What prevent plants from becoming heterotrophic on their mycorrhizal fungi: Insight from the study of achlorophyllous variants in green orchid species

Background/Question/Methods Among green plants, non photosynthetic, achlorophyllous variants (AV) do not survive in the wild, except in few orchid species where they even produce fruits. These orchid species are phylogenetically related to fully heterotrophic orchids that use their mycorrhizal fungi as unique carbon source (mycoheterotrophy). The content in stable isotopes of AV is identical to that of mycoheterotrophyic orchids, i.e. enriched in 13C and 15N  (exactly as their mycorrhizal fungi) as compared to fully autotrophic plants. A similar, but lower, trend exists in the green individual of these orchid species, suggesting that (i) green individual in part relies on carbon from their mycorrhizal fungi and (ii) AV solely use this carbon source. Although both green individuals and AV look morphologically similar, the frequency of AV in populations is barely higher than 5 %. This raises the question of what impairs AV fitness. We studied the dynamics, reproduction and physiology of AV an dgreen individuals in the orchid Cephalanthera damasonium for several populations. We monitored over a growing season for both phenotypes (i) the isotopic content, revealing the level of use of fungal carbon, (ii) gas exchange physiology (CO2, H2O), together with (iii) comparative analysis of phenology and morphology.

            Results/Conclusions First, over three years, AV frequency was stable, and AVs did not invade. Like green individuals, half of AVs were able to stay underground during one or two years. Their phenology was synchronous for shoots growth, flowering and fruiting. No significant difference in growth rate, morphology, pollination rate, or germination rate was detected. Nevertheless, some differences occurred at fruit maturation: AVs more often exhibited an early shoot drying, leading to fruit abortion. Isotopic content showed that green individuals use fungal carbon all over the whole vegetative season, including during fruiting, so that a decrease of the flow of fungal carbon is unlikely to explain AV drying. Rather, AVs transpire five times less than green individuals, entailing overheating of leaves in summer. Since stomata did not respond well to CO2 and light, we suggest that non-functional plastids in stomatal cells entail dysfunction of stomatal exchanges. This underlines the cost of the persistence of otherwise useless leaves in AV. Interestingly, mycoheterotrophic orchids are leafless. At rise of full mycoheterotrophy, previously green plants are limited by their previous adaptations to photosynthesis (leaves, stomata). This probably contributes to limit the emergence of mycoheterotrophic plants.