Variation among biosynthetic gene clusters, secondary metabolite profiles, and cards of virulence across Aspergillus species

<i>Aspergillus fumigatus </i>is a major human pathogen. In contrast, <i>Aspergillus</i> <i>fischeri</i> and the recently described <i>Aspergillus</i> <i>oerlinghausenensis</i>, the two species most closely related to <i>A. fumigatus</i>, are not known to be pathogenic. Some of the genetic determinants of virulence (or “cards of virulence”) that <i>A. fumigatus </i>possesses are secondary metabolites that impair the host immune system, protect from host immune cell attacks, or acquire key nutrients. To examine whether secondary metabolism-associated cards of virulence vary between these species, we conducted extensive genomic and secondary metabolite profiling analyses of multiple <i>A. fumigatus</i>, one <i>A. oerlinghausenensis</i>, and multiple<i> A. fischeri</i> strains. We identified two cards of virulence (gliotoxin and fumitremorgin) shared by all three species and three cards of virulence (trypacidin, pseurotin, and fumagillin) that are variable. For example, we found that all species and strains examined biosynthesized gliotoxin, which is known to contribute to virulence, consistent with the conservation of the gliotoxin biosynthetic gene cluster (BGC) across genomes. For other secondary metabolites, such as fumitremorgin, a modulator of host biology, we found that all species produced the metabolite but that there was strain heterogeneity in its production within species. Finally, species differed in their biosynthesis of fumagillin and pseurotin, both contributors to host tissue damage during invasive aspergillosis. <i>A. fumigatus</i> biosynthesized fumagillin and pseurotin, while <i>A. oerlinghausenensis</i> biosynthesized fumagillin and <i>A. fischeri</i> biosynthesized neither. These biochemical differences were reflected in sequence divergence of the intertwined fumagillin/pseurotin BGCs across genomes. These results delineate the similarities and differences in secondary metabolism-associated cards of virulence between a major fungal pathogen and its nonpathogenic closest relatives, shedding light onto the genetic and phenotypic changes associated with the evolution of fungal pathogenicity.

烟曲霉（Aspergillus fumigatus）是一类重要的人类致病菌。与之相对，与烟曲霉亲缘关系最近的两个物种——费希尔曲霉（Aspergillus fischeri）与新近报道的厄林豪森曲霉（Aspergillus oerlinghausenensis），目前尚未被发现具有致病性。烟曲霉所携带的部分毒力遗传决定因子（亦称“毒力因子”）为次级代谢产物，这些产物可损害宿主免疫系统、抵御宿主免疫细胞的攻击，或获取关键营养物质。为探究与次级代谢相关的毒力因子在这三个物种间是否存在差异，我们对多株烟曲霉、1株厄林豪森曲霉以及多株费希尔曲霉开展了全面的基因组分析与次级代谢产物谱分析。我们鉴定出两类在三个物种中均存在的毒力因子——胶霉毒素（gliotoxin）与富马替莫菌素（fumitremorgin），以及三类存在物种间差异的可变毒力因子：锥曲菌素（trypacidin）、假尿嘧啶素（pseurotin）与烟曲霉素（fumagillin）。例如，我们发现所有受试物种与菌株均能生物合成胶霉毒素——该物质已被证实可促进致病进程——这与胶霉毒素生物合成基因簇（biosynthetic gene cluster, BGC）在各基因组间的保守性相符。对于其他次级代谢产物，例如宿主生理调节因子富马替莫菌素，我们发现所有物种均可产生该代谢物，但同一物种内的不同菌株在其产量上存在异质性。最终，三个物种在烟曲霉素与假尿嘧啶素的生物合成上存在显著差异，这两种物质均会在侵袭性曲霉病进程中造成宿主组织损伤。烟曲霉可生物合成烟曲霉素与假尿嘧啶素，厄林豪森曲霉仅可合成烟曲霉素，而费希尔曲霉则无法合成这两种物质。这些生化层面的差异与各基因组间相互交织的烟曲霉素/假尿嘧啶素生物合成基因簇的序列分化特征相一致。本研究阐明了主要致病真菌病原体与其非致病性近缘物种间，与次级代谢相关的毒力因子的异同，为解析真菌致病性演化相关的遗传与表型变化提供了新的认知。