Impact of stem cell graft on early viral infections and immune reconstitution after allogeneic transplantation in adults.

BACKGROUND: Viral infections are well-known complications after allogeneic stem cell transplant (allo-SCT). OBJECTIVES: We compared prospectively incidences of DNAemia and active infections (AI) for five opportunistic viruses (Human Herpesvirus 6 (HHV-6), Epstein-Barr virus (EBV), BK polyomavirus (BKPyV), Cytomegalovirus (CMV) and Adenovirus (ADV)) and kinetics of immune reconstitution (IR) in adults receiving either double umbilical cord blood (dUCB group) or unrelated peripheral blood stem cell (uPBSC group) allo-SCT after a reduced-intensity conditioning (RIC) regimen. STUDY DESIGN: Whole blood samples were collected at transplant, every 15days during the first 3 months and at 4, 5 and 6 months post-transplant. RESULTS: Sixty-five patients were enrolled (uPBSC n=34; dUCB n=31). Incidences of HHV-6 and BKPyV DNAemia were significantly higher for dUCB (97% vs 23.5% and 58% vs 32%, respectively) while EBV DNAemia was more frequently detected in uPBSC (71% vs 26%). The incidence of CMV DNAemia was similar between both groups. ADV AI developed only in dUCB. HHV-6 AI were also higher in dUCB (84% vs 21%). In multivariate analysis, dUCB graft was the only independent factor associated with HHV-6 DNAemia (OR: 19.0; 95%CI: 5.2-69.1; p<0.0001) while EBV DNAemia were significantly associated with uPBSC (OR: 29.9; 95%CI: 5.68-158; p <0.0001). dUCB graft was also the only factor associated with HHV-6 AI. Finally, higher counts and faster recoveries of B lymphocytes (p<0.0001) and monocytes (p=0.0007) were observed in the dUCB group. CONCLUSION: We demonstrate a strong correlation between sources of graft and patterns of viral DNAemia and AI and IR after RIC allo-SCT.

Human herpesvirus 6 reactivation before engraftment is strongly predictive of graft failure after double umbilical cord blood allogeneic stem cell transplantation in adults.

Our main objective was to determine new factors associated with engraftment and single-unit predominance after double umbilical cord blood (UCB) allogeneic stem-cell transplantation. Engraftment occurred in 78% of cases in this retrospective study including 77 adult patients. Three-year overall survival, disease-free survival, relapse incidence, and nonrelapse mortality were 55 ± 6%, 44 ± 6%, 33 ± 5%, and 23 ± 4%, respectively. In multivariate analysis, Human herpesvirus 6 reactivation during aplasia (hazard ratio [HR] = 2.63; 95% confidence interval [CI]: 1.64-4.17; p < 0.001), younger recipient age (<53 years) (HR = 1.97; 95% CI: 1.16-3.35; p = 0.012), and lower human leukocyte antigen matching between the two units (3 of 6 or 4 of 6) (HR = 2.09; 95% confidence interval: 1.22-3.59; p = 0.013) were the three factors independently associated with graft failure. Also, factors independently predicting the losing UCB unit were younger age of the UCB unit (odds ratio [OR] = 1.01; 95% CI: 1-1.02; p = 0.035), lower CD34(+) cell dose contained in the UCB unit (≤ 0.8 × 10(5)/kg) (OR = 2.55; 95% CI: 1.05-6.16; p = 0.04), and presence of an ABO incompatibility between the UCB unit and the recipient (OR = 2.53; 95% CI: 1.15-5.53; p = 0.02). Thus, Human herpesvirus 6 reactivation during aplasia, lower unit-unit human leukocyte antigen matching, and younger UCB unit age, as new unfavorable predictive factors, may represent new parameters to take into account after double UCB allogeneic stem-cell transplantation in adults. These results need to be confirmed prospectively, as they may influence unit selections and patient outcomes.

Long-lasting HHV-6 reactivation in long-term adult survivors after double umbilical cord blood allogeneic stem cell transplantation.

Higher incidence of human herpesvirus 6 (HHV-6) infection has been documented after umbilical cord blood allo-transplant in adults. Here we demonstrate that HHV-6 reactivation persists for a very long time in half of the patients after this type of graft. Long-term immune reconstitution does not explain this event, which remains to be explained.

Quantification of two viral transcripts by real time PCR to investigate human herpesvirus type 6 active infection.

