Plastomes from tribe Plantagineae (Plantaginaceae) reveal infrageneric structural synapormorphies and localized hypermutation for Plantago and functional loss of ndh genes from Littorella.

Tribe Plantagineae (Plantaginaceae) comprises ~ 270 species in three currently recognized genera (Aragoa, Littorella, Plantago), of which Plantago is most speciose. Plantago plastomes exhibit several atypical features including large inversions, expansions of the inverted repeat, increased repetitiveness, intron losses, and gene-specific increases in substitution rate, but the prevalence of these plastid features among species and subgenera is unknown. To assess phylogenetic relationships and plastomic evolutionary dynamics among Plantagineae genera and Plantago subgenera, we generated 25 complete plastome sequences and compared them with existing plastome sequences from Plantaginaceae. Using whole plastome and partitioned alignments, our phylogenomic analyses provided strong support for relationships among major Plantagineae lineages. General plastid features-including size, GC content, intron content, and indels-provided additional support that reinforced major Plantagineae subdivisions. Plastomes from Plantago subgenera Plantago and Coronopus have synapomorphic expansions and inversions affecting the size and gene order of the inverted repeats, and particular genes near the inversion breakpoints exhibit accelerated nucleotide substitution rates, suggesting localized hypermutation associated with rearrangements. The Littorella plastome lacks functional copies of ndh genes, which may be related to an amphibious lifestyle and partial reliance on CAM photosynthesis.

Symbiosis of isoetid plant species with arbuscular mycorrhizal fungi under aquatic versus terrestrial conditions.

Arbuscular mycorrhizal fungi (AMF) colonize the roots of numerous aquatic and wetland plants, but the establishment and functioning of mycorrhizal symbiosis in submerged habitats have received only little attention. Three pot experiments were conducted to study the interaction of isoetid plants with native AMF. In the first experiment, arbuscular mycorrhizal (AM) symbiosis did not establish in roots of Isoëtes echinospora and I. lacustris, while Littorella uniflora roots were highly colonized. Shoot and root biomass of L. uniflora were, however, not affected by AMF inoculation, and only one of nine AMF isolates significantly increased shoot P concentration. In the second experiment, we compared colonization by three Glomus tetrastratosum isolates of different cultivation history and origin (aquatic versus terrestrial) and their effects on L. uniflora growth and phosphorus nutrition under submerged versus terrestrial conditions. The submerged cultivation considerably slowed, but did not inhibit mycorrhizal root colonization, regardless of isolate identity. Inoculation with any AMF isolate improved plant growth and P uptake under terrestrial, but not submerged conditions. In the final experiment, we compared the communities of AMF established in two cultivation regimes of trap cultures with lake sediments, either submerged on L. uniflora or terrestrial on Zea mays. After 2-year cultivation, we did not detect a significant effect of cultivation regime on AMF community composition. In summary, although submerged conditions do not preclude the development of functional AM symbiosis, the contribution of these symbiotic fungi to the fitness of their hosts seems to be considerably less than under terrestrial conditions.

Molecular and Morphological Data Improve the Classification of Plantagineae (Lamiales).

The tribe Plantagineae (Lamiales) is a group of plants with worldwide distribution, notorious for its complicated taxonomy and still unresolved natural history. We describe the result of a broadly sampled phylogenetic study of tribe. The expanded sampling dataset is based on the trnL-F spacer, rbcL, and ITS2 markers across all three included genera (Aragoa, Littorella and Plantago) and makes this the most comprehensive study to date. The other dataset uses five markers and provides remarkably good resolution throughout the tree, including support for all of the major clades. In addition to the molecular phylogeny, a morphology database of 114 binary characters was assembled to provide comparison with the molecular phylogeny and to develop a means to assign species not sampled in the molecular analysis to their most closely related species that were sampled. Based on the molecular phylogeny and the assignment algorithm to place unsampled species, a key to sections is presented, and a revised classification of the tribe is provided. We also include the description of new species from North America.

The impact of NO inf3sup- loading on the freshwater macrophyte Littorella uniflora: N utilization strategy in a slow-growing species from oligotrophic habitats.

