Epstein-Barr Virus Monitoring after an Allogeneic Hematopoietic Stem Cell Transplant: Review of the Recent Data and Current Practices in Canada.

Epstein-Barr virus-related post-transplantation lymphoproliferative disorder (EBV-PTLD) is a serious complication following hematopoietic stem cell transplantation (HSCT). A pre-emptive strategy using rituximab, which aims to manage patients early at the time of EBV reactivation to avoid PTLD, has been recommended by the most recent ECIL-6 guidelines in 2016. However, there is still a great heterogeneity of viral-load monitoring protocols, targeted patient populations, and pre-emptive treatment characteristics between centers, making precise EBV monitoring recommendations difficult. We conducted a literature review from the most recent publications between 1 January 2015 and 1 August 2023, to summarize the emerging data on EBV-PTLD prevention strategies in HSCT recipients, including the EBV-DNA threshold and use of rituximab. We also present the results of a survey of current practices carried out in 12 of the main HSCT centers across Canada. We confirm that pre-emptive rituximab remains an efficient strategy for EBV-PTLD prevention. However, there is an urgent need to perform prospective, randomized, multicentric trials with larger numbers of patients reflecting current practices to determine the best clinical conduct with regards to rituximab dosing, timing of treatment, and criteria to initiate treatments. Longer follow-ups will also be necessary to assess patients' long-term outcomes.

A Rapid and Efficient Method for the Extraction of Histone Proteins.

Current protocols used to extract and purify histones are notoriously tedious, especially when using yeast cells. Here, we describe the use of a simple filter-aided sample preparation approach enabling histone extraction from yeast and mammalian cells using acidified ethanol, which not only improves extraction but also inactivates histone-modifying enzymes. We show that our improved method prevents N-terminal clipping of H3, an artifact frequently observed in yeast cells using standard histone extraction protocols. Our method is scalable and provides efficient recovery of histones when extracts are prepared from as few as two million yeast cells. We further demonstrate the application of this approach for the analysis of histone modifications in fungal clinical isolates available in a limited quantity. Compared with standard protocols, our method enables the study of histones and their modifications in a faster, simpler, and more robust manner.

Real-life comparison of posaconazole versus fluconazole for primary antifungal prophylaxis during remission-induction chemotherapy for acute leukemia.

Background: Patients undergoing remission-induction intensive chemotherapy for acute leukemia are at high risk for life-threatening invasive fungal infections (IFIs). Primary antifungal prophylaxis with posaconazole has been shown to reduce the incidence of IFI compared to fluconazole, but real-life data are limited and the effect on mortality remains unclear. Methods: This retrospective cohort study compared fluconazole and posaconazole as primary prophylaxis in real-life practice over a 10-year period, in a Canadian hospital. Results: A total of 299 episodes were included (fluconazole, n = 98; posaconazole, n = 201), of which 68% were first inductions. The underlying hematologic malignancy was acute myeloid leukemia or myelodysplastic syndrome in 88% of episodes and acute lymphoblastic leukemia in 9%. Overall, 20 cases of IFI occurred (aspergillosis, n = 17; candidiasis, n = 3) and 14 were considered as breakthrough IFI. IFI incidence was significantly lower in the posaconazole group (3.5% versus 13.2%; p = 0.001). Empirical or targeted antifungal therapy was also reduced in the posaconazole cohort. Mortality was similar in both groups. Conclusions: In a real-life setting in Canada, primary posaconazole prophylaxis reduces the incidence of IFI during remission-induction chemotherapy, compared to fluconazole.

