Diagnosis and Treatment of Invasive Mold Diseases

1. Host factors

The EORTC/MSG definitions were originally created for use in patients with cancer and hematologic malignancy and HSCT recipients [10]. Therefore, in the 2002 EORTC/MSG definitions, host factors were mainly composed of neutropenia, persistent fever refractory to broad-spectrum antimicrobial treatment, graft-versus-host disease, and prolonged (>3 weeks) use of corticosteroids. In the 2008 revised definitions, various immunosuppressant treatments were also included to include SOT recipients [16]. The 2020 definitions listed more specific situations [17]. Table 1 compares the 2008 and 2020 definitions and presents them in detail.

Table 1
Comparison of the 2008 and 2020 EORTC/MSG definitions for probable invasive pulmonary mold diseases

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2. Clinical criteria

Clinical criteria of the 2002 EORTC/MSG definitions consisted of radiologic findings and clinical symptoms/signs [10]. For example, to diagnose pulmonary IMDs, chest CT findings required a halo sign, air-crescent sign, or cavity within the area of consolidation. Clinical symptoms/signs were coughs, chest pain, hemoptysis, dyspnea, and pleural rub. In the 2008 definitions, clinical symptoms/signs were excluded from clinical criteria because they were usually non-specific for the diagnosis of pulmonary IMDs [16]. In the 2008 definitions, radiologic findings were also revised; with only dense, well-circumscribed lesions, radiologic criteria for pulmonary IMDs were satisfied regardless of the presence of a halo sign (Table 1). It was because halo signs appear between the initial 7 - 10 days and might not be visible thereafter [18, 19].

There were some additional changes in the 2020 definitions [17]. Radiologic patterns of pulmonary IMDs were classified as either angio-invasive or airway-invasive forms [20, 21]. Angio-invasive forms, such as halo sign, air-crescent sign, and cavity within an infarct-shaped consolidation, were observed more often in neutropenic patients [22, 23]. Because the EORTC/MSG definitions were originally focused on patients with cancer and hematologic malignancy, radiologic patterns of angio-invasive form were mainly included even in the 2008 definitions. Airway-invasive forms, such as small airway lesions, peribronchial consolidations, and bronchiectasis, are often observed in non-neutropenic transplant patients [22, 23]. Based on the results of these studies, the findings of wedge-shaped and segmental or lobar consolidation were added to the 2020 definitions (Table 1).

The 2020 definitions were characterized by separating the clinical criteria for pulmonary aspergillosis and other pulmonary mold diseases (Table 1). The reverse halo sign was added as the clinical criteria for other pulmonary mold diseases such as mucormycosis. The reverse halo sign is more common in patients with pulmonary mucormycosis than in those with pulmonary aspergillosis [24]. The histology of the reverse halo sign includes an infarcted lung with a greater amount of hemorrhage in the peripheral solid ring than in the center ground-glass region [25].

While the clinical criteria of the EORTC/MSG definitions are presented by pulmonary diseases, tracheobronchitis, sinonasal diseases, and central nervous system infection, Table 1 only summarizes pulmonary mold diseases, which are the most representative type.

3. Mycological criteria

According to the mycological criteria of the EORTC/MSG definitions, direct tests (i.e., cytology, direct microscopy, or culture) or indirect tests (i.e., detection of antigen or cell-wall constituents) can be used for the diagnosis of IMD (Table 1) [10, 16, 17]. If an appropriate specimen can be obtained, we can identify the presence of mold from the smear and culture, otherwise, indirect tests could be used. Galactomannan (GM) assay can be used for the diagnosis of aspergillosis, but there is no specific test for other mold diseases.

In experimental assays, the detection of Aspergillus antigenemia seems to correlate with clinical diagnosis and treatment response of antifungal therapy in invasive aspergillosis [26, 27]. A sandwich ELISA technique that uses a monoclonal antibody to GM has been developed [28]. The GM assay (Platelia Aspergillus, Bio-Rad, Hercules, CA, USA) could be applied to cerebrospinal fluid and bronchoalveolar lavage fluid as well as plasma and serum [16]. The GM assay is performed with an optical read-out that is interpreted as a ratio relative to the optical density (OD) of a threshold control provided by the manufacturer; this ratio is called the OD index. The cut-off index for the GM assay was originally set at 1.0 - 1.5 in order to minimize false-positive results [10], but it was lowered to 0.5 in the 2008 EORTC/MSG definitions [16]. The lowered cut-off value improved the overall performance of the test for adult hematology patients [29]. In the 2020 EORTC/MSG definitions, however, the cut-off index has been raised again (1.0 in serum, plasma, or bronchoalveolar lavage fluid) in order to ensure a higher likelihood of diagnostic certainty for clinical trial purposes (Table 1) [17, 30].

