Because MICs were not available at the time of A. calidoustus isolation, changes in antifungals were up to the treating physician. Therefore, three of eight patients were initiated on combination antifungal therapy. Others were started on voriconazole (n=2), increased dosing of voriconazole (n=1), or kept on the same dose (n=1). In two patients, immunosuppression was reduced. We analyzed patient outcomes at 90 days after infection. Of the six patients with radiologic changes at the onset of infection, four remained stable, one progressed, and one death occurred within 90 days.
Table 3 summarizes the in vitro antifungal susceptibility results. Voriconazole MICs ranged from 2 to 8 µg/mL. Greater variability was observed with caspofungin minimum effective concentrations (range, 0.03–2 µg/mL). Voriconazole MICs were greater in patients with previous exposure to voriconazole (median, 8 vs. 3 µg/mL in those without previous exposure).
Incidence and Risk Factors in Lung Transplant Recipients
From 2006 to 2007, a total of 74 lung transplant recipients were followed up for a total of 103,821 days, and from 2008 to 2011, a total of 197 patients were followed up for a total of 193,588 days. Incidence rates in lung transplants showed an increase for A. calidoustus (0/1000 vs. 11.3/1000 transplant-years in 2006–2007 and 2008–2011, respectively; P=0.018), whereas A. fumigatus cases decreased (73.9/1000 vs. 49.0/1000 transplant-years, P=0.0066), reflecting a reduction of 53.5%.
In the case-control study of lung transplant recipients, each of the six cases had two controls who were matched for age, gender, baseline lung disease, type of lung transplant (single/double), and follow-up time. When analyzed for other variables that could affect infection, most factors were similar between the two groups, including induction therapy or immunosuppressive regimens, number of respiratory samples taken, length of intensive care unit stay, and rejection rates (Table 4). However, 4 (33.3%) of 12 control patients had received third-generation azole prophylaxis compared with five (83.3%) of six cases (P=0.13). The median duration of exposure to third-generation azoles was greater in cases compared with control patients (3 vs. 0 months, P=0.045).
Table 5 shows the review of literature on solid organ transplant recipients and HSCT patients with A. ustus–associated IA. A. calidoustus cases have not been described, but in many cases, details of species identification are not described. Therefore, many of the A. ustus cases could have been caused by A. calidoustus. For example, the initial description of A. calidoustus by Varga et al. (16) included the isolates described by Panackal et al. (12), which were eventually noted to be all A. calidoustus. Most cases have been reported in HSCT patients (n=15) with only 6 (28.6%) of 21 previous cases reported in organ transplant recipients. There were 9 cases of cutaneous disease, 17 cases of pulmonary disease, and 6 cases of disseminated disease mainly with cerebral infection. Overall mortality was high with death reported in 13 (65%) of 20 cases with 100% mortality in the 4 cases of cerebral disease. Excluding those with cerebral disease, the eight HSCT patients with fatal outcome received either amphotericin B formulation alone (5/8) or the combination of amphotericin B and caspofungin (3/8).
We performed a 5-year review of A. calidoustus infection in transplant recipients at our institution. We found eight patients who were predominantly lung transplant recipients, half of whom presented with underlying chronic lung disease and had received either voriconazole or posaconazole before the isolation of the fungus. In our series, A. calidoustus was always isolated from the respiratory tract, in many cases along with copathogens. Four of six lung transplant patients fulfilled ISHLT criteria for probable IA (17). Approximately half had recent augmentation of immunosuppression, either induction therapy with T-cell depleting agents or therapy for acute rejection. In the case-control study, the duration of azole exposure was associated with A. calidoustus infection. In clinical practice, it would be important to identify Aspergillus isolates to the species level. A. calidoustus can be differentiated based on colony and cellular morphology and temperature requirements for growth. Optimal antifungal therapy is not well defined, although isolates tend to be resistant to azoles but sensitive to polyenes and echinocandins.
Our study is the largest series published to date of transplant recipients specifically with the newly described A. calidoustus species. The Transplant-Associated Infection Surveillance Network study showed an 8.6% cumulative 1-year incidence of IA in lung transplant recipients (2). In addition, there are increasing reports of emerging fungal infections after SOT especially in lung transplant recipients. One factor may be the universal use of antifungal prophylaxis. Starting in 2008, our lung transplant program instituted universal voriconazole prophylaxis beginning on the day of transplantation to 3 months after transplantation in all new lung transplants. Before this, voriconazole was only used for preemptive therapy in those with Aspergillus specie colonization or therapy for IA. We did not find A. calidoustus isolates in transplant recipients before 2008. We also document a significant 53% reduction of A. fumigatus cases after the start of prophylaxis.
Observational studies and our own epidemiological analysis indicate that voriconazole prophylaxis may lead to significant reduction of IA associated with A. fumigatus (5–7); however, in our patients, breakthrough infection with A. calidoustus raises concerns about selecting for strains with decreased antifungal susceptibility. A. calidoustus is reported to have higher MICs to azoles (16, 26, 27). In our center, we observed a significant increase of A. calidoustus cases since the introduction of third-generation azole prophylaxis; six of our eight patients had previous third-generation azole exposure. Voriconazole MICs in patients who previously received voriconazole (patients 1, 3, 4, 5, 6, and 8) were elevated compared with those who had not been exposed (patients 2 and 7). In addition, the patient receiving posaconazole (patient 3) showed an elevated MIC to this antifungal.
