Invasive pulmonary aspergillosis: comparative analysis in cancer patients with underlying haematologic malignancies versus solid tumours.

BACKGROUND: Invasive pulmonary aspergillosis (IPA) is commonly associated with haematologic malignancies but also occurs with solid tumours. AIM: To compare the diagnostic approaches and therapeutic outcomes for IPA between patients with haematologic malignancies and solid cancers. METHODS: A retrospective study was conducted evaluating consecutive cases of proven and probable IPA from 2004 to 2016. Patients >18 years of age with an underlying solid tumour, haematologic malignancy, or haematopoietic cell transplantation (HCT) within one year of IPA diagnosis were included. FINDINGS: Of the 311 patients analysed, 225 had haematologic malignancies and 86 had solid tumours. Patients with solid tumours were more likely to have had chronic obstructive pulmonary disease (COPD) or other pulmonary diseases, have Aspergillus fumigatus infections, and have received radiotherapy before IPA occurrence than were those with haematologic malignancies (all P<0.01). Antifungal monotherapy and voriconazole-based therapy were more often prescribed in the solid group (87% vs 56%, P<0.0001, and 77% vs 53%, P=0.0002, respectively). The median duration of primary antifungal therapy was longer in the solid group (64 days vs 20 days, P<0.0001). Complete or partial response to antifungal therapy was recorded in 66% of the solid group and 40% of the haematologic group (P=0.0001). At 12 weeks, overall mortality was similar in both groups, but IPA-attributable mortality was higher in the haematologic group (30% vs 18%, P=0.04). CONCLUSIONS: Monotherapy was more often prescribed in patients with solid tumours than in patients with haematologic malignancies. Patients with solid tumours had better antifungal therapy response and lower 12-week IPA-attributable mortality than did those with haematologic malignancies.

Whole-genome sequencing of Staphylococcus epidermidis bloodstream isolates from a prospective clinical trial reveals that complicated bacteraemia is caused by a limited number of closely related sequence types.

OBJECTIVES: The significance of isolating Staphylococus epidermidis from a blood culture is highly heterogeneous, ranging from contamination to an indication of a serious infection. Herein we sought to determine whether there is a relationship between S. epidermidis genotype and clinical severity of bacteraemia. METHODS: S. epidermidis bacteraemias from a prospective, multicentre trial at 15 centres in the United States and one in Spain were classified as simple (including possible contamination), uncomplicated, and complicated. Whole-genome sequencing (WGS) was performed on 161 S. epidermidis isolates, and clinical outcomes were correlated with genotypic information. RESULTS: A total of 49 S. epidermidis sequence types (STs) were identified. Although strains of all 49 STs were isolated from patients with either simple or uncomplicated infection, all strains causing complicated infections were derived from five STs: ST2, ST5, ST7, ST16, and ST32. ST2 and ST5 isolates were significantly more likely to cause uncomplicated and complicated bloodstream infections compared to simple bacteraemia (odds ratio 2.0, 95%CI 1.1-3.9, p 0.04). By multivariate regression analysis, having an ST2 or ST5 S. epidermidis bacteraemia was an independent predictor of complicated bloodstream infection (odds ratio 3.7, 95%CI 1.2-11.0, p 0.02). ST2/ST5 strains carried larger numbers of antimicrobial resistance determinants compared to non-ST2/ST5 isolates (6.34 ± 1.5 versus 4.4 ± 2.5, p < 0.001). CONCLUSION: S. epidermidis bacteraemia was caused by a genetically heterogeneous group of organisms, but only a limited number of STs-particularly multidrug-resistant ST2 and ST5 strains-caused complicated infections.

Banff Lung Report: Current knowledge and future research perspectives for diagnosis and treatment of pulmonary antibody-mediated rejection (AMR).

The Lung session of the 2017 14th Banff Foundation for Allograft Pathology Conference, Barcelona focused on the multiple aspects of antibody-mediated rejection (AMR) in lung transplantation. Multidimensional approaches for AMR diagnosis, including classification, histological and immunohistochemical analysis, and donor- specific antibody (DSA) characterization with their current strengths and limitations were reviewed in view of recent research. The group also discussed the role of tissue gene expression analysis in the context of unmet needs in lung transplantation. The current best practice for monitoring of AMR and the therapeutic approach are summarized and highlighted in this report. The working group reached consensus of the major gaps in current knowledge and focused on the unanswered questions regarding pulmonary AMR. An important outcome of the meeting was agreement on the need for future collaborative research projects to address these gaps in the field of lung transplantation.

The role of C4d deposition in the diagnosis of antibody-mediated rejection after lung transplantation.