The determination of (1,3)-β-d-glucan (BDG) serum levels is a noninvasive test for circulating fungal cell wall components. The BDG assay is not specific for Aspergillus species and can produce positive results in patients with a variety of IFDs, including candidiasis and Pneumocystis jirovecii. However, it is typically negative in patients with mucormycosis or cryptococcosis [31, 32]. There are several different commercial assays available in different countries. A commercial BDG assay (Fungitell assay, Associates of Cape Cod, East Falmouth, MA, USA) has been approved by the FDA. However, BDG was not considered to provide mycological evidence of any IMD [17] since BDG detection is not specific to a single IMD.

In the 2008 EORTC/MSG definitions, molecular methods of detecting fungi in clinical specimens (e.g., polymerase chain reaction [PCR]) were not included in the definitions because there was as yet to be a standard, and none of the techniques had been clinically validated [16]. Since then, much progress has been made in PCR assays for Aspergillus. In a meta-analysis of 25 studies, the sensitivity and specificity of PCR to detect invasive aspergillosis were 84.0% and 76.0%, respectively. When at least two PCR results were positive, the sensitivity was 64.0% and the specificity was 95.0% [33]. Many other meta-analyses showed similar results. Standardized protocols for nucleic acid extraction, sample types, volumes, and processing have been developed by the European Aspergillus PCR Initiative/Fungal PCR Initiative [34]. As a result of these efforts, 2 or more consecutive positive results of Aspergillus PCR was included in the mycological criteria of the 2020 EORTC/MSG definitions (Table 1) [17].

4. Diagnostic criteria for other host groups

In recent years, there has been an increasing number of reports of IMDs accompanying various infectious diseases. Invasive pulmonary aspergillosis can develop in patients admitted to intensive care units due to severe influenza or severe fever with thrombocytopenia syndrome [35, 36, 37]. The category of proven IMD can apply to any patient, regardless of whether the patient is immunocompromised. However, probable IMD requires the presence of at least 1 host factor, clinical and mycological criteria, and is proposed for immunocompromised patients only [16, 17]. As severe influenza or severe fever with thrombocytopenia syndrome are not a host factor for IMD, efforts to create new diagnostic algorithms are ongoing [36, 38, 39]. In the coronavirus disease 2019 (COVID-19) pandemic situation, many cases of coronavirus disease-associated pulmonary aspergillosis (CAPA) were reported [40, 41]. In order to diagnose CAPA in patients with COVID-19, diagnostic criteria are being created by applying criteria for influenza-associated pulmonary aspergillosis (IAPA) and the EORTC/MSG definitions [42].

1. Initial therapy for invasive aspergillosis

For initial primary therapy of invasive aspergillosis, voriconazole is recommended (Table 2) [43, 44, 45]. Even in the early 2000s, conventional amphotericin B deoxycholate was the standard therapy for invasive aspergillosis, although responses were suboptimal (less than 40.0%) and it had multiple adverse effects [46]. Voriconazole gained its current status based on a randomized, unblinded trial published in 2002 [47] that compared 144 patients in the voriconazole group and 133 patients in the amphotericin B group with definite or probable aspergillosis. This trial reported that compared with amphotericin B, initial therapy with voriconazole led to better responses (successful outcome at week 12, 52.8% vs. 31.6%) and a better survival rate at 12 weeks (70.8% vs. 57.9%) while resulting in fewer severe side effects such as nephrotoxicity.

Table 2
Antifungal treatment of invasive aspergillosis

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During voriconazole therapy, therapeutic drug monitoring (TDM) is recommended [43, 44, 45]. The target trough level of voriconazole is 1 - 5.5 mg/L. Routine TDM of voriconazole may reduce the incidence of drug discontinuation due to adverse events and improve the treatment response [48]. Considering that voriconazole interacts with various drugs such as cyclosporine, tacrolimus, and sirolimus, TDM of potentially interacting drugs is also needed. Elevation of hepatic enzymes is the most common adverse event of voriconazole, followed by hallucination and visual disturbance [48].

Isavuconazole is the primary or primary alternative drug for the treatment of invasive aspergillosis [43, 44, 45]. In phase 3, randomized-controlled trial, isavuconazole was non-inferior to voriconazole for the primary therapy of IMDs including invasive aspergillosis [12]. In the isavuconazole group of this trial, hepatotoxicity, visual disturbance, and rash were less common than in the voriconazole group. However, the incidence of audio or visual hallucinations was similar between the two groups. TDM of isavuconazole is not routinely recommended because there is no clear exposure-response relationship and low variability of plasma level [49].

