3. Discussion
We report the first case of azole-resistant CAPA, which occurred in an immunocompetent host during ICU support without a previous history of azole therapy. The A. fumigatus cyp51A gene TR34/L98H mutation found in this patient has been well described as an environmentally acquired mutation [16], which is in line with data from clinical studies where two-thirds of patients with azole-resistant infections had no history of azole pretreatment [10]. This case underscores the importance of early diagnosis and the need for resistance surveillance, comparable to what has been described in influenza patients [9,17], given the emergence of triazole resistance [18,19].
The sensitivity for detection of resistance in primary cultures with the VIPcheck plate depends on the number of A. fumigatus colonies that are tested, as clinical cultures may contain both mixed azole-susceptible and azole-resistant isolates during an infection [20]. We suspect that A. fumigatus isolated in the first tracheal aspirate was already a mixed culture but was missed in initial fungal cultures due to abundance of azole-susceptible A. fumigatus spores. Molecular detection could have given a suggestion to the presence of a mixed culture [21] but PCR could not be performed due to absence of material. The TR34/L98H had a phenotype with high itraconazole MIC (>16 mg/L) and low voriconazole MIC (2 mg/L), similar to strains which have been described only recently in the Netherlands [22].
IPA is known to be problematic to diagnose in the non-neutropenic ICU host [23]. Regardless of the compelling evidence for CAPA in this patient, the EORTC/MSGERC [24] host criteria for invasive fungal disease were not met, nor did the patient meet the AspICU algorithm because we tested tracheal aspirates instead of bronchoalveolar lavage (BAL) fluid [25]. This is in line with findings from other groups, where CAPA patients did not meet the EORTC/MSGERC host criteria either [3,4,5,6]. In addition, the American Association for Bronchology and Interventional Pulmonology (AABIP) has issued a statement advising against routine bronchoscopy in COVID-19 patients, as it poses substantial risk to patients and staff [26]. BAL should only be considered in intubated patients if upper respiratory samples are negative and BAL would significantly change clinical management. Tracheal aspirate cultures, as performed twice weekly in our patient, repeatedly identified A. fumigatus as the only micro-organism present. In the first positive culture, five colonies were tested for resistance with the VIPcheck plate as is recommended to exclude azole resistance [15]. When surveillance cultures of tracheal aspirates were persistently cultured positive with A. fumigatus during voriconazole therapy, we suspected the selection of resistant isolates which were probably already present in the first samples, albeit in undetectable numbers. An autopsy to confirm IPA was not done.
Serum galactomannan testing has been shown to be a fairly sensitive diagnostic tool (70%) in neutropenic patients with pathology-proven invasive aspergillosis [27,28]. However, in patients who are non-neutropenic, serum galactomannan sensitivity of around 25% has been reported [27], which may explain the low number of serum galactomannan positive findings in recently published case reports [6,7] and case series [3,4,5]. The role of β-d-glucan and the Aspergillus-specific lateral flow device (LFD) as an adjunct to the diagnosis of IPA in COVID-19 is not yet clear [2,23]. Serum β-d-glucan was persistently strongly positive in this patient over the course of a week. The specificity for invasive fungal disease of β-d-glucan testing in a mixed ICU population has been shown to be high (86%), with two consecutive positive results [29] compared to those with only fungal colonization and no invasive fungal disease. In addition, multiple other studies report a good sensitivity for the diagnosis of invasive aspergillosis in critically ill patients [30,31,32,33,34]. BAL β-d-glucan in the ICU setting is, however, not recommended, due to its poor specificity and confounders causing false positive results [35].
The LFD is particularly interesting in the ICU due to its short turnaround time. It has demonstrated a higher sensitivity but lower specificity in BAL fluids compared to galactomannan [36] and β-d-glucan [37] in IPA-probable and proven immunocompromised patients. In the ICU setting, however, LFD is suggested to have a lower sensitivity but comparable specificity to galactomannan testing in BAL fluids [35,38]. Noteworthily, a negative predictive value of >96% has been reported in the ICU setting [39]. We used the OLM lateral flow device (AspLFD) on sequential patient tracheal aspirates, yielding positive results on all samples confirming the positive galactomannan result. Although suitable for its negative predictive value or as an additional diagnostic measure, further evaluation of lateral flow technology in critically ill patients is warranted.
Altogether, we describe the clinical course of the first reported patient with azole-resistant CAPA. The contribution of A. fumigatus to this fatal COVID-19 course is highly likely, although autopsy was not performed, as in all previously reported CAPA cases [3,4,5,6,7].