The decline and disappearance of Littorella uniflora from oligotrophic waters which have become eutrophic has been associated with shading or reduced CO2 supply. However NO inf3sup- concentrations can reach very high levels (100-2000 mmol m-3 compared with <1-3 in oligotrophic habitats). To investigate the impact of NO inf3sup- loading alone, plants were grown under three NO inf3sup- regimes (very low, near-natural and high). The interactive effects of NO inf3sup- and photon flux density (low and high regimes) on N assimilation and accumulation, CO2 concentrating mechanisms, C3 photosynthesis and growth were also examined. The results were unexpected. Increased NO inf3sup- supply had very little effect on photosynthetic capacity, crassulacean acid metabolism (CAM) or lacunal CO2 concentrations ([CO2]i), although there was considerable plasticity with respect to light regime. In contrast, increased NO inf3sup- supply resulted in a marked accumulation of NO inf3sup- , free amino acids and soluble protein in shoots and roots (up to 25 mol m-3, 30 mol m-3 and 9 mg g-1 fresh weight respectively in roots), while fresh weight and relative growth rate were reduced. Total N content even under the very low NO inf3sup- regime (1.6-2.3%) was mid-range for aquatic and terrestrial species (and 3.1-4.3% under the high NO inf3sup- regime). These findings, together with field data, suggest that L. uniflora is not growth limited by low NO inf3sup- supply in natural oligotophic habitats, due not to an efficient photosynthetic nitrogen use but to a slow growth rate, a low N requirement and to the use of storage to avoid N stress. However the increased NO inf3sup- concentrations in eutrophic environments seem likely have detrimental effects on the long-term survival of L. uniflora, possibly as a consequence of N accumulation.

Land plants of amphibious Littorella uniflora (L.) Aschers. maintain utilization of CO2 from the sediment.

Submerged macrophytes of the isoetid life form derive the majority of their CO2 for photosynthesis from the sediment. The experiments described here were designed to test the hypothesis that root uptake of CO2 is important also in the terrestrial form of Littorella uniflora. The results of 14CO2 experiments showed that sediment CO2 contributed 56% of the total fixation at 0.1MM CO2 in the rhizosphere, 83% at 0.5MM and 96% at 2.5MM. Sediment CO2 in emergent Littorella stands ranged from 0.1 to 1.0MM and averaged 0.5MM. Measurements of the net CO2 exchange over the leaves showed an even higher dependence of the sediment as CO2 source. Littorella leaves had no stomata at the base and densities (ca. 100 mm-2) typical of terrestrial plants at the tip, allowing sediment-derived CO2 to be supplied along the length of the leaf. The stomata permit supply of CO2 from the air during periods of reduced sediment CO2 concentrations (e.g. if the sediment dries up) and regulate transpiration.

Photosynthesis of Littorella uniflora grown under two PAR regimes: C3 and CAM gas exchange and the regulation of internal CO2 and O2 concentrations.

The submersed aquatic macrophyte Littorella uniflora was grown under 50 and 300 µmol m-2 s-1 photosynthetically active radiation (PAR) (low and high PAR regimes) but identical sediment CO2 supply (1.0 mol m-3). The interactions between plant morphology, whole plant CO2 and O2 exchange, CAM activity, [CO2] i and [O2] i have been investigated in comparison with in vitro CO2 and PAR response characteristics (using 1 mm leaf sections). In terms of morphology, high-PAR-grown plants were smaller and leaves contained less chlorophyll, although root growth was proportionally larger. Gas exchange fluxes over roots and shoots of intact plants were similar in direction under the two PAR regimes, with the majority of CO2 uptake via the roots. Photosynthetic O2 evolution from intact plants was greater in high-PAR-grown L. uniflora (2.18 compared with 1.49 µmol O2g-1 fresh weight h-1 for the low PAR regime). Although net daytime CO2 uptake was similar for both PAR regimes (0.79 and 0.75 µmol g-1 fwt h-1), net dark CO2 uptake was at a higher rate (0.92 compared with 0.52 µmol CO2 g-1 fwt h-1), and dark fixation (as malic acid) was threefold greater in high PAR plants (ΔH+ 117 compared with 42 µmol H+ g-1 fwt). Comparison of dark CO2 uptake with dark fixation suggested that much of the CO2 fixed at night and regenerated during the day may be respiratory in origin (60% low PAR plants, 71% high PAR plants). Regeneration of CO2 from CAM could account for 62% of daytime CO2 supply in low PAR plants and 81% in high PAR plants. [CO2] i values (ranging from 0.42 to 1.03 mol m-3) were close to or above the concentration required to saturate photosynthesis in vitro (0.5 mol m-3) under both PAR regimes, and combined with the low [O2] i (2.6-4.3 mol m-3) should have suppressed photorespiration. However, PAR inside leaves would have been well below the in vitro light saturation requirement (850-1000 µmol m-2 s-1 for both treatments). Thus PAR rather than CO2 supply appeared to limit photosynthesis even in high PAR grown plants, and CAM appears to have an important role in the regulation of CO2 supply for photosynthesis in response to variation in light regime.