Introduction

Filamentous fungal infections are increasing in hematology patients,¹ due in largely to the use of more aggressive chemotherapy regimens, with longer periods of severe neutropenia, and the utilization of different modalities of transplant. Unrelated and mismatched donors, and reduced intensity conditioning regimens, promote more graft versus leukemia, and more GVHD with increased risk of filamentous fungal infections, particularly aspergillosis.

Fungal infections are the most significant complication in the management of acute leukemia patients, not only because of high mortality rates² but also because they often mandate the use of lower doses of antineoplastic drugs and delay chemotherapy completion and transplant procedures. Moreover, a proven or probable diagnosis of mycosis is frequently difficult owing to lack of positive cultures and reduced sensitivity of serologic tests, such as galattomannan antigen, in patients receiving prophylactic adsorbable antifungal drugs or those being treated empirically.³ In allogeneic bone marrow transplant recipients, particularly, mortality rates owing to aspergillosis are very high, approaching 90%;^{2, 4} for this reason, secondary prophylaxis with antifungal drugs is usual.⁵ In particular, in patients scheduled for bone marrow transplantation, surgical procedures should be considered when there is persistence of only one pulmonary nodule or when there is cavitation. Such lesions can persist and be colonized by other bacteria or by the same mycotic agent, despite prolonged antifungal treatment. A possible complication of cavitation is hemoptysis, which can occur when the cavity is near a large vessel; historically, this complication was the cause of death in half of the affected patients;⁶ more recently, in acute leukemia patients with aspergillosis, mortality owing to hemoptysis accounted for 20% of deaths.⁷ During recovery from post-chemotherapy aplasia, leukocytes increase the process of necrosis and the risk of hemorrhage by arterial erosion.^{8, 9} Finally, the presence of pulmonary lesions at the time of transplant seems to favor fungal relapse,^{5, 10} not only during the neutropenic phase but also in the presence of GVHD, the main risk factor for aspergillosis in allogeneic bone marrow transplant recipients.^{11, 12}

The aim of this study was to identify retrospectively fungal relapses after pulmonary resection of localized filamentous fungal infections, in particular after elective surgery carried out immediately before transplantation, to remove possible sites of active infection.

Patients and methods

Between 1997 and 2004, 407 patients with acute leukemia (335 myeloid, 72 lymphoid) were admitted to our Hematology Division. Fifty of them had documented (30) or probable (20) filamentous fungal infection according to Mycoses Study Group/European Organisation for Research and Treatment of Cancer (MSG/EORTC) criteria.¹³ Among the proven filamentous fungal infections, 21 were aspergillosis, eight were zygomycosis and one was fusariosis. Ten of these 50 patients, median age 48 years (range 28–65), seven male patients and three female patients, with acute myeloid leukemia (AML) in nine cases and acute lymphoblastic leukemia in one case, underwent pulmonary surgery for invasive mycosis. All 10 patients received induction therapy for newly diagnosed acute leukemia before invasive fungal infection was suspected or diagnosed. Empirical antifungal therapy with amphotericin B deoxycholate 1 mg/kg was added in febrile neutropenic patients with negative cultures and resistant to empiric broad spectrum antibiotic therapy after 5–7 days. Within 48 h of starting antifungal drugs, a high resolution thoracic (HRCT) scan was performed to reveal any localized infiltrates, with the presence of the 'halo sign' and later the appearance of cavities, both signs strongly suggestive of filamentous fungal infection in neutropenic patients. HRCT scan was repeated (1) when clinically indicated, (2) at the increase of neutrophil count, (3) at the beginning of each subsequent chemotherapy cycle and (4) before pulmonary surgery. Bronchoscopy with bronchoalveolar lavage (BAL) was performed routinely in all patients. Since 2002, when fungal infection was suspected, the galattomannan antigen test was undertaken twice weekly from the beginning of empiric antifungal therapy. If both cultures and galattomannan antigen were negative, patients underwent percutaneous biopsy of the pulmonary lesions as soon as possible. A flow chart of our diagnostic procedures is presented in Figure 1.

Figure 1.

Diagnostic procedures in acute leukemia patients with suspected invasive filamentous fungi infection.

Full figure and legend (17K)

Amphotericin B deoxycholate cessation and change to a lipid formulation was undertaken in the presence of nephrotoxicity (twice the baseline creatinine value). Indications for surgery were classified as emergency and elective. Elective surgical resection was considered for patients with a good hematologic response to chemotherapy at the end of their chemotherapy program, immediately before transplantation, to reduce the risk of fungal relapse and to make an accurate diagnosis of the fungus involved. Emergency surgery was performed when the mycotic lesion was situated close to the main pulmonary artery or its branches, in the presence of hemoptysis. Platelet transfusions before surgery were administered when the count was <50 10⁹/l. Informed consent was given by each patient before surgery.