Antibody-mediated rejection (AMR) is an increasingly recognized form of lung rejection. C4d deposition has been an inconsistent finding in previous reports and its role in the diagnosis has been controversial. We conducted a retrospective single-center study to characterize cases of C4d-negative probable AMR and to compare these to cases of definite (C4d-positive) AMR. We identified 73 cases of AMR: 28 (38%) were C4d-positive and 45 (62%) were C4d-negative. The two groups had a similar clinical presentation, and although more patients in the C4d-positive group had neutrophilic capillaritis (54% vs. 29%, P = .035), there was no significant difference in the presence of other histologic findings. Despite aggressive antibody-depleting therapy, 19 of 73 (26%) patients in the overall cohort died within 30 days, but there was no significant difference in freedom from chronic lung allograft dysfunction (CLAD) or survival between the two groups. We conclude that AMR may cause allograft failure, but that the diagnosis requires a multidisciplinary approach and a high index of suspicion. C4d deposition does not appear to be a necessary criterion for the diagnosis, and although some cases may respond initially to therapy, there is a high incidence of CLAD and poor survival after AMR.

Donor-Derived Exosomes With Lung Self-Antigens in Human Lung Allograft Rejection.

The immunological role of exosomes in allograft rejection remains unknown. We sought to determine whether exosomes are induced during lung allograft rejection and to define the antigenic compositions of HLA, lung-associated self-antigens (SAgs) and microRNAs (miRNAs). Exosomes were isolated from sera and bronchoalveolar lavage fluid from 30 lung transplant recipients (LTxRs) who were stable or who had acute rejection (AR) or bronchiolitis obliterans syndrome (BOS). Exosomes were defined by flow cytometry for CD63 and western blotting for annexin V SAgs, collagen V (Col-V) and Kα1 tubulin were examined by electron microscopy; miRNAs were profiled by a miRNA array. Donor HLA and SAgs were detected on exosomes from LTxRs with AR and BOS but not from stable LTxRs. Exosomes expressing Col-V were isolated from sera from LTxRs 3 mo before AR and 6 mo before BOS diagnosis, suggesting that exosomes with SAgs may be a noninvasive rejection biomarker. Exosomes isolated from LTxRs with AR or BOS also contained immunoregulatory miRNAs. We concluded that exosomes expressing donor HLA, SAgs and immunoregulatory miRNAs are present in the circulation and local site after human lung transplantation and play an important role in the immune pathogenesis of acute allograft rejection and BOS.

Long-Term Persistence of Donor Alveolar Macrophages in Human Lung Transplant Recipients That Influences Donor-Specific Immune Responses.

Steady-state alveolar macrophages (AMs) are long-lived lung-resident macrophages with sentinel function. Evidence suggests that AM precursors originate during embryogenesis and populate lungs without replenishment by circulating leukocytes. However, their presence and persistence are unclear following human lung transplantation (LTx). Our goal was to examine donor AM longevity and evaluate whether AMs of recipient origin seed the transplanted lungs. Origin of AMs was accessed using donor-recipient HLA mismatches. We demonstrate that 94-100% of AMs present in bronchoalveolar lavage (BAL) were donor derived and, importantly, AMs of recipient origin were not detected. Further, analysis of BAL cells up to 3.5 years post-LTx revealed that the majority of AMs (>87%) was donor derived. Elicitation of de novo donor-specific antibody (DSA) is a major post-LTx complication and a risk factor for development of chronic rejection. The donor AMs responded to anti-HLA framework antibody (Ab) with secretion of inflammatory cytokines. Further, in an experimental murine model, we demonstrate that adoptive transfer of allogeneic AMs stimulated humoral and cellular immune responses to alloantigen and lung-associated self-antigens and led to bronchiolar obstruction. Therefore, donor-derived AMs play an essential role in the DSA-induced inflammatory cascade leading to obliterative airway disease of the transplanted lungs.

B Cell-Activating Transcription Factor Plays a Critical Role in the Pathogenesis of Anti-Major Histocompatibility Complex-Induced Obliterative Airway Disease.