BACKGROUND: Human herpesvirus 6 (HHV-6) causes exanthema subitum and is associated with symptomatic reactivations in immunocompromised patients, particularly after hematopoietic stem cell transplantation. The detection of viral mRNA can help to make the difference between latent, chromosomally integrated and true replicating virus. It can also be a useful tool to investigate viral multiplication in different cell types. OBJECTIVES: To develop molecular tools for the detection and quantification HHV-6 transcripts that can be used in a clinical setting. STUDY-DESIGN: Two one-step reverse-transcriptase quantitative real-time PCR (RT-qPCR) were developed for the quantification of the immediate early U90 and the late U100 mRNAs. Viral mRNA loads were compared to viral DNA loads during infection in vitro and in blood samples collected from stem cell transplanted patients. RESULTS: Analytical performances of the two quantitative real-time PCR were good. In vitro, kinetics of both transcripts was well correlated with DNA levels. Sixty blood samples from patients with active HHV-6 infection were analyzed. Overall agreement of qualitative results for HHV-6 DNA, U90 RNA and U100 RNA was good. HHV-6 DNA loads were significantly higher than mRNA loads. In clinical samples, the amounts of U100 and U90 mRNAs were low and their detection was mainly associated to viral DNA loads upper than 1000 copies/ml of blood. CONCLUSION: The new assays are sensitive and reliable methods for the monitoring of viral transcription in vitro and in vivo. As their detection is associated to high DNA loads in vivo, they can be helpful tools for the diagnosis of active infection.

HHV-6 cell receptor CD46 expression on various cell subsets of six blood and graft sources: a prospective series.

BACKGROUND: Cord Blood (CB) are increasingly used as an alternative stem cells source in adults for allogeneic Stem Cell Transplantation (allo-SCT). The risk of human herpesvirus (HHV-6) reactivation is significantly higher after CB transplant vs unrelated peripheral blood stem cells (PBSC) allo-SCT. Higher HHV-6 cell receptor CD46 expression on progenitor cells in CB may explain this difference. OBJECTIVES: To prospectively compare the HHV-6 cell receptor CD46 expression on various cell subsets of three freshly harvested blood sources on one hand and of three graft sources on the other hand. STUDY DESIGN: 52 samples were used for the purpose of this study. They were issued from peripheral blood (PB, n = 10), G-CSF mobilised PB (GCSF-PB, n = 10), cord blood (CB, n = 10), unmanipulated bone marrow (uBM, n = 5), leukapheresis product (LP, n = 10) and thawed CB graft (n = 7). CD46 expression was assessed by FACS analysis on total lymphocytes, monocytes, NK cells, T and B cells subsets, plasmacytoid (pDCs) dendritic cells and stem cells. RESULTS: As all cell subsets were found CD46 positive, CD46 mean fluorescence intensity (MFI) was then considered for comparison between the three blood sources and the three graft sources. The most impressive result observed was that HHV-6 cell receptor CD46 expression was significantly reduced in almost all cell components of thawed CB graft compared to other graft sources. CONCLUSIONS: This original study shows strong differences in term of quantitative CD46 expression between several blood and grafts samples. Our results suggest that other factors than the qualitative CD46 expression play a role in the higher HHV-6 reactivation observed after CB transplant in adults.

Characterization of various blood and graft sources: a prospective series.

BACKGROUND: Studies comparing cell components of blood and graft sources are very scarce. We present here a thorough study examining the cellular content of various sources of blood and cell therapy products. STUDY DESIGN AND METHODS: We have prospectively compared by fluorescence-activated cell sorting analyses the cellular composition of three blood sources on the one hand--peripheral blood (PB; n = 10) versus granulocyte-colony-stimulating factor (G-CSF)-mobilized PB (GCSF-PB, n = 10) versus cord blood (CB, n = 10)--and of three graft sources on the other hand--unmanipulated bone marrow (uBM, n = 5) versus leukapheresis product (LP, n = 10) versus thawed CB graft (n = 7). RESULTS: All median absolute numbers of cell subsets were found significantly higher in GCSF-PB and LP, except for monocytoid dendritic cells (mDCs) in CB and uBM. The most impressive results were the median quantities of memory T and B lymphocytes but also of plasmacytoid DCs (pDCs) contained in LP compared to thawed CB graft, with ratios of 375, 318, and 247, respectively. The proportions of naive and CD4+/CD8- T cells, transitional B cells, and CD5+ and naive B lymphocytes were found significantly higher in CB samples while the proportions of mDCs and pDCs were found significantly lower. CONCLUSION: Our study shows strong differences in terms of quantitative and qualitative cellular composition between several blood or graft sources, possibly explaining the differences observed in terms of outcomes after transplant.