All surgical specimens were examined postoperatively by standard pathologic techniques. Hyphae were visualized in tissue sections stained with periodic acid-Schiff or Grocott-Gomori methanamine-silver nitrate. A histologic diagnosis of Aspergillus was made when typical septate hyphae with dichotomous branches appeared in biopsy specimens; mucorales were diagnosed when irregularly shaped, broad non-septate hyphae with branches occurring at right angles were present. All samples were cultured for bacteria and fungi.

Results

Results of the diagnostic procedures and outcomes of our 10 patients who underwent pulmonary surgery are summarized in Table 1. Fungal cultures of sputum gave a positive result before surgery in only 2/10 patients (Aspergillus flavus and Aspergillus fumigatus, respectively). Histologic diagnosis of invasive moulds was obtained by BAL in 2/10 cases of aspergillosis and by percutaneous lung biopsy in 2/3 cases; aspergillosis and mucormycosis respectively. This diagnosis of mucormycosis was confirmed on pulmonary lobectomy. Patient 10, the only one positive for galattomannan antigen among our four patients in whom the test was performed, had no positive cultures or histology to confirm the diagnosis.

Table 1 - Diagnosis, surgery and outcome of patients.

Full table

The median time from diagnosis of mycosis to surgery was 135 days (range 21–147 days) and before surgery, the median total administered dosage of amphotericin B was 850 mg (range 170–1700 mg). During chemotherapy subsequent to infection, all patients except two were treated with lipid formulations because of nephrotoxicity owing to amphotericin B. One patient received amphotericin B lipid complex (5 mg/kg/day, total dose 2450 mg followed by oral voriconazole 400 mg/day). Seven patients received liposomal amphotericin (L-Amb) (3 mg/kg/day, total dose, median, 6100 mg, range 4200–9350 mg, followed by oral voriconazole 400 mg/day in one patient, and oral itraconazole 400 mg/day in the remaining six cases).

Three patients underwent emergency lobectomies (patients 5, 7 and 8), one before marrow recovery, because of hemoptysis; on HRCT chest scans these patients had wide, cavitating lesions that came into contact with the pulmonary vessels. In the other seven patients, who had only one nodule/cavitation remaining after prolonged antifungal treatment, pulmonary surgery (three wedge resections, four lobectomies) was scheduled before transplantation to eliminate residual pulmonary lesions and, in the case where the fungus had not previously been identified, to make a definitive diagnosis. Histology confirmed vessel invasion and thrombosis by hyphae in 5/10 patients, two with aspergillosis and three with zygomycosis. All three patients who underwent emergency surgery owing to hemoptysis had histological confirmation of their mycosis (two mucormycosis, one aspergillosis). Positive cultures from the resected lung tissue were obtained in only two patients (one aspergillosis, one mucormycosis). The only side effect was pneumothorax, which required tube insertion, in one patient after a wedge resection. Duration of post-surgery hospitalization was a median of 7 days (range 5–10).

Nine of 10 patients were considered cured of their mycosis. The remaining patient, a young woman with resistant AML, underwent lobectomy owing to hemoptysis (histologic diagnosis: mucormycosis) and died owing to leukemia progression in the presence of additional pulmonary infiltrates. Six patients underwent bone marrow transplantation (two allogeneic, three autologous, one syngeneic) with secondary antifungal prophylaxis with L-Amb 3 mg/kg/day, until bone marrow recovery in five patients, and with oral voriconazole 400 mg/day in one patient; none had fungal relapse, although grade II–III GVHD was present. Seven patients had a hematologic relapse, three after chemotherapy only, three after autologous and one after syngeneic bone marrow transplantation, and all were treated with aggressive salvage chemotherapy. Of these, only one 65 years old patient with previous zygomycosis had a fungal relapse during the aplasia after salvage chemotherapy. The main cause of death was relapse of acute leukemia; only one patient died owing to both leukemia progression and relapsed mucomycosis; none of the other patients suffered from recurrent fungal infection. The median duration of survival after diagnosis of IPA was 13.5 months (range 5–85).

