Carbon for nutrient exchange between Lycopodiella inundata and Mucoromycotina fine root endophytes is unresponsive to high atmospheric CO2.

Non-vascular plants associating with arbuscular mycorrhizal (AMF) and Mucoromycotina 'fine root endophyte' (MFRE) fungi derive greater benefits from their fungal associates under higher atmospheric [CO2] (a[CO2]) than ambient; however, nothing is known about how changes in a[CO2] affect MFRE function in vascular plants. We measured movement of phosphorus (P), nitrogen (N) and carbon (C) between the lycophyte Lycopodiella inundata and Mucoromycotina fine root endophyte fungi using 33P-orthophosphate, 15 N-ammonium chloride and 14CO2 isotope tracers under ambient and elevated a[CO2] concentrations of 440 and 800 ppm, respectively. Transfers of 33P and 15 N from MFRE to plants were unaffected by changes in a[CO2]. There was a slight increase in C transfer from plants to MFRE under elevated a[CO2]. Our results demonstrate that the exchange of C-for-nutrients between a vascular plant and Mucoromycotina FRE is largely unaffected by changes in a[CO2]. Unravelling the role of MFRE in host plant nutrition and potential C-for-N trade changes between symbionts under different abiotic conditions is imperative to further our understanding of the past, present and future roles of plant-fungal symbioses in ecosystems.

Mucoromycotina Fine Root Endophyte Fungi Form Nutritional Mutualisms with Vascular Plants.

Fungi and plants have engaged in intimate symbioses that are globally widespread and have driven terrestrial biogeochemical processes since plant terrestrialization >500 million years ago. Recently, hitherto unknown nutritional mutualisms involving ancient lineages of fungi and nonvascular plants have been discovered, although their extent and functional significance in vascular plants remain uncertain. Here, we provide evidence of carbon-for-nitrogen exchange between an early-diverging vascular plant (Lycopodiella inundata) and Mucoromycotina (Endogonales) fine root endophyte fungi. Furthermore, we demonstrate that the same fungal symbionts colonize neighboring nonvascular and flowering plants. These findings fundamentally change our understanding of the physiology, interrelationships, and ecology of underground plant-fungal symbioses in modern terrestrial ecosystems by revealing the nutritional role of Mucoromycotina fungal symbionts in vascular plants.

Prevalence and phenology of fine root endophyte colonization across populations of Lycopodiella inundata.

Mycorrhizal fungi are critical components of terrestrial habitats and agroecosystems. Recently, Mucoromycotina fine root endophyte fungi (MucFRE) were found to engage in nutritional mutualism with Lycopodiella inundata, which belongs to one of the earliest vascular plant lineages known to associate with MucFRE. The extent to which this mutualism plays a role in resilient plant populations can only be understood by examining its occurrence rate and phenological patterns. To test for prevalence and seasonality in colonization, we examined 1305 individual L. inundata roots from 275 plants collected during spring and autumn 2019 across 11 semi-natural heathlands in Britain and the Netherlands. We quantified presence/absence of fine root endophyte (FRE) hyphae and vesicles and explored possible relationships between temperature and precipitation in the months immediately before sampling. Fine root endophyte hyphae were dominant in all of the examined heathlands, and every colonized root had FRE in both cortical cells and root hairs. However, we found significant differences in colonization between the two seasons at every site. Overall, 14% of L. inundata roots were colonized in spring (2.4% with vesicles) compared with 86% in autumn (7.6% with vesicles). Colonization levels between populations were also significantly different, correlating with temperature and precipitation, suggesting some local environments may be more conducive to root and related hyphal growth. These marked seasonal differences in host-plant colonization suggest that results about FRE from single time point collections should be carefully interpreted. Our findings are relevant to habitat restoration, species conservation plans, agricultural bio-inoculation treatments, and microbial diversity studies.

From rhizoids to roots? Experimental evidence of mutualism between liverworts and ascomycete fungi.

