Juan Pedro Ferrio

Mesophyll diffusion conductance to CO2: An unappreciated central player in photosynthesis

Mesophyll diffusion conductance to CO(2) is a key photosynthetic trait that has been studied intensively in the past years. The intention of the present review is to update knowledge of g(m), and highlight the important unknown and controversial aspects that require future work. The photosynthetic limitation imposed by mesophyll conductance is large, and under certain conditions can be the most significant photosynthetic limitation. New evidence shows that anatomical traits, such as cell wall thickness and chloroplast distribution are amongst the stronger determinants of mesophyll conductance, although rapid variations in response to environmental changes might be regulated by other factors such as aquaporin conductance. Gaps in knowledge that should be research priorities for the near future include: how different is mesophyll conductance among phylogenetically distant groups and how has it evolved? Can mesophyll conductance be uncoupled from regulation of the water path? What are the main drivers of mesophyll conductance? The need for mechanistic and phenomenological models of mesophyll conductance and its incorporation in process-based photosynthesis models is also highlighted.

Drought effects on allocation of recent carbon: from beech leaves to soil CO2 efflux

*Recent studies have highlighted a direct, fast transfer of recently assimilated C from the tree canopy to the soil. However, the effect of environmental changes on this flux remains largely unknown. *We investigated the effects of drought on the translocation of recently assimilated C, by pulse-labelling 1.5-yr-old beech tree mesocosms with (13)CO(2). (13)C signatures were then measured daily for 1 wk in leaves, twigs, coarse and fine root water-soluble and total organic matter, phloem organic matter, soil microbial biomass and soil CO(2) efflux. *Drought reduced C assimilation and doubled the residence time of recently assimilated C in leaf biomass. In phloem organic matter, the (13)C label peaked immediately after labelling then decayed exponentially in the control treatment, while under drought it peaked 4 d after labelling. In soil microbial biomass, the label peaked 1 d after labelling in the control treatment, whereas under drought no peak was measured. Two days after labelling, drought decreased the contribution of recently assimilated C to soil CO(2) efflux by 33%. *Our study showed that drought reduced the coupling between canopy photosynthesis and belowground processes. This will probably affect soil biogeochemical cycling, with potential consequences including slower soil nitrogen cycling and changes in C-sequestration potential under future climate conditions.

Δ13C and tree-ring width reflect different drought responses in Quercus ilex and Pinus halepensis

Holm oak (Quercus ilex L.) and Aleppo pine (Pinus halepensis Mill) are representative of two different functional types of trees extensively found in the Mediterranean: evergreen sclerophyllous and drought-adapted conifers. The former is considered a partially drought-tolerant species, whereas the latter is a typically drought-avoiding, water-saving species. We postulated that contrasting strategies in response to water deficits in Q. ilex and P. halepensis would lead to a differential sensitivity to changes in water availability. To test this hypothesis, we compared the response of both species in growth rate (measured as radial increments) and intrinsic water use efficiency [WUEi, as inferred from carbon isotope discrimination (Δ13C) in wood samples] among sites from different provenance regions in NE Spain. We found significant differences in Δ13C and growth among provenance regions, partly explained by contrasting water availability. Wood Δ13C was positively related with precipitation and the ratio between precipitation and potential evapotranspiration (P / E). However, these relationships were stronger in P. halepensis (for P / E, r2=0.67, P <0.001) than in Q. ilex (r 2=0.42, P <0.01). In addition, radial growth was positively related with precipitation and Δ13C in P. halepensis (r 2=0.32 and r2=0.35, respectively, P <0.01), but not in Q. ilex. We concluded that P. halepensis was more sensitive than Q. ilex to water availability, showing faster increase in WUEi in response to water stress. We also found that the effect of north/south aspect on Δ13C and growth was site-specific, and unrelated to climatic variables.

Temporal dynamics of the carbon isotope composition in a Pinus sylvestris stand: from newly assimilated organic carbon to respired carbon dioxide

The 13C isotopic signature (C stable isotope ratio; δ13C) of CO2 respired from forest ecosystems and their particular compartments are known to be influenced by temporal changes in environmental conditions affecting C isotope fractionation during photosynthesis. Whereas most studies have assessed temporal variation in δ13C of ecosystem-respired CO2 on a day-to-day scale, not much information is available on its diel dynamics. We investigated environmental and physiological controls over potential temporal changes in δ13C of respired CO2 by following the short-term dynamics of the 13C signature from newly assimilated organic matter pools in the needles, via phloem-transported organic matter in twigs and trunks, to trunk-, soil- and ecosystem-respired CO2. We found a strong 24-h periodicity in δ13C of organic matter in leaf and twig phloem sap, which was strongly dampened as carbohydrates were transported down the trunk. Periodicity reappeared in the δ13C of trunk-respired CO2, which seemed to originate from apparent respiratory fractionation rather than from changes in δ13C of the organic substrate. The diel patterns of δ13C in soil-respired CO2 are partly explained by soil temperature and moisture and are probably due to changes in the relative contribution of heterotrophic and autotrophic CO2 fluxes to total soil efflux in response to environmental conditions. Our study shows that direct relations between δ13C of recent assimilates and respired CO2 may not be present on a diel time scale, and other factors lead to short-term variations in δ13C of ecosystem-emitted CO2. On the one hand, these variations complicate ecosystem CO2 flux partitioning, but on the other hand they provide new insights into metabolic processes underlying respiratory CO2 emission.

