Intra and Inter-Spore Variability in Rhizophagus irregularis AOX Gene

Arbuscular mycorrhizal fungi (AMF) are root-inhabiting fungi that form mutualistic symbioses
with their host plants. AMF symbiosis improves nutrient uptake and buffers the plant
against a diversity of stresses. Rhizophagus irregularis is one of the most widespread AMF
species in the world, and its application in agricultural systems for yield improvement has
increased over the last years. Still, from the inoculum production perspective, a lack of
consistency of inoculum quality is referred to, which partially may be due to a high genetic
variability of the fungus. The alternative oxidase (AOX) is an enzyme of the alternative respiratory
chain already described in different taxa, including various fungi, which decreases
the damage caused by oxidative stress. Nevertheless, virtually nothing is known on the
involvement of AMF AOX on symbiosis establishment, as well on the existence of AOX variability
that could affect AMF effectiveness and consequently plant performance. Here, we
report the isolation and characterisation of the AOX gene of R. irregularis (RiAOX), and
show that it is highly expressed during early phases of the symbiosis with plant roots. Phylogenetic
analysis clustered RiAOX sequence with ancient fungi, and multiple sequence alignment
revealed the lack of several regulatory motifs which are present in plant AOX. The
analysis of RiAOX polymorphisms in single spores of three different isolates showed a
reduced variability in one spore relatively to a group of spores. A high number of polymorphisms
occurred in introns; nevertheless, some putative amino acid changes resulting from
non-synonymous variants were found, offering a basis for selective pressure to occur within
the populations. Given the AOX relatedness with stress responses, differences in gene variants
amongst R. irregularis isolates are likely to be related with its origin and environmental
constraints and might have a potential impact on inoculum production.