Historique: Les patients soumis à une chimiothérapie intensive visant à induire la rémission d'une leucémie aiguë sont très vulnérables à des infections fongiques invasives (IFI) au potentiel mortel. Il est démontré qu'une prophylaxie antifongique primaire au posaconazole réduit l'incidence d'IFI davantage que le fluconazole, mais les données sur le terrain sont limitées et l'effet de ce médicament sur la mortalité demeure nébuleux. Méthodologie: La présente étude de cohorte rétrospective a comparé le fluconazole au posaconazole comme prophylaxie primaire sur une période de dix ans dans un hôpital canadien. Résultats: Au total, 299 épisodes ont été inclus (fluconazole, n = 98; posaconazole, n = 201), dont 68 % étaient des premières occurrences. Dans 88 % des épisodes, la leucémie myéloïde était le cancer hématologique sous-jacent, et dans 9 % des cas, il s'agissait plutôt d'une leucémie aiguë lymphoblastique. Dans l'ensemble, 20 cas d'IFI ont été observés (aspergillose, n = 17; candidose, n = 3) et 14 étaient considérés comme des IFI qui avaient percé malgré une médication. L'incidence d'IFI était beaucoup plus faible dans le groupe prenant du posaconazole (3,5 % par rapport à 13,2 %; p = 0,001). Le traitement antifongique empirique ou ciblé était également limité dans cette cohorte. La mortalité était semblable dans les deux groupes. Conclusions: Sur le terrain au Canada, la prophylaxie primaire au posaconazole réduit l'incidence d'IFI davantage que le fluconazole pendant une chimiothérapie visant à induire une rémission.

Multicenter Evaluation of Attenuated Total Reflectance Fourier Transform Infrared (ATR-FTIR) Spectroscopy-Based Method for Rapid Identification of Clinically Relevant Yeasts.

Fourier transform infrared (FTIR) spectroscopy has demonstrated applicability as a reagent-free whole-organism fingerprinting technique for both microbial identification and strain typing. For routine application of this technique in microbiology laboratories, acquisition of FTIR spectra in the attenuated total reflectance (ATR) mode simplifies the FTIR spectroscopy workflow, providing results within minutes after initial culture without prior sample preparation. In our previous central work, 99.7% correct species identification of clinically relevant yeasts was achieved by employing an ATR-FTIR-based method and spectral database developed by our group. In this study, ATR-FTIR spectrometers were placed in 6 clinical microbiology laboratories over a 16-month period and were used to collect spectra of routine yeast isolates for on-site identification to the species level. The identification results were compared to those obtained from conventional biochemical tests and/or matrix-assisted laser desorption/ionization-time of flight mass spectrometry. Isolates producing discordant results were reanalyzed by routine identification methods, ATR-FTIR spectroscopy, and PCR gene sequencing of the D1/D2 and internal transcribed spacer (ITS) regions. Among the 573 routine clinical yeast isolates collected and identified by the ATR-FTIR-based method, 564 isolates (98.4%) were correctly identified at the species level, while the remaining isolates were inconclusive with no misidentifications. Due to the low prevalence of Candida auris in routine isolates, additional randomly selected C. auris (n = 24) isolates were obtained for evaluation and resulted in 100% correct identification. Overall, the data obtained in our multicenter evaluation study using multiple spectrometers and system operators indicate that ATR-FTIR spectroscopy is a reliable, cost-effective yeast identification technique that provides accurate and timely (∼3 min/sample) species identification promptly after the initial culture.

SARS-CoV-2 seroprevalence in health care workers from 10 hospitals in Quebec, Canada: a cross-sectional study.