Carbon and oxygen isotope ratios in wood constituents of Pinus halepensis as indicators of precipitation, temperature and vapour pressure deficit

Carbon and oxygen isotope compositions (δ13C, δ18O) in tree rings have been shown to bear relevant climatic signals. However, little is known about the interrelationship between both isotopes in wood constituents for species from other than relatively wet climates. We hypothesized that in a species adapted to temporary droughts (e.g. Pinus halepensis Mill.) the signal derived from δ18O in precipitation would be hidden by the strong variability in leaf transpirative enrichment. To test this assumption, we compared the effect of precipitation, temperature and vapour pressure deficit (VPD) on δ18O and δ13C along 23 sites covering the ecological range for this species. We extracted the cores from the south side of four to six adult dominant trees per aspect (north/south) within each site. For each aspect and site, fragments of the period 1975–1999 were pooled and milled to a fine powder. To further test the postulated need for cellulose purification in the assessment of climatic information, we studied these relationships in whole and extracted wood, holocellulose and lignin. In all wood fractions, δ13C was related to annual precipitation [r=−0.58 (P < 0.01) to −0.78 (P < 0.001)] and VPD [r= 0.53 (P < 0.01) to 0.57 (P < 0.01)]. In contrast, for δ18O only holocellulose showed consistent relationships with climatic data, being strongly significant for VPD [r= 0.66 (P < 0.001)]. However, it was unrelated to modelled δ18O in precipitation, confirming that transpirative enrichment (driven by VPD) dampened the source signal in P. halepensis. The relationships between δ13C and δ18O were generally poor, regardless of the wood constituent, suggesting that although both variables were somewhat related to transpirative demand, they were relatively independent. This was further confirmed by building stepwise models using both isotopes to predict annual and seasonal precipitation [r2= 0.34 (P < 0.01) to 0.68 (P < 0.001)], temperature [r2= 0.15 (P < 0.05) to 0.37 (P < 0.01)] and VPD [r2= 0.31 (P < 0.01) to 0.55 (P < 0.001)]. We concluded that, even when partially describing the same climate variables, the information underlying the two isotopes can be regarded as complementary.

On the metabolic origin of the carbon isotope composition of CO2 evolved from darkened light‐acclimated leaves in Ricinus communis

The (13)C isotopic signature (delta(13)C) of CO(2) respired from plants is widely used to assess carbon fluxes and ecosystem functioning. There is, however, a lack of knowledge of the metabolic basis of the delta(13)C value of respired CO(2). To elucidate the physiological mechanisms driving (12)C/(13)C fractionation during respiration, the delta(13)C of respired CO(2) from dark-acclimated leaves during the night, from darkened leaves during the light period, and from stems and roots of Ricinus communis was analysed. The delta(13)C of potential respiratory substrates, the respiratory quotient and the activities of phosphoenolpyruvatecarboxylase (PEPc) and key respiratory enzymes were also measured. It is shown here that the CO(2) evolved from darkened light-acclimated leaves during the light period is (13)C-enriched, and that this correlates with malate accumulation in the light and rapid malate decarboxylation just after the onset of darkness. Whilst CO(2) evolved from leaves was generally (13)C-enriched (but to a lesser extent during the night), CO(2) evolved from stems and roots was depleted compared with the putative respiratory substrates; the difference was mainly caused by intensive PEPc-catalysed CO(2) refixation in stems and roots. These results provide a physiological explanation for short-term variations of delta(13)C in CO(2), illustrating the effects of variations of metabolic fluxes through different biochemical pathways.

The long way down—are carbon and oxygen isotope signals in the tree ring uncoupled from canopy physiological processes?

The carbon (δ(13)C) and oxygen (δ(18)O) stable isotope composition is widely used to obtain information on the linkages between environmental drivers and tree physiology over various time scales. The tree-ring archive can especially be exploited to reconstruct inter- and intra-annual variation of both climate and physiology. There is, however, a lack of information on the processes potentially affecting δ(13)C and δ(18)O on their way from assimilation in the leaf to the tree ring. As a consequence, the aim of this study was to trace the isotope signals in European beech (Fagus sylvatica L.) from leaf water (δ(18)O) and leaf assimilates (δ(13)C and δ(18)O) to tree-ring wood via phloem-transported compounds over a whole growing season. Phloem and leaf samples for δ(13)C and δ(18)O analyses as well as soil water, xylem water, leaf water and atmospheric water vapour samples for δ(18)O analysis were taken approximately every 2 weeks during the growing season of 2007. The δ(13)C and δ(18)O samples from the tree rings were dated intra-annually by monitoring the tree growth with dendrometers. δ(18)O in the phloem organic matter and tree-ring whole wood was not positively related to leaf water evaporative enrichment and δ(18)O of canopy organic matter pools. This finding implies a partial uncoupling of the tree-ring oxygen isotopic signal from canopy physiology. At the same time, internal carbon storage and remobilization physiology most likely prevented δ(13)C in tree-ring whole wood from being closely related to intra-annual variation in environmental drivers. Taking into account the post-photosynthetic isotope fractionation processes resulting in alterations of δ(13)C and δ(18)O not only in the tree ring but also in phloem carbohydrates, as well as the intra-annual timing of changes in the tree internal physiology, might help to better understand the meaning of the tree-ring isotope signal not only intra- but also inter-annually.

