First evidence of mutualism between ancient plant lineages ( H aplomitriopsida liverworts) and M ucoromycotina fungi and its response to simulated P alaeozoic changes in atmospheric CO 2

Summary The discovery that M ucoromycotina, an ancient and partially saprotrophic fungal lineage, associates with the basal liverwort lineage H aplomitriopsida casts doubt on the widely held view that G lomeromycota formed the sole ancestral plant–fungus symbiosis. Whether this association is mutualistic, and how its functioning was affected by the fall in atmospheric CO 2 concentration that followed plant terrestrialization in the P alaeozoic, remains unknown. We measured carbon‐for‐nutrient exchanges between H aplomitriopsida liverworts and M ucoromycotina fungi under simulated mid‐ P alaeozoic (1500 ppm) and near‐contemporary (440 ppm) CO 2 concentrations using isotope tracers, and analysed cytological differences in plant–fungal interactions. Concomitantly, we cultured both partners axenically, resynthesized the associations in vitro , and characterized their cytology. We demonstrate that liverwort– M ucoromycotina symbiosis is mutualistic and mycorrhiza‐like, but differs from liverwort– G lomeromycota symbiosis in maintaining functional efficiency of carbon‐for‐nutrient exchange between partners across CO 2 concentrations. Inoculation of axenic plants with M ucoromycotina caused major cytological changes affecting the anatomy of plant tissues, similar to that observed in wild‐collected plants colonized by M ucoromycotina fungi. By demonstrating reciprocal exchange of carbon for nutrients between partners, our results provide support for M ucoromycotina establishing the earliest mutualistic symbiosis with land plants. As symbiotic functional efficiency was not compromised by reduced CO 2 , we suggest that other factors led to the modern predominance of the G lomeromycota symbiosis.