Adaptive horizontal transfer of a bacterial gene to an invasive insect pest of coffee

Abstract

Horizontal gene transfer (HGT) involves the nonsexual transmission of genetic material across species boundaries. Although often detected in prokaryotes, examples of HGT involving animals are relatively rare, and any evolutionary advantage conferred to the recipient is typically obscure. We identified a gene (HhMAN1) from the coffee berry borer beetle, Hypothenemus hampei, a devastating pest of coffee, which shows clear evidence of HGT from bacteria. HhMAN1 encodes a mannanase, representing a class of glycosyl hydrolases that has not previously been reported in insects. Recombinant HhMAN1 protein hydrolyzes coffee berry galactomannan, the major storage polysaccharide in this species and the presumed food of H. hampei. HhMAN1 was found to be widespread in a broad biogeographic survey of H. hampei accessions, indicating that the HGT event occurred before radiation of the insect from West Africa to Asia and South America. However, the gene was not detected in the closely related species H. obscurus (the tropical nut borer or "false berry borer"), which does not colonize coffee beans. Thus, HGT of HhMAN1 from bacteria represents a likely adaptation to a specific ecological niche and may have been promoted by intensive agricultural practices.

Fig. 1.

The mannanase gene HhMAN1 from the coffee berry borer (Hypothenemus hampei). (A) An adult H. hampei burrows into and lays its eggs in coffee berries (B). (C) Best maximum likelihood (ML) phylogenetic analysis of HhMAN1 gene (red) with homologs from fungi (blue), bacteria (black), animals (purple), and plants (green). GenBank accession numbers are shown adjacent to each gene and bootstrap values are indicated on key branches. Where relevant, bootstrap values are notated with maximum likelihood followed by maximum parsimony (MP). Nodes with only one bootstrap value are from ML analyses only and were not supported in the MP bootstrap analysis.

Fig. 2.

HhMAN1 is integrated into the H. hampei genome. (A) Structure of the HhMAN1 gene highlighting the ORF, as well as flanking untranslated regions (gray) and predicted adjacent transposase-associated sequences. Positions (bases) within the genomic fragment (GenBank GQ375156) are indicated. (B) PCR amplification of HhMAN1 from DNA isolated from various H. hampei body parts. A, whole adult; C+, positive control (HhMAN1 cDNA plasmid); C−, negative control (empty plasmid); E, elytra; L, legs; MG, midgut of second-stage larvae. The arrow indicates the diagnostic 1,007-bp HhMAN1 PCR product. (C) PCR amplification of HhMAN1 from DNA isolated from H. hampei, H. obscurus, and A. fasciculatus. Molecular mass markers (M) in base pairs are indicated.

Fig. 3.

Detection of HhMAN1 in genomic DNA from a geographically broad range of H. hampei accessions. The arrows indicate the diagnostic amplified band. A positive control (C+, HhMAN1 cDNA plasmid) and a negative control (C−, empty vector) are shown. Molecular mass markers in base pairs (bp) are indicated.

Fig. 4.

Mannanase activity from extracts of the midgut of H. hampei, H. obscurus, and A. fasciculatus and recombinant HhMAN1. (A) Hydrolytic activity of midgut extracts from H. hampei, H. obscurus, and A. fasciculatus acting on coffee galactomannan substrate. (B) SDS/PAGE of recombinant HhMAN1-His₆ purified from transfected insect cells by nickel affinity chromatography. Molecular mas standards are indicated in kilodaltons. (C) Hydrolytic activity of purified HhMAN1-His₆ on coffee galactomannan substrate. Error bars are SD for n = 3.