A Madurella mycetomatis Grain Model in Galleria mellonella Larvae.

Eumycetoma is a chronic granulomatous subcutaneous infectious disease, endemic in tropical and subtropical regions and most commonly caused by the fungus Madurella mycetomatis. Interestingly, although grain formation is key in mycetoma, its formation process and its susceptibility towards antifungal agents are not well understood. This is because grain formation cannot be induced in vitro; a mammalian host is necessary to induce its formation. Until now, invertebrate hosts were never used to study grain formation in M. mycetomatis. In this study we determined if larvae of the greater wax moth Galleria mellonella could be used to induce grain formation when infected with M. mycetomatis. Three different M. mycetomatis strains were selected and three different inocula for each strain were used to infect G. mellonella larvae, ranging from 0.04 mg/larvae to 4 mg/larvae. Larvae were monitored for 10 days. It appeared that most larvae survived the lowest inoculum, but at the highest inoculum all larvae died within the 10 day observation period. At all inocula tested, grains were formed within 4 hours after infection. The grains produced in the larvae resembled those formed in human and in mammalian hosts. In conclusion, the M. mycetomatis grain model in G. mellonella larvae described here could serve as a useful model to study the grain formation and therapeutic responses towards antifungal agents in the future.

Scedosporium aurantiacum is as virulent as S. prolificans, and shows strain-specific virulence differences, in a mouse model.

Several Scedosporium species are clinically important emerging pathogens. Scedosporium prolificans is reported to be the most virulent of the species, while the recently described species Scedosporium aurantiacum, which accounts for a substantial proportion of Australian clinical isolates is capable of causing a range of serious infections. In addition, environmental surveys have revealed a high prevalence of S. aurantiacum in the urban Sydney region. This study was conducted to assess the virulence of selected S. aurantiacum strains recovered from patients who are colonized or have invasive disease, as well as those from environmental sources, in comparison with S. prolificans. PCR fingerprinting with the primer M13 revealed high genetic variation among the S. aurantiacum strains. We evaluated the virulence of eight S. aurantiacum and two S. prolificans strains in a murine model using an infectious dose of 2 × 105 conidia. S. aurantiacum was noted to be as virulent as S. prolificans, causing death in 60-100% of mice (P > 0.05). There were significant strain-specific virulence differences (P < 0.005), indicating a possible link between genotype and virulence in S. aurantiacum.

Mucormycosis, pseudallescheriasis, and other uncommon mold infections.

Serious infections due to non-Aspergillus molds are being encountered with increasing frequency. Factors likely responsible for the rise of these infections include aging populations in countries with advanced medical technologies, the resultant increase in incidence of many cancers, increasingly intensive myeloablative therapies for these cancers, increasingly intensive care for critically ill patients, and increases in the frequency of solid organ and hematopoietic stem cell transplantation. Although diagnostic and therapeutic modalities have improved, mortality rates for invasive mold infections remain high. In this review, we summarize current knowledge about non-Aspergillus mold infections of the chest, with a focus on risk factors, clinical features, diagnosis, and treatment.

Effects of double and triple combinations of antifungal drugs in a murine model of disseminated infection by Scedosporium prolificans.

We have evaluated the efficacies of micafungin, amphotericin B, and voriconazole, alone and in double and triple combinations, in a murine model of systemic infection by Scedosporium prolificans. Micafungin combined with voriconazole or amphotericin B was the most effective, these being the only treatments able to prolong survival and to reduce the fungal load in the kidneys and brain. Triple combinations of these drugs did not improve the results obtained with double combinations.

Scedosporiosis in patients with acute leukemia: a retrospective multicenter report.

We retrospectively analyzed 542 proven/probable mould infections registered, in the course of 2 studies, in 8,633 patients with acute leukemia, focusing on scedosporiosis. We aimed to define scedosporiosis incidence and mortality rate over a 15-year period. Only 5 cases of scedosporiosis were identified, all of them involving patients with acute myeloid leukemia (AML). We also reviewed all cases of Scedosporium spp. infections in acute leukemia reported to date in the international literature. The 52 cases analyzed confirmed that acute myeloid leukemia is the category with the highest risk of scedosporiosis. Clinical features of scedosporiosis were extremely variable and closely related to patient immune status. Infection disseminated to multiple sites in a very high percentage of patients and outcome was confirmed to be very poor. In our surveys all patients died, in spite of Amphotericin B compounds or voriconazole administration. Our review of literature found scedosporiosis attributable mortality rate (AMR) to be 77%. In conclusion, scedosporiosis, although extremely rare, represents a big problem for clinicians because of its aggressive clinical presentation and the lack of an effective therapy. New drugs with in vitro activity against Scedosporium spp (voriconazole, posaconazole) should be considered. However, their clinical activity should be more widely demonstrated.