Background and Aims: The rhizoids of leafy liverworts (Jungermanniales, Marchantiophyta) are commonly colonized by the ascomycete fungus Pezoloma ericae. These associations are hypothesized to be functionally analogous to the ericoid mycorrhizas (ErMs) formed by P. ericae with the roots of Ericaceae plants in terms of bi-directional phosphorus for carbon exchange; however, this remains unproven. Here, we test whether associations between the leafy liverwort Cephalozia bicuspidata and P. ericae are mutualistic. Methods: We measured movement of phosphorus and carbon between C. bicuspidata and P. ericae using [33P]orthophosphate and 14CO2 isotope tracers in monoxenic cultures. We also measured leafy liverwort growth, with and without P. ericae. Key Results: We present the first demonstration of nutritionally mutualistic symbiosis between a non-vascular plant and an ErM-forming fungus, showing transfer of fungal-acquired P to the liverwort and of liverwort-fixed C to the fungus alongside increased growth in fungus-colonized liverworts. Conclusions: Thus, this ascomycete-liverwort symbiosis can now be described as mycorrhiza-like, providing further insights into ericoid mycorrhizal evolution and adding Ascomycota fungi to mycorrhizal fungal groups engaging in mutualisms with plants across the land plant phylogeny. As P. ericae also colonizes the rhizoids of Schistochilaceae liverworts, which originated in the Triassic and are sister to all other jungermannialean liverworts associated with fungi, our findings point toward an early origin of ascomycete-liverwort symbioses, possibly pre-dating their evolution in the Ericales by some 150 million years.

The biology of Marchantia polymorpha subsp. ruderalis Bischl. & Boissel. Dub in nature.

Introduction: Though used as the model liverwort in culture for several decades, the biology of Marchantia polymorpha subsp. ruderalis in nature has never been documented in detail in a single account. Methods: Here we synthesize routine field observations documented with hundreds of images of M. ruderalis colonies (or groups) showing sex differentiation over 3 years on two populations of M. ruderalis after major heathland fires in 2020. Results: Initial post-fire establishment is from airborne spores rather than a spore bank but thereafter spread is via gemmae which have less exacting germination requirements. Young sporelings are highly gemmiferous but gemmae production becomes less frequent after sex organ formation. Over the course of a year there are up to three waves of carpocephalum production with the overwhelming majority of antheridiophores appearing 2-3 months ahead of the archegoniophores though no differences in growth rates were apparent between male and female thalli. Spermatozoids are produced almost continuously throughout the year, whilst sporophyte maturation is restricted to the summer months. Discussion: Because of the asynchrony between antheridiophore and archegoniophore production a 1:1 sex ratio is only apparent over this period. The spring months see an excess of males with more females in the summer. An almost 100% fertilization rate, with fertilization distances of up to 19 m far exceeding those in all other bryophytes, is attributed to vast spermatozoid production for most of the year, dispersal on surface oil films between thalli and highly effective intra-thallus spermatozoid transport via the pegged-rhizoid water-conducting system. Archegoniophores do develop on female-only populations but have shorter stalks than those where fertilization has occurred. Eventual disappearance post fires is attributed to a fall in topsoil nutrient levels preventing new sporeling establishment and competition from Ceratodon purpureus and Polytrichum spp. A major drought in the summer of 2022 almost wiped out the heathland Marchantia populations but all the other bryophytes survived.

Acidified Blue Ink-staining Procedure for the Observation of Fungal Structures Inside Roots of Two Disparate Plant Lineages.

Identifying microscopic mycorrhizal fungal structures in roots, i.e., hyphae, vesicles and arbuscules, requires root staining procedures that are often time consuming and involves chemicals known to present health risks from exposure. By modifying established protocols, our root staining method stains roots using a safe ink- and vinegar-based staining solution, followed by a 2-16 h-long de-staining period. The entire procedure can be completed in less than 6 h (plus up to 16 h de-staining overnight) and roots are suitable for semi-permanent and permanent slide mounting for light microscopy. We tested our method on hundreds of wild-sourced roots from two different plant species: Lycopodiella inundata, a herbaceous clubmoss with tough water-resistant roots, and Sambucus nigra, a temperate woody shrub. Both plants associate with endomycorrhizae, L. inundata predominantly with Mucoromycotina fine root endophytes (MucFRE) and S. nigra with Glomeromycota arbuscular mycorrhizal fungi (AMF). Here we describe a simple, efficient, repeatable and safe method to detect the presence of fungal structures using light microscopy.

A mycorrhizal revolution.

It has long been postulated that symbiotic fungi facilitated plant migrations onto land through enhancing the scavenging of mineral nutrients and exchanging these for photosynthetically fixed organic carbon. Today, land plant-fungal symbioses are both widespread and diverse. Recent discoveries show that a variety of potential fungal associates were likely available to the earliest land plants, and that these early partnerships were probably affected by changing atmospheric CO2 concentrations. Here, we evaluate current hypotheses and knowledge gaps regarding early plant-fungal partnerships in the context of newly discovered fungal mutualists of early and more recently evolved land plants and the rapidly changing views on the roles of plant-fungal symbioses in the evolution and ecology of the terrestrial biosphere.