Calcium (Ca2 ) deficiency is rare in nature, but excessive Ca2+ restricts plant communities on calcareous soils (White and Broadley, 2003). Ecologists have classified plant species into calcifuges, which occur on acid soils (with low effective cation exchange capability and so low Ca2+) and do not grow well in lime-rich (calcareous) soils due to limitations in mineral nutrition (Zohlen and Tyler, 1997), whereas calcicoles occur on calcareous soils (soils with high Ca2+ content) (Lee, 1999). Also, the Ca2+ concentrations in calcifuge and calcicole plants growing in their natural habitat differ markedly. An interest in particular edaphic constraints of calcicole and calcifuge plants extends back to studies in the 18th century (Rorison, 1960). Calcifuges generally grow well at low rhizospheric Ca2+ concentrations and, conversely, the mechanisms that enable calcicole plants to maintain low [Ca2+]cyt in their natural habitat are believed to restrict their growth at low rhizospheric Ca2+ by inducing Ca2+ deficiency (Jefferies and Willis, 1964; Lee, 1999). De Silva (1934) carried out the initial studies to understand the distribution of a few plant species on both calcareous as well as acidic soils and determined that the amount of exchangeable Ca2+ in the soils correlated with the distribution of calcicoles and soil condition being the chief factor for calcifuges.
Soils normally have large amounts of exchangeable Ca2 in the range of 300-5000 ppm (Kelling and Schulte, 1998). A broad variety of studies report
2+
variable Ca2+ concentrations in soils from regions across the UK. Total Ca levels in dried (80°C) soil from four sites including two sites in Gordano Valley (N. Somerset), Cadbury Camp (N. Somerset) and in Braunton Burrows (N.
Devon) ranged from 6,650 ppm to 12,520 ppm (Jefferies and Willis, 1964). The mean extractable Ca2+ concentration of soil collected from Royston,
Hertfordshire was 5840 ppm (approx.) (Wilson et a/., 1995). Total soil Ca2+
from soils collected from eight sites within the Westerleigh-Yate area (northeast of Bristol, England) ranged from 1,250 ppm to 5,540 ppm (Morgan
et a/., 2001).
Calcareous grasslands in the UK and Europe are sites of conservation importance that support a diverse and often specialised flora and fauna,
including many rare or threatened species (Van Helsdingen et a/., 1996;
Rodwell, 1992; Wallis DeVries et a/., 2002; Bobbink and Willems, 1987). Also,
the most species-rich plant populations are found among the calcareous
grasslands around Europe (Hillier et a/., 1999; Rodwell, 1991). The question
of why some soil substrates host a greater diversity of plant species than others is old, and generations of plant ecologists have put emphasis on these studies (Wohlgemuth and Gigon, 2003). The adaptations shown by plants that grow in calcareous soils (calcicoles) to the distinctive habitats have long fascinated ecologists (Lee, 1999), yet our understanding of the physiological basis of the calcicolous habit remains limited. The derived question of whether plant species richness differs depending on calcareous or acidic substrate has thus far been of trivial scientific interest in Europe (Kinzel, 1983; Wohlgemuth and Gigon, 2003). Huge losses among these grasslands and their continuing vulnerability to either agricultural improvement or neglect have focussed attention on the need for conservation of their biodiversity (DETR, 2000).
De Silva et at. (1994a) established that transpiration rate was
unchanged with increasing concentrations of rhizospheric Ca up to 20 mM
in three calcicole species, Campanula Qlomerata, Centaurea scabiosa and
Leontodon hispidus grown at high concentrations of rhizospheric Ca2+, although concentrations of Ca2+ in the xylem sap were very close to those in the rhizosphere. These results were in marked contrast to those observed for
Lupinus luteus, a known calcifuge, subjected to similar experimental conditions (De Silva et al., 1994b). In this species xylem concentrations were maintained at 2 mM even when rhizospheric Ca2+ concentrations were 15 mM. Further to this, they observed that high rhizospheric Ca2+ disrupted stomatal behaviour in L. luteus, resulting in a significant reduction of leaf conductance, transpiration rate and net assimilation rate which was again in contrast with that observed in calcioles (De Silva et al., 1994a), where the stomatal opening was unperturbed by high rhizospheric Ca2+. These studies suggested that a strategy has been developed in calcicoles for protecting the guard cells from deleterious concentrations of this cation by regulating the distribution of Ca2+ reaching the epidermal tissues. X-ray microanalysis in the
leaves of Centaurea scabiosa and Leontodon hispidus showed that the bulk
of the Ca2+ entering the epidermis is limited within the trichomes, most likely
as insoluble Ca2+ oxalate (De Silva et al., 2001, 1996). The Ca2+
concentration in the region of stomata in these species thus remains minimal, allowing efficient intracellular signaling to continue in the guard cells without any hindrance. This study also provides evidence of a very specific regulatory
2+ mechanism operating in calcicoles where the trichomes act as Ca sequestering structures. A similar mechanism was reported in plants with an induced expression of Ca2+-transporters demonstrating Ca -deficiency
symptoms at low rhizospheric Ca2+, where the Ca2+ is removed from the cytoplasm to the vacuole (Hirschi, 2001).
It is likely that part of the adaptation of plants to survival on high Ca2+ soils would be echoed at the molecular level by changes in the expression of various genes underlying the many biochemical pathways, including Ca2+ regulation and Ca2+ signaling, that are operating in these plants. For example, the plant H+/Ca2+ antiporter (CAX1) was identified by its ability to re-establish growth on high Ca2+ media of a yeast mutant defective in vacuolar Ca2+
transport (Hirschi et al., 1996). Additionally, tobacco plants over-expressing
CAX1 showed an alleviated response to Ca2+ deficiency by increasing Ca2+
uptake from the media (Hirschi, 2001). Also, Chan et al (2003) demonstrated
a hypersensitive response by CNGC2 mutants of Arabidopsis in response to
increased rhizospheric Ca2+ which they ascribed to Ca2+ toxicity, perhaps due to accumulation of Ca2+ or to a defect in signaling pathways that facilitate normal growth on high Ca2+.