BACKGROUND: The COVID-19 pandemic has disproportionately affected health care workers. We sought to estimate SARS-CoV-2 seroprevalence among hospital health care workers in Quebec, Canada, after the first wave of the pandemic and to explore factors associated with SARS-CoV-2 seropositivity. METHODS: Between July 6 and Sept. 24, 2020, we enrolled health care workers from 10 hospitals, including 8 from a region with a high incidence of COVID-19 (the Montréal area) and 2 from low-incidence regions of Quebec. Eligible health care workers were physicians, nurses, orderlies and cleaning staff working in 4 types of care units (emergency department, intensive care unit, COVID-19 inpatient unit and non-COVID-19 inpatient unit). Participants completed a questionnaire and underwent SARS-CoV-2 serology testing. We identified factors independently associated with higher seroprevalence. RESULTS: Among 2056 enrolled health care workers, 241 (11.7%) had positive SARS-CoV-2 serology. Of these, 171 (71.0%) had been previously diagnosed with COVID-19. Seroprevalence varied among hospitals, from 2.4% to 3.7% in low-incidence regions to 17.9% to 32.0% in hospitals with outbreaks involving 5 or more health care workers. Higher seroprevalence was associated with working in a hospital where outbreaks occurred (adjusted prevalence ratio 4.16, 95% confidence interval [CI] 2.63-6.57), being a nurse or nursing assistant (adjusted prevalence ratio 1.34, 95% CI 1.03-1.74) or an orderly (adjusted prevalence ratio 1.49, 95% CI 1.12-1.97), and Black or Hispanic ethnicity (adjusted prevalence ratio 1.41, 95% CI 1.13-1.76). Lower seroprevalence was associated with working in the intensive care unit (adjusted prevalence ratio 0.47, 95% CI 0.30-0.71) or the emergency department (adjusted prevalence ratio 0.61, 95% CI 0.39-0.98). INTERPRETATION: Health care workers in Quebec hospitals were at high risk of SARS-CoV-2 infection, particularly in outbreak settings. More work is needed to better understand SARS-CoV-2 transmission dynamics in health care settings.

Long-lasting cluster of nosocomial pneumonia with a single Pneumocystis jirovecii genotype involving different organ allograft recipients.

Pneumocystis pneumonia (PCP) outbreaks may occur in solid organ transplant (SOT) patients. Transmissibility of Pneumocystis jirovecii among SOT and non-SOT patients has not been investigated. Ten SOT (ie, 4 heart, 4 kidney, 2 liver allograft recipients) and 11 non-SOT (ie, 7 HIV-infected, 3 hematologic malignancies, and 1 stem cell transplant) patients with PCP were admitted to London Health Sciences Center (LHSC) from October 2014 to August 2016. We investigated the course of illness and outcome of PCP in SOT and non-SOT patients. Post-transplant PCP was frequently an acute-onset disease (90% vs. 18.2%, p = .01) requiring ICU admission (70% vs. 20%, p = .03) and hemodialysis (60% vs. 0, p = .003). Mortality was more frequent in SOT patients (40% vs. 18.1%, p = .36). Multilocus sequence typing (MLST) demonstrated circulation of a single genotype of P. jirovecii among SOT patients. However, 8 different genotypes were detected from non-SOT patients. Reinstitution of prophylaxis successfully controlled post-transplant cluster until end of observation period in October 2019. No transmission was detected from non-SOT patients to SOT recipients. Detection of a single P. jirovecii genotype from all SOT recipients highlights the likelihood of nosocomial transmission. No source control method is recommended by current guidelines. Improvement of preventive strategies is required.

Lymphocytic Vasculitis Associated With Mild Rejection in a Vascularized Composite Allograft Recipient: A Clinicopathological Study.

BACKGROUND: Histologic criteria for diagnosing acute rejection in vascularized composite tissue allograft (VCA) have been established by the Banff 2007 Working Classification of Skin-Containing Composite Tissue Allograft, but the role of early vascular lesions in graft rejection warrants additional analysis. METHODS: We performed a retrospective study of 34 skin biopsies performed over 430 d for rejection surveillance, in Canada's first face allotransplant recipient. Three observers reviewed all biopsies to assess the nature and intensity of the inflammatory skin infiltrate. A complete histological and immunohistochemical review of the vascular components was performed with a focus on lymphocytic vasculitis, intravascular fibrin, vessel caliber, extent of injury, C4d positivity, and inflammatory cell phenotyping. We then correlated these data points to clinical and immunosuppression parameters. RESULTS: Acute vascular damage in biopsies that would be classified as mild acute rejection correlates with troughs in immunosuppression and subsides when immunosuppressive tacrolimus doses are increased. Grade 0 Banff rejection and Grade I without lymphocytic vasculitis were almost indistinguishable, whereas Grade I with lymphocytic vasculitis was an easy and reproducible histologic finding. CONCLUSIONS: Our results highlight the possible relevance of vascular injury in the context of VCA, as its presence might underlie a more aggressive form of immune rejection. If these findings are validated in other VCA patients, vascular injury in mild rejection might warrant a different clinical approach.