Climate-related variability in carbon and oxygen stable isotopes among populations of Aleppo pine grown in common-garden tests

The Aleppo pine (Pinus halepensis Mill.) is found in the Mediterranean under a broad range of moisture and thermal conditions. Differences in severity and duration of water stress among native habitats may act as selective forces shaping the populations’ genetic make-up in terms of contrasting drought strategies. We hypothesised that these strategies should translate into intraspecific variation in carbon isotope composition (δ13C, surrogate of intrinsic water-use efficiency, WUEi) of wood holocellulose, and such variation might be linked to changes in oxygen isotope composition (δ18O, proxy of stomatal conductance) and to some climatic features at origin. Thus, we evaluated δ13C, δ18O, growth and survival for 25 Aleppo pine populations covering its geographic range and grown in two common-garden tests. We found intraspecific variability for δ13C and growth, with high-WUEi populations (which showed 18O-enriched holocellulose) having low growth. These results suggest stomatal regulation as common control for δ13C and productivity. We also detected sizeable relationships between δ13C and climate factors related to the magnitude and timing of drought such as the ratio of summer to annual rainfall. The main climate variable associated with δ18O was minimum temperature, but only in the coldest trial, suggesting differences in growth rhythms among sources. Overall, slow growing populations from highly-seasonal dry areas of the western Mediterranean exhibited a conservative water-use, as opposed to fast growing sources from the northernmost distribution range. The particular behaviour of the Mediterranean Aleppo pine as compared with other conifers demonstrates different selective roles of climate variables in determining intraspecific fitness.

Stable carbon and nitrogen isotopes and quality traits of fossil cereal grains provide clues on sustainability at the beginnings of Mediterranean agriculture

We present a novel approach to study the sustainability of ancient Mediterranean agriculture that combines the measurement of carbon isotope discrimination (Delta(13)C) and nitrogen isotope composition (delta(15)N) along with the assessment of quality traits in fossil cereal grains. Charred grains of naked wheat and barley were recovered in Los Castillejos, an archaeological site in SE Spain, with a continuous occupation of ca. 1500 years starting soon after the origin of agriculture (ca. 4000 BCE) in the region. Crop water status and yield were estimated from Delta(13)C and soil fertility and management practices were assessed from the delta(15)N and N content of grains. The original grain weight was inferred from grain dimensions and grain N content was assessed after correcting N concentration for the effect of carbonisation. Estimated water conditions (i.e. rainfall) during crop growth remained constant for the entire period. However, the grain size and grain yield decreased progressively during the first millennium after the onset of agriculture, regardless of the species, with only a slight recovery afterwards. Minimum delta(15)N values and grain N content were also recorded in the later periods of site occupation. Our results indicate a progressive loss of soil fertility, even when the amount of precipitation remained steady, thereby indicating the unsustainable nature of early agriculture at this site in the Western Mediterranean Basin. In addition, several findings suggest that barley and wheat were cultivated separately, the former being restricted to marginal areas, coinciding with an increased focus on wheat cultivation.

The Peclet effect on leaf water enrichment correlates with leaf hydraulic conductance and mesophyll conductance for CO2

Leaf water gets isotopically enriched through transpiration, and diffusion of enriched water through the leaf depends on transpiration flow and the effective path length (L). The aim of this work was to relate L with physiological variables likely to respond to similar processes. We studied the response to drought and vein severing of leaf lamina hydraulic conductance (K(lamina) ), mesophyll conductance for CO(2) (g(m) ) and leaf water isotope enrichment in Vitis vinifera L cv. Grenache. We hypothesized that restrictions in water pathways would reduce K(lamina) and increase L. As a secondary hypothesis, we proposed that, given the common pathways for water and CO(2) involved, a similar response should be found in g(m) . Our results showed that L was strongly related to mesophyll variables, such as K(lamina) or g(m) across experimental drought and vein-cutting treatments, showing stronger relationships than with variables included as input parameters for the models, such as transpiration. Our findings were further supported by a literature survey showing a close link between L and leaf hydraulic conductance (K(leaf) = 31.5 × L(-0.43) , r(2) = 0.60, n = 24). The strong correlation found between L, K(lamina) and g(m) supports the idea that water and CO(2) share an important part of their diffusion pathways through the mesophyll.