA loads >1500IU/mL. At least one episode of CMV DNAemia occurred in 109 (72.2%) patients, and 67 of these patients (61.5%) required one or more courses of antiviral therapy. The cumulative incidence of 1-year overall and non-relapse mortality (NRM) was 28.5% (95% confidence interval [CI], 18.4%-39.5%) and 23.2% (95% CI 12.81%-35.4%), respectively. The occurrence of either CMV DNAemia or CMV recurrences had no apparent effect on 1-year overall mortality and NRM; nevertheless, a trend towards an increased risk of death was seen in patients with one or more episodes of CMV DNAemia requiring antiviral therapy (hazard ratio [HR], 2.10; 95% CI, 0.96-4.61; P=.06 for overall mortality, and HR, 2.36; 95% CI, 0.96-5.76; P=.06 for NRM) but not in those displaying one or more self-resolving episodes. Therefore, the data suggest that withholding preemptive antiviral therapy until the plasma CMV DNA load reaches 1500 IU/mL has no apparent detrimental effect on patient survival.

Lack of evidence for a reciprocal interaction between bacterial and cytomegalovirus infection in the allogeneic stem cell transplantation setting.

Pathogenic interactions between bacteria and cytomegalovirus (CMV) may potentially occur early after allogeneic stem cell transplantation (allo-SCT). This possibility nevertheless has not been investigated in depth. This was a retrospective study that included 170 consecutive patients who underwent 173 allo-SCTs. Both bacterial infection (most of which were bacteremic) and CMV DNAemia were detected in 78 allo-SCTs (62.9%). In total, 51 and 32 episodes of bacterial infection preceded or occurred after CMV DNAemia detection, respectively. Both events were diagnosed concurrently in four allo-SCTs. The cumulative incidence of bacterial infection (of any type) over the study period was comparable in patients with or without a preceding episode of CMV DNAemia (P = 0.321). Cox proportional hazards regression analysis failed to identify CMV DNAemia as a significant risk factor for bacterial infection. Likewise, the cumulative incidence of CMV DNAemia within the study period was not significantly different in patients with or without a preceding episode of bacterial infection (P = 0.189). Furthermore, the occurrence of bacterial infection within episodes of active CMV infection had no apparent impact on the kinetics of CMV DNAemia. Our data, thus, do not support the existence of a bidirectional synergistic effect between bacterial infection and active CMV infection in the allo-SCT setting.

Role of cytomegalovirus (CMV)-specific polyfunctional CD8+ T-cells and antibodies neutralizing virus epithelial infection in the control of CMV infection in an allogeneic stem-cell transplantation setting.

The role of cytomegalovirus (CMV)-specific polyfunctional CD8+ T-cells and that of antibodies neutralizing virus epithelial infection (AbNEI) in the control of CMV DNAemia were investigated in 39 CMV-seropositive allogeneic stem-cell transplant (Allo-SCT) recipients with (n = 24) or without (n = 15) CMV DNAemia. AbNEI levels were monitored prospectively by means of a neutralization assay employing retinal epithelial cells (ARPE-19) and the recombinant CMV strain BADrUL131-Y4. Quantification of CMV-specific polyfunctional CD8+ T-cells (expressing two or three of the following markers: IFN-γγ, TNF-α and CD107a) in whole blood was performed by flow cytometry for intracellular cytokine staining. We found no differences in the dynamic pattern of AbNEI in patients with or without subsequent CMV DNAemia. Baseline and peak AbNEI titres were not predictive of the dynamics of CMV replication within episodes. No correlation was found between CMV DNA loads and AbNEI levels during episodes of CMV DNAemia (ρ = 0.09; 95 % confidence interval - 0.52 to 0.64; P = 0.78). The detection of pp65/IE-1 CMV-specific polyfunctional CD8+ T-cells was associated with low-level virus replication within subsequent episodes of CMV DNAemia. Interestingly, the presence of AbNEI titres (inverse) >4.7 log2 was predictive of the occurrence of CMV DNAemia (sensitivity, 83 %; specificity, 80 %). Our findings provide an insight to the role of humoral and cellular immunity in the control of CMV infection in an Allo-SCT setting.

Enumeration of NKG2C+ natural killer cells early following allogeneic stem cell transplant recipients does not allow prediction of the occurrence of cytomegalovirus DNAemia.

The role of Natural killer (NK) cells in the control of cytomegalovirus (CMV) infection in allogeneic stem cell transplant recipients has not been precisely characterized. The current study is aimed at investigating the potential role of NK cells expressing the activating receptor NKG2C in affording protection against the development of CMV DNAemia in patients exhibiting detectable CMV-specific CD8(+) T-cell responses early following transplantation. A total of 61 nonconsecutive patients were included in the study. Peripheral levels of CD56(bright) CD16(-/low) and CD56(dim) CD16(+) NKG2C(+) NK cells and CMV pp65/IE-1-specific IFN-γ-producing CD8(+) T-cells were enumerated by flow cytometry at days +30 and +60 after transplant. Neither the absolute number of NKG2C(+) NK cells, nor that of CD56(bright) CD16(-/low) and CD56(dim) CD16(+) NKG2C(+) NK-cell subsets at day 30 differed significantly between patients with or without subsequent CMV DNAemia. No significant correlation was found between levels of both NKG2C(+) NK-cell populations and the peak CMV DNA load within subsequent episodes of CMV DNAemia. The data indicate that enumeration of NKG2C(+) NK cells early after transplant is unlikely to be helpful in identifying those patients at highest risk of developing CMV DNAemia. Moreover, the data do not support a direct implication of NKG2C(+) NK cells in preventing the development of CMV DNAemia.