Antibodies (Abs) against major histocompatibility complex (MHC) results in T helper-17 (Th17)-mediated immunity against lung self-antigens (SAgs), K-α1 tubulin and collagen V and obliterative airway disease (OAD). Because B cell-activating transcription factor (BATF) controls Th17 and autoimmunity, we proposed that BATF may play a critical role in OAD. Anti-H2K(b) was administered intrabronchially into Batf (-/-) and C57BL/6 mice. Histopathology of the lungs on days 30 and 45 after Ab administration to Batf (-/-) mice resulted in decreased cellular infiltration, epithelial metaplasia, fibrosis, and obstruction. There was lack of Abs to SAgs, reduction of Sag-specific interleukin (IL)-17 T cells, IL-6, IL-23, IL-17, IL-1ß, fibroblast growth factor-6, and CXCL12 and decreased Janus kinase 2, signal transducer and activator of transcription 3 (STAT3), and retinoid-related orphan receptor γT. Further, micro-RNA (miR)-301a, a regulator of Th17, was reduced in Batf (-/-) mice in contrast to upregulation of miR-301a and downregulation of protein inhibitor of activated STAT3 (PIAS3) in anti-MHC-induced OAD animals. We also demonstrate an increase in miR-301a in the bronchoalveolar lavage cells from lung transplant recipients with Abs to human leukocyte antigen. This was accompanied by reduction in PIAS3 mRNA. Therefore, we conclude that BATF plays a critical role in the immune responses to SAgs and pathogenesis of anti-MHC-induced rejection. Targeting BATF should be considered for preventing chronic rejection after human lung transplantation.

Dysregulated MicroRNA Expression and Chronic Lung Allograft Rejection in Recipients With Antibodies to Donor HLA.

The pathogenesis of chronic rejection, Bronchiolitis Obliterans Syndrome (BOS) following lung transplantation (LT) is poorly understood. We hypothesized that development of antibodies to HLA (DSA) is associated with dysregulation of microRNA (miRNA) that predisposes BOS. Towards this, miRNA profiling of mononuclear cells from 10 stable LT (DSA(-) BOS(-) ), 10 LT with DSA(+) BOS(-) (DSA group) and 10 LT with DSA(+) BOS(+) (BOS group) were performed. Prediction by mirPath indicated that differential miRNAs in DSA(+) BOS(-) compared to stable are significantly up-regulated (relative fold >2, p < 0.05) for TGF-ß and B cell receptor signal pathways. A total of seventy-four miRNAs were up-regulated and six miRNAs were down regulated in LT with DSA(+) BOS(+) when compared to stable (relative fold >2, p < 0.05). There was also significant enrichment of cell cycle and gap junction pathways. An inverse correlation between expression of two key miRNAs and their target genes were observed: miR-369-5p and miR-548d were down regulated in DSA(+) LT while their gene targets in TGF-ß signal pathways were up-regulated. In addition, miR-628-5p and miR-134 were down regulated and their target genes (B cell development) were up-regulated. Therefore, we conclude that alloimmunity induced changes in miRNAs affecting the TGF-ß and B cell receptor signal pathways play important roles in BOS development.

Esophageal motor disease and reflux patterns in patients with advanced pulmonary disease undergoing lung transplant evaluation.

BACKGROUND: Advanced pulmonary disorders are linked to esophageal hypomotility and reflux disease. However, characterization of esophageal function using high resolution manometry (HRM) and ambulatory pH monitoring, segregation by pulmonary pathology, and comparison to traditional reflux disease are all limited in the literature. METHODS: Over a 4 year period, 73 patients (55.2 ± 1.3 years, 44F) were identified who underwent esophageal function testing as part of lung transplant evaluation for advanced pulmonary disease (interstitial lung disease, ILD = 47, obstructive lung disease, OLD = 24, other = 2). Proportions of patients with motor dysfunction (≥ 80% failed sequences = severe hypomotility) and/or abnormal reflux parameters (acid exposure time, AET ≥ 4%) were determined, and compared to a cohort of 1081 patients (48.4 ± 0.4 years, 613F) referred for esophageal function testing prior to antireflux surgery (ARS). KEY RESULTS: The proportion of esophageal body hypomotility was significantly higher within advanced pulmonary disease categories (35.6%), particularly ILD (44.7%), compared to ARS patients (12.1%, P < 0.0001). Abnormal AET was noted in 56.5%, and was similar between ILD and OLD, but less frequent than in the ARS group (P = 0.04). Post-transplant chronic rejection trended towards association with pretransplant elevated AET in OLD (P = 0.08) but not ILD. Mortality was not predicted by esophageal motor pattern or reflux evidence. CONCLUSIONS & INFERENCES: Interstitial lung disease has a highly significant association with esophageal body hypomotility. Consequently, prevalence of abnormal esophageal acid exposure is high, but implications for post lung transplant chronic rejection remain unclear.

Granulocyte transfusions in hematologic malignancy patients with invasive pulmonary aspergillosis: outcomes and complications.