Scedosporium infection in a tertiary care cancer center: a review of 25 cases from 1989-2006.

We reviewed the records of patients with cancer who had Scedosporium infection (due to Scedosporium apiospermum and Scedosporium prolificans in 21 and 4 patients, respectively). The incidence of Scedosporium infection increased from 0.82 cases per 100,000 patient-inpatient days (in 1993-1998) to 1.33 cases per 100,000 patient-inpatient days (in 1999-2005). Cases of S. prolificans infection occurred only after 2000. Dissemination occurred in 16 patients (64%). The 12-week mortality rates were 70% and 100% for S. apiospermum and S. prolificans infection, respectively.

Infections due to Scedosporium apiospermum and Scedosporium prolificans in transplant recipients: clinical characteristics and impact of antifungal agent therapy on outcome.

BACKGROUND: Unique characteristics, impact of therapy with antifungal agents, and outcome of infections with Scedosporium species were assessed in transplant recipients. METHODS: The patients comprised a total of 80 transplant recipients with Scedosporium infections, including 13 patients from our institutions (University of Pittsburgh Medical Center [Pittsburgh, PA], University of Maryland [Baltimore], Duke University Medical Center [Durham, NC], Emory University [Atlanta, GA], and Hospital Gregorio Maranon [Madrid, Spain]) and 67 reported in the literature. The transplant recipients were compared with 190 non-transplant recipients with scedosporiosis who were described in the literature. RESULTS: Overall, 69% of the infections in hematopoietic stem cell transplant (HSCT) recipients and 53% of the infections in organ transplant recipients were disseminated. HSCT recipients, compared with organ transplant recipients, were more likely to have infections caused by Scedosporium prolificans (P=.045), to have an earlier onset of infection (P=.007), to be neutropenic (P<.0001), and to have fungemia (P=.04). Time elapsed from transplantation to Scedosporium infection in transplant recipients has increased in recent years (P=.002). The mortality rate among transplant recipients with scedosporiosis was 58%. In a logistic regression model using amphotericin B as comparison treatment, voriconazole was associated with a trend towards better survival (odds ratio [OR], 10.40; P=.08). Presence of disseminated infection (OR, 0.20; P=.03) predicted lower survival, and receipt of adjunctive surgery as treatment (OR, 5.52; P=.02) independently predicted a better survival in this model. CONCLUSIONS: Scedosporium infections in transplant recipients were associated with a high rate of dissemination and a poor outcome overall. The use of newer triazole agents warrants consideration as a therapeutic modality for these infections.

A novel murine model of cerebral scedosporiosis: lack of efficacy of amphotericin B.

OBJECTIVES: Cerebral scedosporiosis is a life-threatening infection that is difficult to treat. The aim of this work was to develop a murine model of cerebral infection by Scedosporium apiospermum using intracranial inoculation and to use this model to evaluate the efficacy of amphotericin B deoxycholate and liposomal amphotericin B. METHODS: Mice were rendered neutropenic by intraperitoneal cyclophosphamide and intravenous (iv) 5-fluorouracil administration. Animals were infected with iv or intracranial inoculation of 1 x 10(4), 5 x 10(4) or 5 x 10(5) cfu of a clinical strain of S. apiospermum. Tissue burden reduction was determined in kidneys and brain 4 days after the infection. Efficacy of amphotericin B and liposomal amphotericin B (0.8 mg/kg/day intraperitoneally and 40 mg/kg/day iv, respectively) was evaluated in neutropenic mice infected iv or intracranially with 5 x 10(4) cfu. Survival was analysed with the log-rank test. Fungal burden values of different groups were compared using the Mann-Whitney U-test. RESULTS: In our model, intracranial infection produced a higher fungal load in the brain and a lower fungal load in the kidney than iv inoculation. Survival of animals infected intracranially and treated with amphotericin B or liposomal amphotericin B (mean survival time = 8.3 and 9.2 days, respectively) was not different from the control group (P=0.58 and 0.85, respectively). CONCLUSIONS: We have developed a murine model of cerebral scedosporiosis, which may be useful for studying various pathological aspects of this infection and evaluating new therapeutic approaches. Amphotericin B and liposomal amphotericin B were unable to resolve the infection.

