Summary• Experimental evidence demonstrates a higher efficiency of water and nitrogen use in C 4 compared with C 3 plants, which is hypothesized to drive differences in biomass allocation between C 3 and C 4 species. However, recent work shows that contrasts between C 3 and C 4 grasses may be misinterpreted without phylogenetic control.• Here, we compared leaf physiology and growth in multiple lineages of C 3 and C 4 grasses sampled from a monophyletic clade, and asked the following question: which ecophysiological traits differ consistently between photosynthetic types, and which vary among lineages?• C 4 species had lower stomatal conductance and water potential deficits, and higher water-use efficiency than C 3 species. Photosynthesis and nitrogen-use efficiency were also greater in C 4 species, varying markedly between clades. Contrary to previous studies, leaf nitrogen concentration was similar in C 4 and C 3 types. Canopy mass and area were greater, and root mass smaller, in the tribe Paniceae than in most other lineages. The size of this phylogenetic effect on biomass partitioning was greater in the C 4 NADP-me species than in species of other types.• Our results show that the phylogenetic diversity underlying C 4 photosynthesis is critical to understanding its functional consequences. Phylogenetic bias is therefore a crucial factor to be considered when comparing the ecophysiology of C 3 and C 4 species.

Summary• The evolution of C 4 photosynthesis in plants has allowed the maintenance of high CO 2 assimilation rates despite lower stomatal conductances. This underpins the greater water-use efficiency in C 4 species and their tendency to occupy drier, more seasonal environments than their C 3 relatives.• The basis of interspecific variation in maximum stomatal conductance to water (g max ), as defined by stomatal density and size, was investigated in a common-environment screening experiment. Stomatal traits were measured in 28 species from seven grass lineages, and comparative methods were used to test for predicted effects of C 3 and C 4 photosynthesis, annual precipitation and habitat wetness on g max .• Novel results were as follows: significant phylogenetic patterns exist in g max and its determinants, stomatal size and stomatal density; C 4 species consistently have lower g max than their C 3 relatives, associated with a shift towards smaller stomata at a given density. A direct relationship between g max and precipitation was not supported. However, we confirmed associations between C 4 photosynthesis and lower precipitation, and showed steeper stomatal size-density relationships and higher g max in wetter habitats.• The observed relationships between stomatal patterning, photosynthetic pathway and habitat provide a clear example of the interplay between anatomical traits, physiological innovation and ecological adaptation in plants.

Plant growth plays a key role in the functioning of the terrestrial biosphere, and there have been substantial efforts to understand why growth varies among species. To this end, a large number of experimental analyses have been undertaken; however, the emergent patterns between growth rate and its components are often contradictory. We believe that these conflicting results are a consequence of the way growth is measured. Growth is typically characterized by relative growth rate (RGR); however, RGR often declines as organisms get larger, making it difficult to compare species of different sizes. To overcome this problem, we advocate using nonlinear mixed-effects models so that RGR can be calculated at a standard size, and we present easily implemented methods for doing this. We then present new methods for analyzing the traditional components of RGR that explicitly allow for the fact that is log (RGR) the sum of its components. These methods provide an exact decomposition of the variance in . Finally, we use simple analytical log (RGR) and simulation approaches to explore the effect of size variation on growth and its components and show that the relative importance of the components of RGR is influenced by the extent to which analyses standardize for plant size.

C4 photosynthesis is an important example of convergent evolution in plants, having arisen in eudicots, monocots and diatoms. Comparisons between such diverse groups are confounded by phylogenetic and ecological differences, so that only broad generalisations can be made about the role of C4 photosynthesis indetermining ecophysiological traits. However, 60% of C4 species occur in the grasses (Poaceae) and molecular phylogenetic techniques confirm that there are between 8 and 17 independent origins of C4 photosynthesis in the Poaceae. In a screening experiment, we compared leaf physiology and growth traits across several majorindependent C3 & C4 groups within the Poaceae, asking 1) which traits differ consistently between photosynthetictypes and 2) which traits differ consistently between clades within each photosynthetic type.

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