BACKGROUND: Granulocyte transfusions (GTXs) have been used successfully as an adjunctive treatment option for invasive infections in some neutropenic patients with underlying hematologic malignancy (HM). PATIENTS AND METHODS: We sought to determine the impact of GTX as an adjunct to antifungal therapy in 128 patients with HM and prolonged neutropenia (≥14 days) with a proven or probable invasive aspergillosis (IA) infection by retrospectively reviewing our institutional database. RESULTS: Fifty-three patients received GTX and 75 did not. By univariate analysis, patients with invasive pulmonary aspergillosis who received GTX were less likely to respond to antifungal therapy (P = 0.03), and more likely to die of IA (P = 0.009) when compared with the non-GTX group. Among patients who received GTX, 53% developed a pulmonary reaction. Furthermore, IA-related death was associated with the number of GTX given (P = 0.018) and the early initiation of GTX within 7 days after starting antifungal therapy (P = 0.001). By multivariate competing risk analysis, patients who received GTX were more likely to die of IA than patients who did not receive GTX (P = 0.011). CONCLUSIONS: Our study suggests that GTX does not improve response to antifungal therapy and is associated with worse outcomes of IA infection in HM patients, particularly those with pulmonary involvement.

Precision diagnostics in transplantation: from bench to bedside.

The Canadian and American Societies of Transplantation held a symposium on February 22, 2012 in Quebec City focused on discovery, validation and translation of new diagnostic tools into clinical transplantation. The symposium focused on antibody testing, transplantation pathology, molecular diagnostics and laboratory support for the incompatible patient. There is an unmet need for more precise diagnostic approaches in transplantation. Significant potential for increasing the diagnostic precision in transplantation was recognized through the integration of conventional histopathology, molecular technologies and sensitive antibody testing into one enhanced diagnostic system.

Chelator-based catheter lock solutions in eradicating organisms in biofilm.

Two different chelator-based antimicrobial catheter lock solutions, methylene blue-citrate-parabens (MB-CIT) and minocycline-EDTA-25% ethanol (M-EDTA-25% ETOH), were compared in 2-h biofilm eradication experiments. Eradication of both mature and immature Gram-positive, Gram-negative, and fungal biofilms was assessed. M-EDTA-25% ETOH was able to fully eradicate all biofilms within 2 h. MB-CIT was only effective against immature biofilms but was unable to fully eradicate most of the mature biofilms tested.

Antibodies to K-α 1 tubulin and collagen V are associated with chronic rejection after lung transplantation.

Bronchiolitis obliterans syndrome (BOS), the clinical correlate of chronic rejection after lung transplantation, is the leading obstacle to better long-term outcomes. We previously instituted a clinical protocol to screen for donor-specific human leukocyte antigen (HLA) antibodies (DSA) and a preemptive antibody-directed therapy protocol consisting of rituximab and/or intravenous immune globulin. In this study, we retrospectively analyzed serum samples from lung transplant recipients (n = 108) for antibodies to self-antigens (K-α 1 tubulin and collagen V) before and after antibody-directed therapy and correlated the results with the subsequent development of BOS. Seventy-two of the 108 recipients developed antibodies to self-antigens. There was a correlation between the development of antibodies to self-antigens and DSA. Sixteen of the 54 patients who had antibodies to self-antigens and were treated with antibody-directed therapy cleared the antibodies, and they were significantly less likely to develop BOS than those who had persistent antibodies. Furthermore, those who cleared DSA after treatment but had persistent antibodies to self-antigens were significantly more likely to develop BOS than those who cleared these antibodies. We conclude that antibodies to self-antigens are an important risk factor for the development of BOS.

A shift in the collagen V antigenic epitope leads to T helper phenotype switch and immune response to self-antigen leading to chronic lung allograft rejection.

Immune responses to human leucocyte antigen (HLA) and self-antigen collagen V (Col-V) have been proposed in the pathogenesis of chronic rejection (bronchiolitis obliterans syndrome, BOS) following human lung transplantation (LTx). In this study, we defined the role for the shift in immunodominant epitopes of Col-V in inducing T helper phenotype switch leading to immunity to Col-V and BOS. Sera and lavage from BOS(+) LTx recipients with antibodies to Col-V were analysed. Two years prior to BOS, patients developed antibodies to both Col-V,α1(V) and α2(V) chains. However, at clinical diagnosis of BOS, antibodies became restricted to α1(V). Further, lung biopsy from BOS(+) patients bound to antibodies to α1(V), indicating that these epitopes are exposed. Fourteen Col-V peptides [pep1-14, pep1-4 specific to α1(V), pep5-8 to α1,2(V) and pep9-14 to α2(V)] which bind to HLA-DR4 and -DR7, demonstrated that prior to BOS, pep 6, 7, 9, 11 and 14 were immunodominant and induced interleukin (IL)-10. However, at BOS, the response switched to pep1, 4 and 5 and induced interferon (IFN)-γ and IL-17 responses, but not IL-10. The T helper (Th) phenotype switch is accompanied by decreased frequency of regulatory T cells (T(regs) ) in the lavage. LTx recipients with antibodies to α1(V) also demonstrated increased matrix metalloproteinase (MMP) activation with decreased MMP inhibitor, tissue inhibitor of metalloproteinase (TIMP), suggesting that MMP activation may play a role in the exposure of new Col-V antigenic epitopes. We conclude that a shift in immunodominance of self-antigenic determinants of Col-V results in induction of IFN-γ and IL-17 with loss of tolerance leading to autoimmunity to Col-V, which leads to chronic lung allograft rejection.

