The genome of the coffee bean weevil (Araecerus fasciculatus) reveals a cytochrome P450 repertoire as a convergent candidate mechanism of insect-origin caffeine detoxification
The coffee bean weevil, Araecerus fasciculatus (Coleoptera, Curculionoidea, Anthribidae), is a cosmopolitan pest of over 100 stored agricultural commodities, with particular economic impact on coffee (Coffea arabica). Although two chromosome-level anthribid genomes have recently been released as part of the Darwin Tree of Life (DToL) project (Booth et al. 2024; Crowley et al. 2025), no functionally annotated genome has been available for the family. Here we present a draft genome assembly for A. fasciculatus, generated from PacBio HiFi long reads and processed through a three tiered metagenomic filtering pipeline to remove host plant (C. arabica) and microbial contamination. The final assembly spans 475 Mb across 3,617 scaffolds (N50 = 170 kb) with 88.5% BUSCO completeness (insecta_odb10) and only 3.1% duplication. Gene prediction with BRAKER2 identified 22,384 protein-coding genes, of which 11,783 received functional annotations through SwissProt similarity. Notably, we identified 92 cytochrome P450 (CYP) genes, including tandem gene clusters on two scaffolds (4 genes on ptg000464l, 5 genes on ptg001867l), suggestive of lineage-specific expansion through tandem duplication. Homology searches against Drosophila melanogaster caffeine-metabolizing P450s (CYP12D1, CYP6d5, CYP6a8) recovered strong matches (e-values 9.7 x 10-110 to 5.4 x 10-101, 33-38% identity). In stark contrast, comprehensive BLAST searches for bacterial caffeine N-demethylase genes (ndmA/B/C/D), which mediate caffeine degradation via horizontal gene transfer in the coffee berry borer Hypothenemus hampei (Scolytinae), returned zero hits across the A. fasciculatus genome, predicted proteome, and associated bacterial scaffolds. AlphaFold2 structure prediction of four top Araecerus P450 candidates produced high-confidence models (pLDDT 84.5-93.9, pTM 0.735-0.930) with conserved P450 catalytic motifs. Foldseek structural homology searches confirmed that all four candidates adopt cytochrome P450 folds (top hits: human CYP3A4, CYP3A7, CYP11A1; TM-scores 0.90-0.92; probability 1.000), with zero hits to bacterial Rieske-fold enzymes. Molecular docking of caffeine against these structures yielded binding affinities of -5.41 to -5.80 kcal/mol for the Araecerus candidates, comparable to or exceeding the -5.55 kcal/mol obtained for the experimentally validated Drosophila CYP6a8 and substantially stronger than the -3.70 kcal/mol for the bacterial NdmA structural outgroup (PDB: 6ICP). Phylogenetic analysis revealed that all four candidates have clear orthologs in two non-seed-feeding DToL anthribids (Pseudeuparius sepicola and Platystomos albinus), demonstrating that these P450 genes predate the dietary transition to caffeine-containing seeds. The Araecerus candidates predominantly belong to the CYP6 family (clan 3), whereas the primary Drosophila caffeine P450 CYP12D1 belongs to the mitochondrial clan, confirming convergent recruitment of different P450 subfamilies for caffeine metabolism. These results support the hypothesis that A. fasciculatus employs an insect-encoded, P450-mediated caffeine detoxification pathway fundamentally distinct from the bacterial horizontal gene transfer mechanism documented in Scolytinae. This represents convergent evolution of caffeine resistance via independent molecular strategies within Curculionoidea, and provides the first functionally annotated genomic resource for comparative studies across the Anthribidae.