# An Ammonium Transporter Gene Contributes to the Aggressiveness of the Dutch Elm Disease Pathogen Ophiostoma novo-ulmi

## TL;DR

A gene encoding an ammonium transporter in a fungus that causes Dutch elm disease is found to be important for its ability to aggressively infect elm trees.

## Contribution

The study identifies and characterizes an ammonium transporter gene (amtA) as a key factor in the aggressiveness of the Dutch elm disease pathogen Ophiostoma novo-ulmi.

## Key findings

- A knockdown mutant of the amtA gene showed significantly reduced aggressiveness in infecting elm saplings.
- The amtA gene product has features similar to Mep2-type transporters, including conserved amino acid residues and transmembrane helices.
- CRISPR-Cas9 knockout mutants of amtA retained some wild-type traits but were less aggressive towards elm trees.

## Abstract

Molecular mechanisms determining pathogenicity of the Dutch elm disease fungus Ophiostoma novo-ulmi are poorly understood. Prior identification of the pathogenicity locus pat1 prompted a chromosome walking approach to elucidate gene function in this region. Among 17 identified genes, ONUg0282 (amtA) was predicted to encode a high-affinity ammonium transporter. In silico analyses confirmed the presence of four additional amt genes (amtB, amtC, amtD, and amtE) in both O. novo-ulmi and the less aggressive O. ulmi and that amtA and amtB belong to the Saccharomyces cerevisiae mep2 clade. The predicted amtA gene product showed features of Mep2-type transceptors, including amino acid residues corresponding to His-168 and His-318 in Escherichia coli AmtB protein, 11 transmembrane helices, and a conserved 22 amino acid motif immediately downstream of the last transmembrane helix. A knockdown amtA mutant with 25% residual expression was significantly less aggressive than wild-type O. novo-ulmi strain H327 when infecting Ulmus americana × U. parvifolia saplings. Predicted AmtA transporters from two CRISPR-Cas9 knockout mutants contained only five intact transmembrane helices. The ΔamtA mutants retained several wild-type phenotypic traits, including yeast–mycelium dimorphism, but were significantly less aggressive than H327 towards U. americana saplings. We concluded that ONUg0282 is an important determinant of aggressiveness in O. novo-ulmi.