Predictors of catheter-related gram-negative bacilli bacteraemia among cancer patients.

Gram-negative bacillary bacteraemia (GNB) is associated with high morbidity and mortality among cancer patients. We conducted this study to determine the risk factors that may predict the catheter as the source of GNB in cancer patients. From July 2005 to December 2006 all 266 cancer patients with GNB and central venous catheters (CVCs) at The University of Texas M. D. Anderson Cancer Centre in Houston, were classified as catheter-related bloodstream infection (CRBSI) according to Infectious Diseases Society of America criteria. We compared clinical and microbiological features of CRBSIs and non-CRBSIs. We identified 78 CRBSIs and 126 non-CRBSIs. On univariate analysis, polymicrobial bacteraemia, Stenotrophomonas maltophilia bacteraemia, and more than 1000 CFUs in CVC blood cultures, were more common among CRBSI cases. Escherichia coli bacteraemia, haematologic cancer, neutropenia and prior antibiotic use were more common among non-CRBSI cases. On multivariate analysis, S. maltophilia bacteraemia (odds ratio (OR), 5.78; 95% confidence interval (CI), 1.47-22.78; p 0.045), polymicrobial bacteraemia (OR, 4.04; 95% CI, 1.56-10.44; p 0.042), and more than 1000 CFUs from CVC blood cultures (OR, 4.39; 95% CI, 2.02-9.27; p <0.01), were associated with CRBSI. Neutropenia was associated with non-CRBSI (OR, 0.26; 95% CI, 0.13-0.53; p <0.01). Several factors such as S. maltophilia bacteraemia, polymicrobial bacteraemia and more than 1000 CFUs from a blood culture drawn through the CVC may assist the clinicians in assessing whether an indwelling catheter is the source of a GNB and hence CVC removal may be considered.

Comparing clinical and microbiological methods for the diagnosis of true bacteraemia among patients with multiple blood cultures positive for coagulase-negative staphylococci.

We assessed the accuracy of the Centers for Disease Control and Prevention (CDC) clinical criteria as well as other microbiological methods for the diagnosis of coagulase-negative staphylococci bacteraemia. The CDC clinical criteria had low accuracy, which can be improved by speciation, particularly if the patient had more than two positive blood cultures.

Efficacy of caspofungin as salvage therapy for invasive aspergillosis compared to standard therapy in a historical cohort.

In a non-comparative study, caspofungin was effective salvage therapy for approximately half of the patients refractory to or intolerant of standard antifungal agents for invasive aspergillosis. To establish a frame of reference for these results, we compared the response to caspofungin with responses to other antifungal agents in a historical cohort of similar patients. The efficacy could be evaluated in 83 patients who received caspofungin 50 mg daily after a 70-mg loading dose. The historical control group, identified through a retrospective review of medical records, included 214 evaluable patients possibly refractory to or intolerant of ≥1 week of standard antifungal therapy. All patients had documented invasive aspergillosis. Favorable response was defined as a complete or partial response to therapy. Underlying diseases, baseline neutropenia, corticosteroid use, and sites of infection were similar in both studies. Most patients had received amphotericin B formulations and/or itraconazole, and were refractory to standard therapy. Favorable response rates were 45% with caspofungin and 16% with standard therapy. The unadjusted odds ratio for a favorable response (caspofungin/standard therapy) was 4.1 (95% confidence interval: 2.2, 7.5). After adjusting for potential imbalances in the frequency of disseminated infection, neutropenia, steroid use, and bone marrow transplantation between groups, the odds ratio remained at 4.1 (2.1, 7.9). Although only tentative conclusions about relative efficacy can be drawn from retrospective comparisons, caspofungin appeared to be at least as efficacious as an amphotericin B formulation and/or itraconazole for the treatment of invasive aspergillosis in patients refractory to or intolerant of their initial antifungal therapy.