Occurrence and molecular characterization of Klebsiella pneumoniae ST37 clinical isolates producing plasmid-mediated AmpC recovered over a 3-year period.

We investigated the clinical and microbiological epidemiology of AmpC plasmidic cephalosporinases (pAmpC) in Klebsiella pneumoniae strains resistant to ceftazidime, during a 3-year period (2007-2009). Among 1505 K. pneumoniae, 7 were pAmpC producers. Molecular characterization revealed the spread of a ST37 strain producing DHA-1 within intensive care units and the diffusion of the same plasmid among unrelated strains.

In vitro antimicrobial activity of "last-resort" antibiotics against unusual nonfermenting Gram-negative bacilli clinical isolates.

In this prospective multicentric study, we assessed the in vitro antimicrobial activity of carbapenems (imipenem, meropenem, and doripenem), tigecycline, and colistin against 166 unusual nonfermenting Gram-negative bacilli (NF-GNB) clinical isolates collected from nine French hospitals during a 6-month period (from December 1, 2008, to May 31, 2009). All NF-GNB isolates were included, except those phenotypically identified as Pseudomonas aeruginosa or Acinetobacter baumannii. Minimal inhibitory concentrations (MICs) of antimicrobial agents were determined by using the E-test technique. The following microorganisms were identified: Stenotrophomonas maltophilia (n=72), Pseudomonas spp. (n=30), Achromobacter xylosoxidans (n=25), Acinetobacter spp. (n=18), Burkholderia cepacia complex (n=9), Alcaligenes faecalis (n=7), and Delftia spp. (n=5). All isolates of Acinetobacter spp., A. faecalis, and Delftia spp. were susceptible to the three carbapenems. Imipenem exhibited the lowest MICs against Pseudomonas spp., and meropenem, as compared with imipenem and doripenem, displayed an interesting antimicrobial activity against A. xylosoxidans and B. cepacia complex isolates. Conversely, no carbapenem exhibited any activity against S. maltophilia. Except for S. maltophilia isolates, tigecycline and colistin exhibited higher MICs than carbapenems, but covered most of the microorganisms tested in this study. To our knowledge, no prior study has compared antimicrobial activity of these five antibiotics, often considered as "last-resort" treatment options for resistant Gram-negative infections, against unusual NF-GNB clinical isolates. Further studies should be carried out to assess the potential clinical use of these antibiotics for the treatment of infections due to these microorganisms.

Stenotrophomonas maltophilia--the most worrisome threat among unusual non-fermentative gram-negative bacilli from hospitalized patients: a prospective multicenter study.

OBJECTIVES: Isolation rates of unusual non-fermentative Gram-negative bacilli (i.e. other than Pseudomonas aeruginosa and Acinetobacter baumannii) are increasing but studies are limited to few observations. We aimed at determining risk factors for infection and influence of antibiotic treatment on the outcome. METHODS: A six-month (December 1, 2008-May 31, 2009) prospective multicenter cohort study was conducted in nine French teaching hospitals. Characteristics of patients colonized or infected by unusual NF-GNB, adequacy of antimicrobial therapies, and outcome were analyzed. RESULTS: Analysis of 158 patients (median age, 62.7 years) was conducted. Stenotrophomonas maltophilia was the predominant bacterial species isolated (39%) followed by Achromobacter group (15%) and non-baumannii Acinetobacter species (13%). Compared to colonized patients, infected ones were more frequently immunocompromised [relative risk (RR) = 1.63, (95% confidence interval (CI) = 1.02-2.60, P = 0.05)], hospitalized within the last three months [RR 1.67 (95% CI 1.09-2.58, P = 0.02)], admitted in an intensive care unit with central venous catheter [RR 1.74 (95% CI 1.15-2.63, P = 0.01)]. The overall hospital mortality concerned 28 patients (18%) but no association with inadequate antimicrobial treatment was found except in the group of S. maltophilia infected cases [RR 2.81 (95% CI 1.01-7.83, P = 0.02)]. CONCLUSION: Naturally carbapenems-resistant S. maltophilia is the main unusual NF-GNB pathogen in hospitalized patients, leading to inappropriate empirical antibiotic treatment at the time of emerging extended-spectrum ß-lactamase-producing bacteria.