Prevalence and mechanisms of azole resistance in clinical isolates of Aspergillus section Fumigati species in a Canadian tertiary care centre, 2000 to 2013.

OBJECTIVES: Azole resistance among Aspergillus fumigatus isolates is a growing concern worldwide. Induction of mutations during azole therapy, environment-acquired mutations caused by azole fungicides and intrinsic resistance of cryptic Fumigati species all contribute to the burden of resistance. However, there is a lack of data in Canada on this emerging threat. METHODS: To gain insights into the magnitude and mechanisms of resistance, a 14 year collection of Aspergillus section Fumigati comprising 999 isolates from 807 patients at a Montreal hospital was screened for azole resistance, and resistance mechanisms were investigated with the combined use of genome sequencing, 3D modelling and phenotypic efflux pump assays. RESULTS: Overall azole resistance was low (4/807 patients; 0.5%). A single azole-resistant A. fumigatus sensu stricto strain, isolated from a patient with pulmonary aspergillosis, displayed efflux-pump-mediated resistance. Three patients were colonized or infected with azole-resistant cryptic Fumigati species (one Aspergillus thermomutatus, one Aspergillus lentulus and one Aspergillus turcosus). Evidence is presented that azole resistance is efflux-pump-mediated in the A. turcosus isolate, but not in the A. lentulus and A. thermomutatus isolates. CONCLUSIONS: Azole resistance is rare in our geographic area and currently driven by cryptic Fumigati species. Continued surveillance of emergence of resistance is warranted.

Identification of Candida auris by Use of the Updated Vitek 2 Yeast Identification System, Version 8.01: a Multilaboratory Evaluation Study.

Candida auris is an emerging multidrug-resistant yeast that has been systematically incorrectly identified by phenotypic methods in clinical microbiology laboratories. The Vitek 2 automated identification system (bioMérieux) recently included C. auris in its database (version 8.01). We evaluated the performance of the Vitek 2 YST ID card to identify C. auris and related species. A panel of 44 isolates of Candida species (C. auris, n = 35; Candida haemulonii, n = 5; Candida duobushaemulonii, n = 4) were tested by three different hospital-based microbiology laboratories. Among 35 isolates of C. auris, Vitek 2 yielded correct identification in an average of 52% of tested samples. Low-discrimination (LD) results with an inability to distinguish between C. auris, C. duobushaemulonii, and Candida famata were obtained in an average of 27% of samples. Incorrect identification results were obtained in an average of 21% of samples, the majority (91%) of which were reported as C. duobushaemulonii and the remaining 9% of which were reported as Candida lusitaniae/C. duobushaemulonii. The proportion of correct identification was not statistically different across different centers (P = 0.78). Stratification by genetic clades demonstrated that 100% (n = 8) of the strains of the South American clade were correctly identified compared to 7% (n = 10) and 0% (n = 4) from the African and East Asian clades, respectively. None of the non-auris Candida strains (n = 9) were incorrectly identified as C. auris Our results show that the Vitek 2 (version 8.01) yeast identification system has a limited ability to correctly identify C. auris These data suggest that an identification result for C. duobushaemulonii should warrant further testing to rule out C. auris The overall performance of the Vitek 2 seems to differ according to C. auris genetic clade, with the South American isolates yielding the most accurate results.

Draft Genome Sequence of Azole-Resistant Aspergillus thermomutatus (Neosartorya pseudofischeri) Strain HMR-AF-39, Isolated from a Human Nasal Septum Abscess Aspirate.

Here, we present the draft genome sequence of Aspergillus thermomutatus (formerly known as Neosartorya pseudofischeri; strain HMR-AF-39/LSPQ-01276), a cryptic species of Aspergillus section Fumigati. This species is intrinsically resistant to antifungal azoles and is recognized as an agent of invasive aspergillosis among immunocompromised hosts.

Draft Genome Sequences of Azole-Resistant and Azole-Susceptible Aspergillus turcosus Clinical Isolates Recovered from Bronchoalveolar Lavage Fluid Samples.

