Colonization Dynamics and Diversity of Holomycota in a Newly Built Urban Wastewater Treatment Plant: An Eight-Month Time-Series Analysis

High-throughput sequencing of eukaryotic 18S rDNA genes was conducted on 23 time-series samples collected over eight months from a newly constructed urban Seine-Morée-wastewater treatment plant (WWTP), generating 521,031 reads. Among these reads, 74,432 (14.2%) are affiliated with Holomycota , spanning nine phylogenetic groups. Cryptomycota dominated the fungal community (76.01%), followed by Chytridiomycota (11.66%), Ascomycota (5.65%), Zoopagomycota (3.02%), Basidiomycota (1.37%), and Blastocladiomycota (1.36%), while Mucoromycota , Nucleariidae / Fonticula , and Microsporidia each represented < 1% of total Holomycota reads. The fungal community was highly uneven, with 50 abundant operational taxonomic units (OTUs) (≥ 0.1% relative abundance) constituting 96.5% of all sequences, while the vast majority of taxa (298 OTUs) were rare (< 0.1%), forming a "rare biosphere" that contributed only by 3.5% of the total reads. Among the 50 abundant taxa, only fifteen highly dominant operational taxonomic units (OTUs), each with a relative abundance ≥ 1%, overwhelmingly structured the fungal community, and accounted for 85% of the total fungal reads. The cultivable fraction (82 OTUs) made up just 8.5% of fungal reads, with Synchytrium cupulatum (6%) emerging as the most prevalent species. Additionally, nineteen species affiliated with the human gut mycobiota (e.g., Geotrichum, Candida, Saccharomyces, Penicillium ) accounted for 3.62% of fungal reads. Among the 348 identified OTUs, at least 32 are affiliated with taxa exhibiting potential pathogenicity toward humans, plants, or animals, highlighting the importance of monitoring fungal communities within wastewater ecosystems under the One-Global-Health framework. From an ecological perspective, early-stage fungal colonization may contribute to the stabilization and structuring of microbial communities within the WWTP. Hence, these communities contribute to ecosystem construction, stabilization, and functioning. Our findings underscore the predominant role of fungal “dark matter” (e.g., Cryptomycota ) and reveal the substantial potential for novel taxonomic and functional discoveries within the wastewater mycobiome.