Fusarium species are among the leading fungal pathogens to cause invasive mould infections in patients with hematopoietic malignancy. The Fusarium species most frequently involved in human infections are Fusarium solani, Fusarium oxysporum and Fusarium verticillioides. However, identification is a cumbersome and time-consuming task. Fusarium is resistant in vitro to many of the antifungal agents and the management of fusariosis is not well defined.
ObjectivesTo emphasise the difficulty of identifying Fusarium spp. by conventional methods and the need of new rapid molecular tests to achieve earlier diagnosis and appropriate therapy.
MethodsA disseminated Fusarium infection due to F. verticillioides was documented in a neutropenic refractory patient with acute myeloid leukaemia, relapsed after allogeneic hematopoietic stem cell transplantation.
ResultsThe patient died despite liposomal amphotericin B and voriconazole combination and “in vitro” susceptibility of agents employed. Morphological and molecular identification of F. verticillioides was obtained only after the death of the patient.
ConclusionsThis case highlights the poor outcome of an invasive fungal disease caused by Fusarium in aplastic patients. Identification of members of Fusarium genus remains restricted to selected laboratories and should be introduced into routine mycological diagnostics. In immunocompromised patients, diagnosis of fusariosis is directly related to prompt diagnosis and to patient's status. Current diagnosis methods and therapeutic options are discussed.
Fusarium es uno de los principales patógenos fúngicos que provoca infecciones invasoras en pacientes portadores de neoplasias malignas hematopoyéticas. Las especies del género Fusarium implicadas habitualmente en las infecciones del ser humano son Fusarium solani, Fusarium oxysporum y Fusarium verticillioides. No obstante, la identificación es una tarea lenta y que consume mucho tiempo. Fusarium spp. es resistente in vitro a numerosos fármacos antimicóticos y el tratamiento de la fusariosis no está bien definido.
ObjetivosDestacar las dificultades en la identificación de Fusarium spp. por los métodos convencionales y la necesidad de disponer de nuevas técnicas moleculares rápidas para obtener un diagnóstico más precoz y un tratamiento apropiado.
MétodosEn un paciente portador de una leucemia mieloide aguda con neutropenia refractaria, que experimentó recidiva tras alotrasplante de células progenitoras hematopoyéticas se documentó una infección diseminada por Fusarium debida a Fusarium verticillioides.
ResultadosA pesar de recibir un tratamiento combinado con anfotericina B y voriconazol liposómicos y de la sensibilidad in vitro de los preparados administrados, el paciente falleció. Sólo después de su muerte se obtuvo la identificación morfológica y molecular de Fusarium verticillioides.
ConclusionesEl caso descrito en el presente informe destaca el desenlace desfavorable de las micosis invasivas debidas a Fusarium en pacientes con aplasia de médula ósea. La identificación de los miembros del género Fusarium sigue limitándose a laboratorios seleccionados y debe introducirse en el diagnóstico micológico sistemático. En el huésped inmunocomprometido el diagnóstico de fusariosis se relaciona directamente con el estado del paciente. Se describen los métodos diagnósticos y las opciones terapéuticas actuales.
Fusarium species cause severe infections in patients with hematologic malignancies. The Fusarium species most frequently involved in human infections are Fusarium solani, Fusarium oxysporum and Fusarium verticillioides.12,19,27Fusarium spp. are very resistant to single antifungal agents, and new treatment strategies, such as combination therapy, can be considered life-saving for immunocompromised patients.16,27 We report a fatal disseminated F. verticillioides infection in a refractory patient with acute myeloid leukaemia (AML), unresponsive to all antifungal agents, including liposomal amphotericin B and voriconazole combination.
Case reportA 40-year-old Caucasian female was admitted to the haematology unit in January 2010 for AML, M2 FAB subtype, normal karyotype, FLT3 ITD+, NPM1 negative.
After standard induction-chemotherapy and consolidation, in April 2010 the patient underwent a peripheral allogeneic HSCT from HLA-matched family donors, without any serious infectious complications. In August 2010 the patient relapsed and a course of reinduction therapy with Clofarabine and Citarabine was started. Anti-infective prophylaxis included levofloxacin (500mg PO/day), acyclovir (500mg, IV every 12h), and itraconazole (400mg PO/day). Surveillance cultures were taken weekly; Aspergillus galactomannan antigenemia test (GM) was taken 2/times/weekly. No febrile episode was documented during neutropenia, but the treatment failed to achieve a complete remission.5 A new salvage-regimen with MEC (mitoxantrone, etoposide, and cytarabine) plus cyclosporine was started, continuing the same anti-infective prophylaxis. On day +4 post chemotherapy, while neutropenic, the patient experienced fever refractory to broad spectrum antibiotics. Blood cultures, taken from both the peripheral vein and the CVC, chest X-ray and GM were negative. On day 4 of persistent fever, empiric antifungal therapy with liposomal amphotericin B (L-Amb), 3mg/kg/day was added. After 8 days the patient, still febrile, developed multiple skin lesions which had necrotic centres surrounded by spreading erythema; a new chest X-ray, pulmonary CT scan, cultures and GM were negative. It was not possible to obtain cultures through skin biopsy. After 13 days of fever, all new repeated blood cultures tested positive for fungi; L-Amb was raised to a dose of 5mg/kg/day and combined with voriconazole (loading dose 6mg/kg/day, followed by 4mg/kg/day IV every 12h). Five days later the fungus was recognised as a Fusarium species and combination therapy was continued on the basis of data literature susceptibility. A new X-ray and pulmonary CT scan, documented lung involvement with nonspecific findings, although the patient remained asymptomatic for dyspnea, cough and hemoptysis. A new bone marrow aspirate revealed persistence of AML and after 25 days of aplasia and fever the patient died. Six days after the death, following the method and the key proposed by Guarro and Gené,11 the isolated etiological agent was morphologically identified as F. verticillioides.