We present the draft genome sequences of two clinical strains of Aspergillus turcosus, one azole-susceptible (strain HMR-AF-23/LSPQ-01275) and the other azole-resistant (strain HMR-AF-1038/LSPQ-01280), isolated from bronchoalveolar lavage fluid of two adult patients. These two strains are the first reported clinical isolates of A. turcosus.

Comparison of early and recent influenza A(H1N1)pdm09 isolates harboring or not the H275Y neuraminidase mutation, in vitro and in animal models.

After 6 years of circulation in humans, a novel antigenic variant of influenza A(H1N1)pdm09 (i.e., A/Michigan/45/2015) emerged in 2015-16 and has predominated thereafter worldwide. Herein, we compared in vitro and in vivo properties of 2016 wild-type (WT) A/Michigan/45/15-like isolate and its H275Y neuraminidase (NA) variant to the original A/California/07/09-like counterparts. The H275Y mutation induced comparable levels of resistance to oseltamivir and peramivir without altering zanamivir susceptibility in both 2009 and 2016 isolates. In vitro, the two WT isolates had comparable replicative properties. The 2016-H275Y isolate had lower titers at 36 h post-inoculation (PI) (P < 0.05) while the 2009-H275Y titers were lower at both 24 h (P < 0.01) and 36 h PI (P < 0.001) vs the respective WTs. In mice, the 2016-WT isolate caused less weight losses (P < 0.001) and lower lung viral titers (LVTs) (P < 0.01) vs the 2009-WT. The LVTs of 2016-WT and 2016-H275Y groups were comparable whereas the 2009-H275Y LVTs were lower vs the respective WT (P < 0.01). Ferrets infected with the 2016-WT isolate and their contacts had higher nasal viral titers (NVTs) at early time points vs the 2009-WT group (P < 0.01). Also, NVTs of 2016-H275Y animals were lower vs the 2016-WT group at early time points in both infected (P < 0.01) and contact animals (P < 0.001). In conclusion, while the H275Y mutation similarly impacts the A/California/07/2009- and A/Michigan/45/2015-like A(H1N1)pdm09 NAs, the fitness of these isolates differs according to animal models with the 2016 virus being less virulent in mice but slightly more virulent in ferrets, potentially reflecting a period of cumulative changes in surface and internal genes.

High frequency of pathogenic Aspergillus species among nonsporulating moulds from respiratory tract samples.

Nonsporulating moulds (NSM) represent an identification challenge for clinical laboratories. Data on the prevalence of pathogenic species among NSM are lacking. We prospectively investigated consecutive thermotolerant (36°C) clinical NSM isolates from respiratory tract samples. A total of 123 isolates were identified by DNA sequencing and phenotypically characterized. Of those, 13 (11%) were pathogenic species (Aspergillus fumigatus, n = 10; A. flavus, n = 1; A. hiratsukae, n = 1; Schizophyllum commune, n = 1). Presumptive identification of Aspergillus species among NSM can be achieved by simple phenotypic testing.

Successful Treatment of Disseminated Anncaliia algerae Microsporidial Infection With Combination Fumagillin and Albendazole.

Anncaliia algerae myositis is a life-threatening, emerging microsporidiosis among immunocompromised hosts. We report a case of disseminated A algerae infection in a man previously treated with alemtuzumab. Due to failure of albendazole-based therapy, fumagillin was added as a novel approach to management, with a good clinical response and patient survival.

Incidence of invasive aspergillosis following remission-induction chemotherapy for acute leukemia: a retrospective cohort study in a single Canadian tertiary care centre.

