Early oseltamivir reduces risk for influenza-associated aspergillosis in a double-hit murine model

Abstract

Invasive pulmonary aspergillosis (IPA) is a life-threatening fungal infection occurring mainly in immunocompromised patients. We recently identified IPA as an emerging co-infection with high mortality in critically ill, but otherwise immunocompetent influenza patients. The neuraminidase inhibitor oseltamivir is the current standard-of-care treatment in hospitalized influenza patients; however, its efficacy in influenza-associated pulmonary aspergillosis (IAPA) is not known. Therefore, we have established an imaging-supported double-hit mouse model to investigate the therapeutic effect of oseltamivir on the development of IAPA. Immunocompetent mice received intranasal instillation influenza A or PBS followed by orotracheal inoculation with Aspergillus fumigatus 4 days later. Oseltamivir treatment or placebo was started at day 0, day 2, or day 4. Daily monitoring included micro-computed tomography and bioluminescence imaging of pneumonia and fungal burden. Non-invasive biomarkers were complemented with imaging, molecular, immunological, and pathological analysis. Influenza virus-infected immunocompetent mice developed proven airway IPA upon co-infection with Aspergillus fumigatus, whereas noninfluenza-infected mice fully cleared Aspergillus, confirming influenza as a risk factor for developing IPA. Longitudinal micro-CT showed pulmonary lesions after influenza infection worsening after Aspergillus co-infection, congruent with bioluminescence imaging and histology confirming Aspergillus pneumonia. Early oseltamivir treatment prevented severe influenza pneumonia and mitigated the development of IPA and associated mortality. A time-dependent treatment effect was consistently observed with imaging, molecular, and pathological analyses. Hence, our findings underscore the importance of initiating oseltamivir as soon as possible, to suppress influenza infection and mitigate the risk of potentially lethal IAPA disease.

Overcome Double Trouble: Baloxavir Marboxil Suppresses Influenza Thereby Mitigating Secondary Invasive Pulmonary Aspergillosis

“…This drug prevents the release and spread of the influenza virus from the host cell by inhibiting the viral neuraminidase. We found that early oseltamivir modulated the severity of influenza thereby mitigating the susceptibility to IAPA in an immunocompetent murine model [12]. In addition, in the clinical setting, prophylactic oseltamivir proved to be effective in prevention of IAPA in children after haploidentical hematopoietic cell transplantation [14].…”

“…A mouse-adapted influenza virus A/H3N2/Ishikawa/7/82 and a red-shifted thermostable firefly luciferase A. fumigatus strain were prepared, as previously described [12] (manuscript in consideration [28]).…”

“…The experiment was repeated twice. The mouse model was developed as previously described [12]. Briefly, mice received intranasal instillation of 10 plaque forming units (PFU) influenza virus on day 0 (10 µL per nostril) and orotracheally inoculation with 20 µL of A. fumigatus spore solution (5 × 10 9 conidia/mL) under inhalation anesthesia with isoflurane (1.5-2% in 100% oxygen, Primal Critical Care) on day 4.…”

“…Understanding the synergism 2 of 13 between the virus and the fungus is of utmost importance. We recently developed a preclinical animal model of IAPA disease, confirming influenza as a strong risk factor for developing invasive pulmonary aspergillosis (IPA) in immunocompetent mice [12,13].…”

“…In a French multicenter observational cohort study, a highly reduced sensitivity to oseltamivir was found in 23% of the patients with a severe influenza A(H1N1)pdm09 [NA-H275Y] pneumonia [19]. If resistance occurs, it will be important to use other effective antiviral treatments which can lower the severity of the influenza infection and thereby mitigate the susceptibility to IPA [12]. Resistance together with the possible effect of blocking the host neuraminidase indicate the importance to preclinically assess new antiviral drugs, relying on different modes of actions, on the effectiveness in IAPA.…”