Monitoring of trough plasma ganciclovir levels and peripheral blood cytomegalovirus (CMV)-specific CD8+ T cells to predict CMV DNAemia clearance in preemptively treated allogeneic stem cell transplant recipients.

It is uncertain whether monitoring plasma ganciclovir (GCV) levels is useful in predicting cytomegalovirus (CMV) DNAemia clearance in preemptively treated allogeneic stem cell transplant recipients. In this observational study, including 13 episodes of CMV DNAemia treated with intravenous (i.v.) GCV or oral valganciclovir, we showed that monitoring trough plasma GCV levels does not reliably predict response to therapy. Rather, immunological monitoring (pp65 and immediate-early [IE]-1-specific gamma interferon [IFN-γ]-producing CD8+ T cells) appeared to perform better for this purpose.

A single-center experience of cytomegalovirus infections in Asian pediatric patients undergoing allogeneic hematopoietic stem cell transplant for leukemia in Singapore.

Cytomegalovirus (CMV) infection remains a significant cause of morbidity and mortality in pediatric patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT) despite improved surveillance and the current preemptive approach. Few data on its prevalence in the Asian pediatric population exist. We retrospectively reviewed the prevalence of CMV infections in 33 patients with 37 transplants who received HSCT for leukemia from 1998 to 2008, and who were managed preemptively for infections. In the 37 transplants, 16 patients (43%) had CMV DNAemia. Of the patients who were CMV seropositive before transplant and received stem cells from seropositive donors (R+/D+), 69% had DNAemia; of those who received stem cells from seronegative donors (R+/D-), 36% had CMV DNAemia. Of the patients who were CMV naïve before transplant and received stem cells from seropositive donors (R-/D+), 25% had CMV DNAemia. In CMV-seronegative donor-recipient transplants (R-/D-), 20% of patients had CMV DNAemia. The median time to the first episode of CMV DNAemia was 21 (range: 10-107) days after the transplants, and the median duration of CMV DNAemia was 22 (range: 2-315) days. CMV DNAemia recurred in 44% (7 of 16) of these patients. Only 1 patient developed CMV disease (retinitis). No deaths were related to CMV infections. CMV infection manifesting as DNAemia is a common complication in pediatric patients undergoing allogeneic HSCT for leukemia. Pre-transplant serostatus predicts reactivation risks; invasive CMV disease is rare using the preemptive approach in our patient population.

An evaluation of the role of NKG2C+ natural killer cells in protection from cytomegalovirus DNAemia early following allogeneic stem cell transplantation

The role of natural killer (NK) cells in affording protection against human cytomegalovirus (CMV) in allogeneic stem cell transplant recipients is largely unknown. The current study was aimed at determining whether NKG2C+ NK cells confer protection from CMV DNAemia early following transplantation in patients lacking mono and polyfunctional CMV pp65 and IE-1-specific CD4+ and CD8+ T-cell responses, as measured by flow cytometry for intracellular cytokine staining. Fourteen out of the 36 patients included in this study developed CMV DNAemia between days +30 and +60 after transplant. Three patients did so after day +60. Peripheral blood levels of CD56(bright) CD16(-/low) and CD56(dim) CD16+ NKG2C+ NK cells measured at day +30 and at day +60 in patients who had or had not subsequent CMV DNAemia did not differ significantly. In addition, no significant correlation was found between CD56(bright) CD16(-/low) (σ = -0.229; P = 0.39) and CD56(dim) CD16+ (σ = -0.285; P = 0.28) NKG2C+ NK-cell levels and initial plasma CMV DNA loads. In summary, the data presented do not support a direct implication of NKG2C+ NK cells in preventing the development of CMV DNAemia or modulating the magnitude of CMV replication at early stages during episodes of CMV DNAemia in allogeneic stem cell transplant patients with unreconstituted CMV-specific T-cell responses.

Functional profile of cytomegalovirus (CMV)‐specific CD8+ T cells and kinetics of NKG2C+ NK Cells associated with the resolution of CMV DNAemia in allogeneic stem cell transplant recipients

Immune mechanisms involved in control of cytomegalovirus (CMV) infection in the allogeneic stem cell transplantation setting have not been fully disclosed. CMV pp65 and IE-1-specific CD8(+) T cells expressing IFN-γ, TNF-α, and CD107a, alone or in combination, and NKG2C(+) NK cells were prospectively enumerated during 13 episodes of CMV DNAemia. The expansion of monofunctional and polyfunctional CD8(+) T cells was associated with CMV DNAemia clearance. The size and functional diversity of the expanding CD8(+) T-cell population was greater in self-resolved episodes than in episodes treated with antivirals. These differences were related to the magnitude of expansion of cognate antigen IFN-γ CD4(+) T cells. The resolution of CMV DNAemia was associated frequently with a marked expansion of both CD56(dim) /CD16(+) NK cells and NKG2C(+) CD56(bright) /CD16(-) NK cells. The data lend support to the role of polyfunctional CD8(+) T cells in controlling CMV replication in the allogeneic stem cell transplantation setting, and suggest that NKG2C(+) NK cells may be involved critically in the resolution of CMV DNAemia episodes.