The confirmation of this species was obtained 20 days later by molecular methods. Fungal DNA was extracted by the Bowman method22 and 5ng of fungal template DNA were amplified in a PCR assay using F. verticillioides specific primers (VER1: 5′-CTTCCTGCGATGTTTCTCC-3′ and VER2: 5′-AATTGGCCATTGGTATTATATATCTA-3′) according to Mulé et al.18 PCR products, consisting of a single band of about 580bp specific for F. verticillioides, confirmed the morphological identification.
For the antifungal susceptibility, the broth microdilution assay was performed according to the Clinical and Laboratory Standards Institute (CLSI).6 The concentrations of amphotericin B, itraconazole, and voriconazole assayed ranged from 64 to 0.06mg/l. The strain was tested in duplicate and the control strains Candida parapsilosis ATCC 22019 and Candida albicans ATCC 90028 were included. The MIC of the strain studied was 8mg/l to voriconazole and 2mg/l to amphotericin B, while it was resistant to the itraconazole maximum tested concentration (64mg/l).
DiscussionDisseminated fusariosis is a serious invasive mould infection in hematologic patients. The most commonly found pathogens are F. solani and F. oxysporum, although other species such as F. verticilloides, have been frequently reported as aetiological agents of human infections.17,28 This fungus in neutropenic patients may affect multiple organs and frequently also the skin as the primary or the metastatic site. The skin is often the single source of diagnosis and cutaneous lesions may precede positive blood cultures for up 5 days.4,9
Currently, the identification to species level of Fusarium is based on the production of macroconidia. However, recognition may be difficult when the macroconidia are not produced in culture; in this case the isolates can be confused with other genera such as Acremonium and Verticillium. In order to solve this issue, new rapid molecular methods are developed. The majority of molecular methods are PCR-based techniques that ensure high sensitivity and specificity and are fully discriminative even for closely related species.2,14 However identification is a time-consuming task, only reserved to trained mycologists.
In our case, recognition of the Fusarium species was obtained only after the death of the patient, while microbiological cultures were still awaiting.
The high mortality caused by Fusarium is attributed to high resistance to many antifungal agents.1 Amphotericin B is the most effective of the antifungal drugs. Fluconazole, itraconazole and flucytosine have no activity against Fusarium spp., and ketoconazole, miconazole, terbinafine and echinocandins have limited activity. F. solani and F. verticillioides are usually resistant to azoles and exhibit higher amphotericin B MICs than other Fusarium species21. The new triazole agents, voriconazole, posaconazole and ravuconazole, exhibit activity against these Fusarium species and have been reported as a successful treatment in oncohematologic patients or in refractory fungal infections.7,15,25
Our patient developed a disseminated fusariosis, after receiving itraconazole prophylaxis. On the other hand, breakthrough fusariosis is not an unexpected event in the immunocompromised patients and may occur also employing agents as voriconazole or posaconazole.3,8
The management of fusariosis is not well defined. There are case reports 10 where the early described F. verticillioides (moniliforme) infections resulted in the death of the patients, and more recently successful cases with patients, treated with the combination of amphotericin B and other agents as caspofungin, voriconazole, terbinafine or posaconazole,13,15,24,26,27,29 suggesting a promising role for this approach, even in the case of late identification to the species level. At present, treatment with voriconazole+amphotericin B is the main alternative.
However patients with severe and prolonged immunosuppression are at high risk for refractory disseminated fusariosis; in fact our patient died after a persistent fungemia, despite early aggressive combination therapy and at least “in vitro” susceptibility of the agents employed.
The failure of antifungal therapy in neutropenic patients with invasive fusariosis has been reported before in a multicentre study.20 Despite the overall relative efficacy, all antifungal agents, including new azoles as posaconazole, were effective only in the case of recovery from myelosuppression.23
In conclusion, morphological and molecular identification of Fusarium spp. remains cumbersome and restricted to selected laboratories; further rapid molecular methods should be introduced into routine mycological diagnostics to achieve earlier diagnosis and appropriate therapy.
In the immunocompromised patients, prognosis of fusariosis remains directly related to prompt diagnosis and to patient's status.
Conflict of interestThe authors declare no conflict of interest.