A murine model of Madurella mycetomatis eumycetoma.

Eumycetoma due to Madurella mycetomatis is a major mycological health problem in endemic areas. We infected BALB/c mice (male or female) with various amounts of M. mycetomatis mycelium, containing sterilized soil as a natural adjuvant or Freund's incomplete adjuvant. Mice differed with respect to age and immune status. Intraperitoneal, subcutaneous and intravenous inoculation was explored and survival was monitored. Mice were killed at various intervals after inoculation, checked for the presence of the characteristic black grains, and organs were cultured for M. mycetomatis. Infected organs were subjected to histopathological examination. Immunocompetent male mice were as susceptible as immunocompromised female mice, but showed higher mortality rates. In conclusion, a reproducible mouse model of intraperitoneal M. mycetomatis infection with characteristic black grains in immunocompetent adult or young female mice was developed. Although this experimental model does not simulate macroscopic features of the subcutaneous M. mycetomatis infection in humans, the histopathological characteristics of the lesions and the development of black grains are clearly representative for the human infection. This model will enable further studies on the pathogenesis as well as prevention and treatment of the fungal infection.

Experimental murine model of disseminated Pseudallescheria infection.

Pseudallescheria boydii is found in soil and has a worldwide distribution. This fungus was initially identified as a pathogen targeting a variety of tissues. There are fragmentary data in the literature on the in vitro susceptibility of P. boydii to different antifungal compounds. P. boydii is highly refractory to antifungal treatments. In this study, a murine model of disseminated Pseudallescheria infection was developed to evaluate efficacy of different treatment regimens. A clinical strain of P. boydii was studied in normal and neutropenic outbred ICR mice. Several inocula were tested over a range from 1 x 10(3) to 5 x 10(6) cfu. Groups of eight mice were injected with a intravenous dose of one inoculum. Mortality correlated with the dose of the inoculum, and with immunosuppression. Quantitative cultures of various tissues showed initial dissemination of disease in immune competent mice. This was followed by, reduction of tissue burden, except in the brain. In contrast, disseminated infection persisted in most organs in immunosuppressed animals (p < 0.0001). This model should be appropriate for in vivo evaluation of antifungal chemotherapy.

Disseminated phaeohyphomycosis: review of an emerging mycosis.

Disseminated phaeohyphomycosis is an uncommon infection caused by dematiaceous fungi, although the number of case reports about this infection has been increasing in recent years. A total of 72 cases are reviewed. Scedosporium prolificans is by far the most common cause. The presence of melanin in their cell walls may be a virulence factor for these fungi. The primary risk factor is decreased host immunity, although cases in apparently immunocompetent patients have been reported. Eosinophilia was seen in 11% of cases. Endocarditis is mostly reported on bioprosthetic valves, particularly those of porcine origin. The outcome of antifungal therapy remains poor, with an overall mortality rate of 79%. Special precautions taken for immunocompromised patients may help prevent exposure to fungi during the patients' period of greatest risk. The development of newer antifungal agents and combination therapy may hold promise in improving the management of these devastating infections in the future.

Outcome of lung transplantation in patients with mycetomas.

BACKGROUND: Lung transplantation has become an acceptable treatment option for many end-stage lung diseases. Pulmonary mycetomas are found in patients with end-stage lung diseases, especially sarcoidosis. The clinical course and long-term outcome of these patients after transplantation remains unknown. METHODS: We reviewed retrospectively the pathology reports of the explanted lungs from all lung and heart-lung transplantations performed at our institution between January 20, 1992, and June 26, 2000. Patients were included in our study if mycetomas were present on the specimens. Information on transplant date and type, diagnosis, information on antifungal therapy and fungal infections pretransplant and posttransplant, and clinical course after transplantation was recorded. RESULTS: Mycetomas were present in 3.0% of transplant recipients (9 of 303 patients). The underlying pulmonary diagnoses were sarcoidosis (six patients), and emphysema, idiopathic pulmonary fibrosis, and pneumoconiosis (one patient each). Seven patients received bilateral lung transplants, one patient received a heart/lung transplant, and one patient received a single lung transplant. Aspergillus was isolated from culture in five patients pretransplant and from five patients posttransplant. Six patients received treatment with itraconazole, or IV or inhaled amphotericin B prior to transplantation. All patients who survived transplantation received posttransplant antifungal therapy. Four patients died in the first month after transplantation. Two patients died at 17 months and 24 months posttransplant, respectively; one patient received a second transplant 30 months later; and two patients are alive and free from fungal infections 17 months and 18 months, respectively, after transplantation. All of the medium-term survivors received lengthy therapy with inhaled and systemic amphotericin B and itraconazole before and after transplantation. CONCLUSIONS: Lung transplant recipients with mycetomas have significantly reduced posttransplant survival. Careful selection of patients and aggressive antifungal therapies before and after transplantation have led to improved outcomes in patients with mycetomas. Additional research is needed to define the best therapeutic strategy for these patients during transplantation.

