Rhizobial bacteria nodulate legume roots and fix nitrogen in exchange for photosynthates. These symbionts are infectiously acquired from the environment and in such cases selection models predict evolutionary spread of uncooperative mutants. Uncooperative rhizobia – including nonfixing and non‐nodulating strains – appear common in agriculture, yet their population biology and origins remain unknown in natural soils. Here, a phylogenetically broad sample of 62 wild‐collected rhizobial isolates was experimentally inoculated onto Lotus strigosus to assess their nodulation ability and effects on host growth. A cheater strain was discovered that proliferated in host tissue while offering no benefit; its fitness was superior to that of beneficial strains. Phylogenetic reconstruction of Bradyrhizobium rDNA and transmissible symbiosis‐island loci suggest that the cheater evolved via symbiotic gene transfer. Many strains were also identified that failed to nodulate L. strigosus, and it appears that nodulation ability on this host has been recurrently lost in the symbiont population. This is the first study to reveal the adaptive nature of rhizobial cheating and to trace the evolutionary origins of uncooperative rhizobial mutants.

Summary• Arbuscular mycorrhizal fungi (AMF) form symbioses with the majority of plants and form extensive underground hyphal networks simultaneously connecting the roots of different plant species. No empirical evidence exists for either anastomosis between genetically different AMF or genetic exchange.• Five isolates of one population of Glomus intraradices were used to study anastomosis between hyphae of germinating spores. We show that genetically distinct AMF, from the same field, anastomose, resulting in viable cytoplasmic connections through which genetic exchange could potentially occur.• Pairs of genetically different isolates were then co-cultured in an in vitro system. Freshly produced spores were individually germinated to establish new cultures. Using several molecular tools, we show that genetic exchange occurred between genetically different AMF. Specific genetic markers from each parent were transmitted to the progeny. The progeny were viable, forming symbioses with plant roots. The phenotypes of some of the progeny were significantly different from either parent.• Our results indicate that considerable promiscuity could occur in these fungi because nine out of 10 combinations of different isolates anastomosed. The ability to perform genetic crosses between AMF experimentally lays a foundation for understanding the genetics and evolutionary biology of these important plant symbionts.

Summary Arbuscular mycorrhizal fungi (AMF) are highly successful plant symbionts. They reproduce clonally producing multinucleate spores. It has been suggested that some AMF harbor genetically different nuclei. However, recent advances in sequencing the Glomus irregulare genome have indicated very low within‐fungus polymorphism. We tested the null hypothesis that, with no genetic differences among nuclei, no significant genetic or phenotypic variation would occur among clonal single spore lines generated from one initial AMF spore. Furthermore, no additional variation would be expected in the following generations of single spore lines. Genetic diversity contained in one initial spore repeatedly gave rise to genetically different variants of the fungus with novel phenotypes. The genetic changes represented quantitative changes in allele frequencies, most probably as a result of changes in the frequency of genetic variation partitioned on different nuclei. The genetic and phenotypic variation is remarkable, given that it arose repeatedly from one clonal individual. Our results highlight the dynamic nature of AMF genetics. Even though within‐fungus genetic variation is low, some is probably partitioned among nuclei and potentially causes changes in the phenotype. Our results are important for understanding AMF genetics, as well as for researchers and biotechnologists hoping to use AMF genetic diversity for the improvement of AMF inoculum.

Gene copy number polymorphism was studied in a population of the arbuscular mycorrhizal fungus Glomus intraradices by using a quantitative PCR approach on four different genomic regions. Variation in gene copy number was found for a pseudogene and for three ribosomal genes, providing conclusive evidence for a widespread occurrence of macromutational events in the population.