No abstract

Rationale: Lower airway (LAW) infection with Pseudomonas aeruginosa and Staphylococcus aureus is the leading cause of morbidity in cystic fibrosis (CF). The upper airways (UAW) were shown to be a gateway for acquisition of opportunistic bacteria and to act as a reservoir for them. Therefore, tools for UAW assessment within CF routine care require evaluation. Objectives: The aims of the study were non-invasive assessment of UAW and LAW microbial colonisation, and genotyping of P aeruginosa and S aureus strains from both segments. Methods: 182 patients with CF were evaluated (age 0.4-68 years, median 17 years). LAW specimens were preferably sampled as expectorated sputum and UAW specimens by nasal lavage. P aeruginosa and S aureus isolates were typed by informative single nucleotide polymorphisms (SNPs) or by spa typing, respectively. Results: Of the typable S aureus and P aeruginosa isolates from concomitant UAW-and LAW-positive specimens, 31 of 36 patients were carrying identical S aureus spa types and 23 of 24 patients identical P aeruginosa SNP genotypes in both compartments. Detection of S aureus or P aeruginosa in LAW specimens was associated with a 15-or 88-fold higher likelihood also to identify S aureus or P aeruginosa in a UAW specimen from the same patient. Conclusions: The presence of identical genotypes in UAW and LAW suggests that the UAW play a role as a reservoir of S aureus and P aeruginosa in CF. Nasal lavage appears to be suitable for non-invasive UAW sampling, but further longitudinal analyses and comparison with invasive methods are required. While UAW bacterial colonisation is typically not assessed in regular CF care, the data challenge the need to discuss diagnostic and therapeutic standards for this airway compartment. Trial registration number: NCT00266474.

Serial analysis of chimerism reliably identifies patients at highest risk to relapse. The 3-year EFS of patients with increasing MC without immunotherapy was 0%, by which overt relapse could be prevented in a considerable group of patients.

Summary:Within a prospective study we analyzed hematopoietic chimerism in serial peripheral blood samples taken from 55 patients with acute leukemias (ALL 21, AML 20, MDS 14) with a median age of 13.5 years at very short time intervals following allogeneic bone marrow transplantation (allo-BMT). The investigation was performed to determine the implications of mixed hematopoietic chimerism (MC) with regard to the clinical outcome in patients with acute leukemias after allo-BMT. Analysis of chimerism was performed by PCR of variable number of tandem repeat (VNTR) sequences with a maximum sensitivity of 0.8%. Thirteen male patients transplanted with the marrow of a female donor were also studied by amplification of a Y-chromosomespecific alphoid repeat (0.1-0.01% sensitivity). VNTR analysis in 55 patients revealed complete chimerism (CC) in 36 cases, MC in 18 follow-ups and autologous recovery in one patient. Quantitative analysis of MC identified 10/18 patients with increasing autologous patterns in whom 9/10 subsequently relapsed. The patient with autologous recovery relapsed as well. Eight of 18 patients with MC showed decreasing amounts of autologous DNA and became CC upon further follow-up. In contrast, only 7/36 patients with CC in the prior analysis of chimerism status relapsed. However, in 4/7 patients the interval between last CC confirmation and relapse was more than 4 months. In 2/7 patients autologous DNA was not detectable in peripheral blood but in bone marrow aspirates. One of these seven patients developed a fulminant relapse within 3 weeks. The probability of relapse-free survival for patients with CC is 0.67 (n = 36), for patients with decreasing MC 1.0 (n = 8) and for patients with increasing MC 0.1 (n = 10). In summary, the results demonstrate that serial and quantitative chimerism analysis at short time intervals by PCR provides a reliable and rapid screening method for the early detection of recurrence of underlying dis- ease and is therefore a prognostic tool to identify patients at highest risk of relapse.

We have retrospectively investigated the relationship between the level of minimal residual disease (MRD) detected in bone marrow taken prior to conditioning therapy and outcome following stem cell transplantation for high risk childhood ALL. Forty-one patients, in whom both a molecular marker of MRD and sufficient archival material was available, were included in the study. All were in remission at BMT: eight in CR1, 32 in CR2 and five in greater than CR2. MRD was measured by PCR amplification of antigen receptor gene rearrangements and clone-specific oligoprobing, the median sensitivity of detection being one leukaemic cell in 10 000 normals. Results were classified as high-level positive (if a clonal band was evident after electrophoresis), low-level positive (if MRD was detected only after oligoprobing) and negative. MRD was detected at high levels in 17 patients, at low levels in 10 patients and 14 patients were MRD negative at the time of transplant. The 5-year event-free survival for these groups was 23%, 48% and 78%, respectively (P = 0.022). Limited multivariate analysis confirmed the significance of MRD (P = 0.0095) vs CR status, donor type, sex, immunophenotype and acute GvHD. This study confirms the strong relationship between MRD level and outcome following allogeneic transplantation. In contrast to a previous study we observed that a minority of children with high-level pre-BMT MRD can enter long lasting remission. The possible role for acute GVHD coupled with a graft-versus-leukaemia effect in the clearance of high level MRD in patients with ALL is discussed.