Discussion

Filamentous fungal infections are a frequent complication in the management of both acute myeloid and lymphoid leukemia.^{14, 15} These mycoses are present particularly after aggressive chemotherapy when the hematologic disease is active, such as at diagnosis and during induction therapy,¹⁶ and also during salvage therapy in resistant and relapsed patients. Moreover, aspergillosis is increasing in allogeneic bone marrow transplant recipients, particularly in patients with GVHD.¹² Mycoses result in a high mortality rate of about 40–50% in acute leukemia during standard therapy and autologous transplantation^{2, 16} and 80–90% after allogeneic transplantation.^{2, 12} Early diagnostic procedures, such as periodic HRCT chest scanning, are able to indicate mycotic infection promptly, justifying empiric antifungal treatment with improved outcome.¹⁷ Even if this strategy is likely to have reduced mortality in leukemia patients undergoing chemotherapy, the problem remains of not delaying chemotherapy and eventual transplantation. When bone marrow transplantation is scheduled, pulmonary nodules with possible presence of mycosis increase the risk of fungal relapse after transplant;¹⁰ on the other hand, several publications confirm a lower rate of reactivation of fungal infections in transplanted patients without mycosis.^{10, 18, 19} It is therefore important to commence transplantation free of fungal infection or, if this is not possible, with reduced sites of fungal involvement, which are controllable with an effective prophylaxis. Finally, diagnosis of the fungal species permits appropriate secondary prophylaxis, which seems essential in avoiding fungal relapse during transplant, as shown by some reports.^{10, 20} Cordonnier et al.²⁰ showed no fungal relapse in 11 allotransplanted patients, six in complete and five in partial remission of mycosis, who were treated with voriconazole 400 mg/day as secondary prophylaxis. Another report¹⁰ described the outcome of 18 hematology patients with previous proven invasive fungal infections, who underwent allogeneic stem cell transplantation; aggressive secondary prophylaxis with L-Amb 5 mg/kg/day was given pre and during the transplant period, and surgical resection of the fungal mass before transplant was performed in three cases. In this series of patients, the presence of both active leukemia and active fungal infection increased procedure-related mortality. Three patients (17 %) died owing to reactivation (one patient) or progression (two patients also with active leukemia) of fulminant fungal infection following their allogeneic transplant.

In our group of patients, the presence of sporadic, but not particularly rare, cases of zygomycosis caused us to identify the fungal species with all possible diagnostic procedures, including sputum, broncoscopy with BAL, and percutaneous biopsies.²¹ Although sputum and BAL gave modest results in hematology patients,^{14, 22} in our experience, percutaneous lung biopsies were accurate and specific.²³ However, many patients remain classified as having 'possible' aspergillosis, underestimating the presence of other filamentous fungi, such as zygomycetes, which afford the same radiological picture but grow poorly in culture; in fact, the standard for diagnosing zygomycosis is histopathologic examination of infected tissue.²⁴ After surgery, zygomycosis was diagnosed histologically in three of our patients, two of these with no previous proven diagnosis. These results were very useful in deciding on the appropriate secondary prophylaxis, particularly as new antifungal drugs have become available, which are all effective against aspergillosis, but not against zygomycosis.²⁵

In three patients lobectomy was performed early, as an emergency procedure, owing to hemoptysis, which quite commonly occurs frequently in the phase of granulocyte recovery. Neutrophils are important in clearing mycoses; however, their proteolytic enzymes are believed to cause destruction of vascular structures.^{6, 7, 8, 9, 26} In hematology patients with aspergillosis, hemoptysis is often massive and the first episode can be fatal.⁸ Two of our three patients, who underwent emergency surgery owing to hemoptysis, had zygomycosis. Although it is rarer than aspergillosis, zygomycosis carries a high mortality rate in hematology patients,^{27, 28, 29} as is our experience.³⁰ Its rapid invasion is directly related to neutropenia; in this situation early diagnosis and aggressive treatment are imperative. The three patients, who underwent emergency resection owing to hemoptysis, were the only patients who came to surgery with little previous antifungal treatment, and all had a histological diagnosis of fungal infection. The other seven patients were scheduled for surgery only after consolidation chemotherapy, a median of 4.5 months from diagnosis and after prolonged antifungal treatment, administered until surgery. In five of these seven patients, fungus was not present in the resected nodules; evidently, the prolonged antifungal treatment had eradicated the filamentous fungi from the nodules, which remained only fybrotic. The percentage (29%) of histologic positivity was lower in comparison to other studies, when surgery was performed earlier.^{31, 32}

Antifungal therapy alone might be insufficient to eradicate extended localizations rapidly. Surgery permits the elimination of residual nodules or cavities, which, even if sterile, can be newly colonized by other bacteria or fungi. In our experience, surgical resection of residual fungi combined with antifungal drugs, achieves local control avoiding possible mycotic relapse. Similar positive results were seen after pulmonary surgery in bone marrow transplant patients were seen by Yeghen et al.³³ Among their 39 hematology patients with mycosis treated early with lung resection, only eight underwent bone marrow transplantation; seven allogeneic and one autologous, with 71% survival rate at 2 years. Two of these eight patients died owing to leukemia relapse, but none had relapse of mycosis. In our population, only seven of our 50 patients with mycosis came to transplant in complete remission of their hematologic disease, but with persistence of only one mycotic pulmonary nodule; none had fungal relapse after surgery. In the rest of our patients, the length of antifungal treatment began during the first chemotherapy cycle and continued until transplant, often resulted in resolution of the pulmonary lesions, especially in patients in complete remission of leukemia. The recent availability of oral voriconazole has also allowed effective antifungal therapy after discharge, probably reducing the time needed for resolution of fungal lesions.

In conclusion, surgical resection of residual pulmonary mycosis combined with secondary antifungal prophylaxis seem to be effective in preventing mycotic relapse during chemotherapy and transplantation in selected patients scheduled for bone marrow transplantation, and is not associated with severe complications.

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