In addition, some patients had recently had augmentation of immunosuppression. In our case-control study, the cumulative exposure time to azole prophylaxis was the only significant different risk factor (3 vs. 0 months, P=0.045).
Because A. calidoustus is an emerging species, there are no specific case series for this pathogen in transplant patients. Clearly, these cases occur since isolates of A. calidoustus from transplant patients are often included in larger series of Aspergillus susceptibility testing and molecular identification, but these have not been described in detail (16, 26, 27). It is also likely that these infections were previously classified as A. ustus. Therefore, we reviewed data for both pathogens in the setting of transplantation. We noted that this pathogen is more commonly reported in HSCT patients. High mortality was noted in the reports of HSCT recipients, although this may in part be caused by underlying disease. In our series, we only found pulmonary infections, and no disseminated disease was seen. Similar to our cases, reported patients were generally treated with a variety of antifungal combinations. In the reported literature, survival was variable and not related to any particular therapy. However, the organ transplant recipients who survived were mainly on a terbinafine-based combination. Surviving HSCT patients received a combination of voriconazole and caspofungin. However, surgical resection was also used in many cases as an adjunct therapy and may improve outcomes.
Limitations of our study include the retrospective design. However, our case series is the largest to date in the organ transplant population. Although most of our cases are consistent with probable IA, fungal colonization cannot be excluded. Nevertheless, all our patients were immunosuppressed and showed pulmonary nodules or cavities in several cases suggestive of a fungal cause. Finally, an outbreak of A. calidoustus cannot be excluded given that this organism has previously been described in nosocomial water systems (28). However, all the epidemiological factors including time after transplantation, types of transplant, and the location in which patients underwent bronchoscopy varied, making a single source unlikely.
Overall, our case series suggests the emergence of invasive pulmonary A. calidoustus infection in immunosuppressed transplant recipients during azole use. Because prophylaxis is adopted more widely, unusual drug-resistant fungi are more likely to be seen.
MATERIALS AND METHODS
A retrospective review of the clinical microbiology laboratory database was conducted to identify cases of culture positive for A. ustus or A. calidoustus at our institution between January 2006 and January 2011. From this search, we selected to review in detail the clinical information of solid organ transplant or HSCT recipients and determine susceptibility profiles for isolates from these patients.
All transplant patient charts were reviewed for demographics, type of organ transplant (heart, lung, liver, kidney, or combined) or type of stem-cell transplant (allogeneic or autologous), time after transplantation, immunosuppressive regimen, neutropenia, coinfections, cytomegalovirus viremia in the 30 days before diagnosis, acute rejection in the 30 days before diagnosis, imaging studies, pulmonary function testing, antifungal prophylaxis and treatment, and outcome. Descriptive statistics were used for data analysis.
Fungal infections were defined as colonization, probable infection, or proven infection based on the criteria defined by the ISHLT for thoracic transplants only (17). Infections were also categorized using the European Organization for Research and Treatment of Cancer/Mycoses Study Group criteria for all patients (29).
Literature review of A. ustus and the newly described species A. calidoustus infection in solid organ transplant and HSCT recipients was conducted using PubMed and search terms Aspergillus calidoustus, Aspergillus ustus, and transplantation.
The study was approved by the institutional research ethics board.
Identification of A. calidoustus and Determination of MIC
A. calidoustus was identified based on colony and cellular morphology, including columnar conidial heads and echinulate conidia, and by growth (≥30 mm) on Czapek agar after 7 days incubation at 37°C (16). Antifungal susceptibility testing was performed by broth microdilution according to the Clinical and Laboratory Standards Institute M38-A2 standards document (30). These were reported as MICs for azoles and minimum effective concentration for caspofungin.
Susceptibility results were done as part of this study and were not available to the treating physicians at the time of diagnosis.
A case-control study was performed for only the lung transplant recipients. Lung transplant recipients with A. calidoustus infection (n=6 cases) were matched to a historical cohort of lung transplant recipients without infection and underwent transplantation between January 2006 and December 2007 (2:1 ratio, n=12 controls). Controls and cases were matched for age, gender, baseline disease leading to lung transplantation, and single or double lung transplants. Controls were followed up to the time point that A. calidoustus infection was diagnosed in their respective matched cases.
Incidence rates for A. calidoustus and A. fumigatus in the lung transplant population between the periods 2006 to 2007 and 2008 to 2011 were calculated. This was performed using the date of transplantation, date of death, and date of the last follow-up. SPSS version 19.0 (SPSS Inc., Chicago, IL) was used to perform statistical analysis.
The authors thank Ms. Sandy Shokoples for her technical assistance in susceptibility testing. The authors also thank Ms. Kathy Jackson from the lung transplant program, Mr. Chris Broscheit from pharmacy, and Ms. Leticia Wilson for their assistance in data collection.
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Keywords:© 2012 Lippincott Williams & Wilkins, Inc.
Immunosuppression; Solid organ transplantation; Hematopoietic stem-cell transplant; Antifungal therapy