Mutations in the Second Alternative Oxidase Gene: A New Approach to Group Aspergillus niger Strains

Abstract

Alternative oxidase is a terminal oxidase in the branched mitochondrial electron transport chain of most fungi including Aspergillus niger (subgenus Circumdati, section Nigri). A second, paralogous aox gene (aoxB) is extant in some A. niger isolates but also present in two divergent species of the subgenus Nidulantes—A. calidoustus and A. implicatus—as well as in Penicillium swiecickii. Black aspergilli are cosmopolitan opportunistic fungi that can cause diverse mycoses and acute aspergillosis in immunocompromised individuals. Amongst the approximately 75 genome-sequenced A. niger strains, aoxB features considerable sequence variation. Five mutations were identified that rationally affect transcription or function or terminally modify the gene product. One mutant allele that occurs in CBS 513.88 and A. niger neotype strain CBS 554.65 involves a chromosomal deletion that removes exon 1 and intron 1 from aoxB. Another aoxB allele results from retrotransposon integration. Three other alleles result from point mutations: a missense mutation of the start codon, a frameshift, and a nonsense mutation. A. niger strain ATCC 1015 has a full-length aoxB gene. The A. niger sensu stricto complex can thus be subdivided into six taxa according to extant aoxB allele, which may facilitate rapid and accurate identification of individual species

Generation, Transfer, and Loss of Alternative Oxidase Paralogues in the Aspergillaceae Family

“…We have described one of the four events in our previous paper on the rare aoxB paralogue found in the Aspergillus niger sensu stricto complex and in three other highly divergent species of Aspergillaceae [13]. In the context of this current work, we renamed this rare aox paralogue aoxB1 (Figure 1a, scheme at the left) because its origin is the furthest away from the tips of the terminal branches (i.e., the present), the small, seemingly homogeneous paralogous AoxB1 clade consistently appearing as the sister clade of the Onygenales AoxA in alternative ML phylogenies (For clarity, all Onygenales species only specify one Aox, AoxA).…”

“…The list includes a specimen of the Trichoderma asperellum/asperelloides taxon participating in the lateral transfer of an Aspergillus-born aox gene (see Results and Discussion section). The rare aoxB gene found intact in one clade of the A. niger sensu stricto complex (typified by strains ATCC 1015 and CBS 147482), as well as in A. calidoustus, A. implicatus and Penicillium swiecickii, was previously identified as part of a gene couple with an equally rare type-2 NADH dehydrogenase paralogue gene (andB), unique to these four divergent taxa [13]. In the gene nomenclature adopted to describe the multiple aox gene duplication events in the lineage of the Aspergillaceae, this previously described paralogue is dubbed "aoxB1".…”

“…In addition, Aspergillus calidoustus-all six genomes in the databases-trapped a rare copy of the aoxB1 duplication event (cf. [13]). In A. calidoustus, both paralogous aoxB genes are thus effectively "replaced" by two other aoxB paralogues originating from different duplication events.…”

“…Furthermore, sequence-related but genetically unlinked genes called aod-1 and aod-3 have been described in Neurospora crassa [12]. We recently described a third instance, where a rare second aox paralogue named aoxB (the primary enzyme AoxA encoded by the ubiquitous aoxA gene) was found scattered in four species of Aspergillaceae amongst hundreds of genome-sequenced member species [13]. Importantly, in the Aspergillus niger sensu stricto complex-where more than 80 genome sequences are available for comparison-we identified five mutations in this acquired aoxB gene that could be used to distinguish six taxa in this species complex known to be notoriously difficult to subdivide [13].…”