Heterogeneity among Isolates Reveals that Fitness in Low Oxygen Correlates with Aspergillus fumigatus Virulence

Abstract

Previous work has shown that environmental and clinical isolates of Aspergillus fumigatus represent a diverse population that occupies a variety of niches, has extensive genetic diversity, and exhibits virulence heterogeneity in a number of animal models of invasive pulmonary aspergillosis (IPA). However, mechanisms explaining differences in virulence among A. fumigatus isolates remain enigmatic. Here, we report a significant difference in virulence of two common lab strains, CEA10 and AF293, in the murine triamcinolone immunosuppression model of IPA, in which we previously identified severe low oxygen microenvironments surrounding fungal lesions. Therefore, we hypothesize that the ability to thrive within these lesions of low oxygen promotes virulence of A. fumigatus in this model. To test this hypothesis, we performed in vitro fitness and in vivo virulence analyses in the triamcinolone murine model of IPA with 14 environmental and clinical isolates of A. fumigatus. Among these isolates, we observed a strong correlation between fitness in low oxygen in vitro and virulence. In further support of our hypothesis, experimental evolution of AF293, a strain that exhibits reduced fitness in low oxygen and reduced virulence in the triamcinolone model of IPA, results in a strain (EVOL20) that has increased hypoxia fitness and a corresponding increase in virulence. Thus, the ability to thrive in low oxygen correlates with virulence of A. fumigatus isolates in the context of steroid-mediated murine immunosuppression.

Interleukin 1α Is Critical for Resistance against Highly Virulent Aspergillus fumigatus Isolates

“…Taken together, our data show that initial fungal germination following fungal deposition in the airways is a critical determinant of virulence and the subsequent immune response induced. This further validates the importance of using multiple isolates in studies of pathogenesis and immunity, in order to fully understand the virulence potential of specific pathogens (33,37,38,(53)(54)(55)(56)(57)(58)(59).…”

“…To extend these observations, we screened a number of strains and isolates of A. fumigatus that we previously studied (37) for their ability to germinate in lung homogenate medium. Interestingly, the EVOL20 strain of A. fumigatus, which is a more virulent strain recently generated by in vitro serial passaging of Af293 under low-oxygen conditions (37), was able to germinate more extensively than its parent strain Af293 in lung homogenate medium (Fig. 7C).…”

“…Previous reports indicated variation in virulence among the commonly studied "wild-type" Aspergillus fumigatus strains CEA10 and Af293 (37,38). To examine the overall lung damage induced by each of these strains in an immunocompetent pulmonary inhalation model of Aspergillus fumigatus infection, C57BL/6 mice were challenged with 4 ϫ 10 7 conidia of CEA10 or Af293.…”

“…Preparation of Aspergillus fumigatus conidia. Aspergillus fumigatus strains and isolates CEA10, Af293, 02-10, W72310, and EVOL20 were used for this study, and the origins of these strains and isolates were reported previously (37). Each strain or isolate was grown on glucose minimal medium (GMM) agar plates for 3 days at 37°C.…”

“…Most notably, in those studies, different Aspergillus fumigatus strains were used, CEA10 (22) and Af293 (26,27). It was recently shown that Af293 is significantly less virulent than CEA10 in the triamcinolone model of IA (37). Moreover, both the inflammatory response elicited by these strains and the mortality associated with each strain significantly differed in immunocompetent mice (38).…”