Airborne outbreak of nosocomial Scedosporium prolificans infection.

We describe six inpatients with acute non-lymphocytic leukaemia who developed invasive infection with Scedosporium prolificans resistant to amphotericin B, flucytosine, ketoconazole, fluconazole, and itraconazole. All six patients died. Phenotypic and genotypic assessment of samples from clinical material and ambient air from the isolation rooms where the patients were being treated showed that the epidemic was caused by a single strain. After implementation of aerial control measures, there were no further infections with this organism. We conclude that fatal multidrug-resistant S prolificans epidemics can be aerially transmitted and can be prevented with implementation of appropriate infection-control measures.

Invasive mold infections in allogeneic bone marrow transplant recipients.

Invasive mold infections (IMIs) are an important cause of morbidity and mortality in patients who are undergoing bone marrow transplantation (BMT). To examine the epidemiology, risk factors, and outcome of IMIs in allogeneic BMT recipients, all cases of mold infection among 94 adult patients who underwent allogeneic BMT at this institution from 1 January 1997 through 31 December 1998 were reviewed retrospectively. Fifteen cases of IMI were identified; infection occurred a median of 102 days after BMT. Aspergillus species was the most common cause of disease, and species other than Aspergillus fumigatus were present in 53% of patients. By multivariate analysis, the variable associated with infection risk was systemic glucocorticosteroid use. Prophylactic antifungal therapy that was targeted to high-risk patients had little effect on disease incidence. These observations suggest that early identification of high-risk patients and better approaches to prevention should be explored, to reduce incidence and severity of disease in this population.

Risk of reactivation of a recent invasive fungal infection in patients with hematological malignancies undergoing further intensive chemo-radiotherapy. A single-center experience and review of the literature.

BACKGROUND: Patients with hematologic malignancies and a history of an invasive fungal infection are considered to be at high risk of suffering reactivation of the infection during subsequent intensive chemotherapy. PATIENTS AND METHODS: From January 1993 to September 1996, nine patients with a hematologic malignancy and previous invasive pulmonary aspergillosis (IPA) or Pseudallescheria boydii pneumonia and five with invasive candidiasis received further intensive chemotherapy (n = 3) or a bone marrow or peripheral blood stem cell transplant (n = 11) four days to 13 months (median three months) from the start of therapy for the fungal infection. Five patients with IPA and all five with invasive candidiasis showed complete or good partial radiologic resolution of the infection with the primary antifungal therapy given, which was continued before, during and after the period(s) of subsequent neutropenia. RESULTS: Twelve of the 14 patients showed no signs of progression or reactivation of the fungal infection during therapy, while two patients with active IPA died with progressive aspergillosis shortly after an allogeneic transplant. A review of the literature revealed that in both types of infections the risk of reactivation and dissemination appears low after achieving clinical and radiologic signs of response, which takes several weeks or months before proceeding to further antileukemic therapy. INTERPRETATION AND CONCLUSIONS: Despite lack of definite evidence, administration of an active antifungal drug before, during and after the period of neutropenia appears to be useful. In IPA, residual masses, nodules or cavities in the lung usually contain viable invasive fungal elements and should be resected whenever possible. On the other hand, the risk of reactivation and progression of an active fungal infection during intensive chemoradiotherapy is very high, and novel therapeutic strategies appear warranted in this setting.

Ten-year experience with mycetomas in patients with pulmonary tuberculosis.

We studied 33 consecutive patients with tuberculous pulmonary cavities complicated by fungus balls to evaluate their treatment. Nineteen had surgical resection for massive or recurrent bleeding or possibility of tumor. One patient died of postpneumonectomy empyema (30-day surgical mortality, 5 percent). Fourteen had no surgery. No patient died of hemoptysis. Respiratory failure contributed most often to death. Hepatic complications and other problems of alcoholism were also prominent. Good results can be obtained by resection in these severely ill patients if care is taken to preserve functioning pulmonary tissue and to avoid complications of alcoholic hepatic disease. Within these constraints, tuberculous cavities complicated by mycetomas should be resected for massive or recurrent hemoptysis.