BACKGROUND: The decision to use universal primary antimould prophylaxis to prevent invasive aspergillosis in patients with acute leukemia depends on the incidence of infection at individual centres. We determined our institution's incidence of invasive aspergillosis among patients who received remission-induction chemotherapy for acute leukemia to evaluate the potential benefits of primary antimould prophylaxis. METHODS: We conducted this retrospective cohort study at a Canadian tertiary care centre. From the central pharmacy registries, we retrieved records for all adult patients for whom remission-induction chemotherapy for acute leukemia was prescribed between 2008 and 2010. We retrieved clinical, microbiologic, pathologic and radiologic data from the patients' medical charts. The primary outcome was a diagnosis of probable or proven invasive aspergillosis up to 180 days after resolution of aplasia. RESULTS: We retrieved records for 123 patients with acute leukemia. Twenty-two of these patients did not receive the prescribed chemotherapy and were excluded from the analysis. Of the 101 patients included, 77 (76.2%) had acute myeloid leukemia. Overall, 136 courses of chemotherapy were administered, with more than 1 course administered to 26 (25.7%) of the 101 patients. In 9 of the patients (8.9%; 95% confidence interval 4.2%-16.2%), invasive aspergillosis was diagnosed (3 proven and 6 probable cases) a median of 19 (range 11-34) days after initiation of chemotherapy. In 7 (78%) of these 9 patients, invasive aspergillosis occurred during the first course of chemotherapy. Three patients died within the first year after diagnosis of invasive aspergillosis. INTERPRETATION: We found a high incidence (8.9%) of invasive aspergillosis at our centre. This finding triggered the introduction of targeted antimould prophylaxis for patients with acute leukemia who were undergoing remission-induction chemotherapy.

Epidemiology of Candida kefyr in patients with hematologic malignancies.

Candida kefyr is an emerging pathogen among patients with hematologic malignancies (HM). We performed a retrospective study at Johns Hopkins Hospital to evaluate the epidemiology of C. kefyr colonization and infection in HM patients between 2004 and 2010. Eighty-three patients were colonized and/or infected with C. kefyr, with 8 (9.6%) having invasive candidiasis (IC). The yearly incidence of C. kefyr colonization and candidemia increased over the study period (P < 0.01), particularly after 2009. In 2010, C. kefyr caused 16.7% of candidemia episodes. The monthly incidence of C. kefyr was higher during the summer throughout the study. In a cohort of patients with acute myelogenic leukemia receiving induction chemotherapy, risks for C. kefyr colonization included the summer season (odds ratio [OR], 3.1; P = 0.03); administration of an azole (OR, 0.06; P < 0.001) or amphotericin B (OR, 0.35; P = 0.05) was protective. Fingerprinting of 16 isolates by repetitive sequence-based PCR showed that all were different genotypes. The epidemiology of C. kefyr candidemia was evaluated in another hospital in Montreal, Canada; data confirmed higher rates of C. kefyr infection in the summer. C. kefyr appears to be increasing in HM patients, with prominent summer seasonality. These findings raise questions about the effect of antifungal agents and health care exposures (e.g., yogurt) on the epidemiology of this yeast.

Clostridium difficile infection after adult autologous stem cell transplantation: a multicenter study of epidemiology and risk factors.

We sought to describe the epidemiology of Clostridium difficile infection (CDI) among adult recipients of autologous hematopoietic stem cell transplantation (auto-HSCT) within the first year after HSCT in centers with variable epidemiology of hypertoxigenic strains. A multicenter, retrospective nested case-control study was conducted among 873 auto-HSCT recipients at Johns Hopkins Hospital (JHH) and Hôpital Maisonneuve-Rosemont (HMR) between January 2003 and December 2008. Despite center differences in the prevalence of NAP-1 strains during the study period (21% to 43% at JHH versus 80% to 84% in HMR), the 1-year incidence of CDI was similar in the 2 hospitals (6.2% at JHH versus 5.7% at HMR). The median time to infection was 11 days (interquartile range, 1 to 27 days). In case-control analyses, grade ≥2 mucositis (odds ratio [OR], 3.00; P = .02) and receipt of a fourth-generation cephalosporin (OR, 2.76; P = .04) were identified as predictors for CDI. Mucositis was the strongest predictor of risk for CDI in multivariate analysis (adjusted OR, 2.77; P = .03). CDI is a common and early complication of auto-HSCT. Treatment-related gastrointestinal mucosal damage, along with the potentially modifiable risk of antimicrobial exposure, influence the risk for CDI early after auto-HSCT.