Recently, a phase 3, randomized-controlled trial assessed the non-inferiority of posaconazole to voriconazole for the primary therapy of invasive aspergillosis [13]. In this trial, posaconazole was non-inferior to voriconazole for all-cause mortality until day 42 and had fewer adverse events than voriconazole. Although posaconazole is recommended as salvage therapy in the recent international guidelines (Table 2) [43, 44, 45], it might be included as primary therapy in the next revision of the guidelines.

Liposomal amphotericin B or amphotericin B lipid complex are additional primary alternatives but these agents carry the risk of nephrotoxicity [43, 44, 45]. Although no randomized trial has been performed to evaluate the efficacy of these drugs compared with voriconazole as primary therapy, a series of randomized trials suggest their efficacy. A randomized trial evaluating the primary treatment of invasive aspergillosis using liposomal amphotericin B reported favorable outcomes, especially with regard to minimizing toxicities [50]. Lipid formulations of amphotericin B are useful as initial therapy for invasive aspergillosis in patients using drugs that interact with azoles. They are also useful for patients suspected of invasive aspergillosis and mucormycosis, but accurate diagnosis is difficult, especially in patients who have used prophylactic voriconazole for a long time.

2. Salvage therapy of invasive aspergillosis

Salvage therapy is a form of therapy given after a disease becomes refractory to or intolerant of primary therapy. If voriconazole or isavuconazole was used for primary therapy, lipid formulations of amphotericin B can be used as salvage therapy [44]. Representative antifungal agents for salvage therapy are echinocandins such as caspofungin, micafungin, and anidulafungin [43, 44, 45]. Caspofungin was approved as salvage therapy for invasive aspergillosis due to its high efficacy and acceptable safety [51]. In a study of 225 patients with invasive pulmonary aspergillosis, favorable responses were observed in 50.0% and 41.0% of patients treated with micafungin as primary or salvage therapy, respectively [52]. A unique attribute of anidulafungin is that it is eliminated by non-enzymatic degradation in the blood and does not require dosage adjustments in patients with renal or hepatic dysfunction [53].

Combination therapy for primary therapy of invasive aspergillosis is not routinely recommended, but it may be considered in a salvage situation. Because of their distinct mechanism of action, echinocandins have the potential for use in combination with antifungal agents with different mechanisms of action [14, 54]. Antifungal agent resistance due to antifungal abuse raised concern, and efforts for antifungal stewardship should not be neglected [55].

1. Hyalohyphomycosis

Although infection due to Aspergillus spp. fits the description of colorless hyphae, aspergillosis is typically not included in hyalohyphomycosis, which includes various species such as Fusarium, Scedosporium, Penicillium, and Acremonium [62]. In patients with hematologic malignancy and HSCT recipients, almost all cases of fusariosis are disseminated at presentation [5, 63]. In contrast, fusariosis that occurs after SOT tends to be localized, and the outcome is better than in patients who have neutropenia [64]. Voriconazole or a lipid formulation of amphotericin B is recommended as primary therapy for invasive fusariosis (Table 3) [63].

Scedosporium spp. causes a wide spectrum of conditions, including mycetoma, colonization of the airway, sinopulmonary infections, extrapulmonary localized infections, and disseminated infections [65]. SOT from a nearly-drowned donor resulted in cases of fatal scedosporiosis [4]. As for primary therapy for invasive scedosporiosis, voriconazole is recommended together with surgical debridement when possible [63].

2. Phaeohyphomycosis

Phaeohyphomycosis is the result of infection with various species including Alternaria, Exophiala, Dactylaria, Cladophialophora, and Curvularia [62]. They cause a broad spectrum of diseases including skin and subcutaneous lesions, pneumonia, central nervous system disease, fungemia, and disseminated disease, particularly in immunocompromised patients. Extracutaneous invasive diseases can also occur in immunocompetent patients but are much less common. Skin and subcutaneous diseases were more common in SOT recipients, while pulmonary diseases were more common in HSCT recipients [66, 67].

There is no standard therapy for phaeohyphomycosis, and depends on the clinical disease and status of the patient. Surgical excision of the skin or subcutaneous lesion is often curative, although antifungal therapy is usually given in conjunction with surgery. Voriconazole, posaconazole, and itraconazole showed the most consistent in vitro activity against phaeohyphomycosis. Lipid formulations of amphotericin B have also been useful as an alternative therapy in some cases (Table 3). Combination antifungal therapy is recommended for cerebral abscesses when surgery is not possible and for disseminated infections in immunocompromised patients [66].