CODENET: WORK AREA 1 - EVOLUTIONARY BIODIVERSITY

This work area comprises study of the keystone taxa as representatives of the total biodiversity of coccolithophorids. This page outlines the background, methodology, and objectives of this work area.

BACKGROUND

Coccolithophorid algae belong to the phylum Haptophyta (also known as Prymnesiophyta). These are unicellular algae with one or two large chlorophyll a and c- containing chloroplasts, which are surrounded by four membranes. Motile phases bear two flagella and a haptonema. The haptonema is a unique organelle with an ultrastructure of usually seven microtubules, in contrast to the 9 + 2 microtubular structure of flagella. There are two sub-classes within the Haptophyta: the Pavlovophycideae and Prymnesiophycideae. The Prymnesiophycideae have smooth sub-equal flagella and organic scales. The Pavlovophycideae have unequal flagella and lack scales. Separation of these two sub-classes and of haptophyta from other algae is well supported by a range of cytological, biochemical and molecular genetic data (Green & Jordan 1994, Medlin et al. 1996).

The Prymnesiophycidae includes both coccolithophorids and many non-calcifying taxa (e.g. Chrysochromulina, Isochrysis, Prymnesium, Phaeocystis). Formerly these were regarded as distinct groups but cytological and biochemical evidence suggested that at least some non-calcifying genera were derived from coccolithophorids (Green et al. 1989). Similarly there are calcifying taxa, such as Braarudosphaera, Polycrater , and Florisphaera, which produce nannoliths (i.e. plates very different to typical coccoliths) and which may have variable relations to coccolithophorids. Finally there are two main types of coccoliths: holococcoliths and heterococcoliths which have completely separate structures but which can occur on haploid and diploid stages, respectively, of a single species (e.g. Parke & Adams 1960, Thomsen 1991, Billard 1994).

Available biological and biochemical data is inadequate to resolve this diversity into a coherent phylogenetic scheme (e.g. Green & Jordan 1994). Stratophenetic analysis of the fossil record, however, suggests a consistent large scale phylogeny (Perch-Nielsen 1985a,b, Bown et al. 1991, Young et al. 1994). The morphologically defined heterococcolithophorid families can all at least tentatively be traced to a common origin in the Early Jurassic, when the main radiation of the group occurred (Bown 1987). Supporting this, all heterococcoliths appear to share a common biomineralization pattern, V/R mode nucleation (Young et al. 1992, 1994, Bown & Young in prep.). So, the apparent complications are not incompatible with the hypothesis that coccolith structure/biomineralization pattern can be used as a reliable basis for phylogenetic inference and that the heterococcolithophorids form a natural, albeit paraphyletic, group.

Our proposed research will greatly expand our knowledge of the major evolutionary steps in the diversification of the coccolithophorids. We will be able to test extensively the models established for life cycles strategies, biomineralization, endsymbiosis and its concomitant gene reduction as well as divergencies in the fossil record. In addition, we will be able to provide biological calibrations for palaeontological interpretations of phylogeny and to use the phylogeny to probe biodiversity in biochemical and cell biological characters.

METHODOLOGY

A key part of our strategy is to adopt a phylogenetic approach to the problem of sampling and interpreting biodiversity. Even for a relatively small group such as coccolithophorids it is only practical to investigate in detail a few species. However, by co-ordinating observations on a select set of species our data can be maximised. The basic logic is that if the character distribution in the sampled set of species parallels the phylogeny, then we can predict the character distribution in the unsampled species. The most useful extreme result is that all the sampled species show common characters, in this case we can predict that this result will hold for all taxa within the clade defined by the sampled species. By sampling the broadest clade possible relative to the known phylogeny we maximise the value of such results. Finer resolution sampling allows us to investigate the level at which phylogenetically variable characters are determined.

Broad sampling is achieved in our set of keystone species by including members of the Syracosphaeraceae, Helicosphaeraceae, Coccolithaceae and Noelearhabdaceae (FIG X). We believe these groups diverged during the Liassic radiation (ca. 200 Ma) of coccolithophorids and predict that any characters held in common will be primitive for the coccolithophorids in general - and for derived non-calcifying haptophytes.

An intermediate level of sampling is provided by inclusion in our data set of three members of the Coccolithaceae, which we believe diverged in the mid Tertiary (ca 20 Ma).

The lowest level of sampling is provided by investigation of the genus Gephyrocapsa which is diverse at the present day following radiation during the Pleistocene, beginning approximately 2 Ma. At present it consists of 5 or 6 closely related species, in addition the well-studied species Emiliania huxleyiis a descendant of Gephyrocapsa (divergence approximately 250 ka) and so a member of the same clade. This group will be studied in special detail (ETHZ, AWI, NIOZ) allowing us to investigate the degree to which characters are affected by micro-evolutionary processes.

Our work will consist of three types of study:

1. Analysis of modern diversity: The phylogenetic approach will underpin our study of: life-cycles; lipid biomarkers; photosynthetic pigments; plastid genome size, and; coccolith ultrastructure. In each of these areas detailed examination of the keystone taxa will allow us to make a major advance in our knowledge, and provide a firm grounding for interpreting the more heterogeneous data already available in the literature and from our other studies. Where appropriate these studies may also be extended to include collection of new comparative data from other species.

2. Molecular clock calibration: Our set of taxa provide sampling of widely varying divergence times. The excellent fossil record of the coccolithophorids will enable us to constrain divergence times and so to calibrate molecular clock estimates of divergence rates. Since genetic studies are producing an enormous volume of data suitable for estimation of divergence rates calibration studies such as these exploiting a good geological record are urgently needed. Our results should be of wide applicability within protoctist and particularly phytoplankton studies.

3. Development of phylogeny: In addition to using our existing understanding of coccolithophorid phylogeny the project will develop and test this phylogeny. This will come from studies of molecular genetics, cladistic analysis of morphological and biological data, and palaeontological studies of evolutionary divergences. This work will maximise the robustness of our results and provide a firm framework for interpretation of phylogenetic data from other coccolithophorids.

This set of studies involves a range of advanced methodologies, many at the forefront of modern science - e.g. molecular biology, liquid column chromatography, refined HPLC analysis, and image analysis. These cannot be detailed here, but the participants, are international leaders in the various relevant fields as explained in the research team details.

OBJECTIVES

The overall objective is to understand the variability shown by coccolithophorids in a range of key areas relative to phylogeny. Specifically for each of the keystone taxa our objectives are to:

• 1. Determine their life-cycles. The limited data available suggests that coccolithophorids, and other haptophytes, have a complex life-cycle typically including haploid and diploid phases, each capable of reproducing asexually (Billard 1994, Green et al. 1996). The ecological significance of these phase changes is still unknown and it is unclear how variable the life-cycles are. (RT2; U. Caen)
• 2. Investigate their cytology and scale morphology. Transmission electron microscopy will be used to examine ultrastructure of organic scales, the flagellar apparatus and intracellular formation of scales and coccoliths. (RT7; U. Caen)
• 3. Characterise their lipid composition. Distinctive alkenone lipids formed by coccolithophorids constitute excellent biomarkers with high preservation potential in the geological record (de Leeuw et al. 1980, Conte et al. 1994). In addition variation in saturation ratios of these alkenones form an important palaeothermometer, UK37 index (Brassell et al. 1986, Jordan et al. 1996). Our work will determine the chemotaxonomic significance of variability in these and other potential biomarkers and palaeothermometer calibrations will be performed on species containing the alkenones. (RT4A; NIOZ, CSIC)
• 4. Determine coccolith ultrastructure and ontogeny. Work on the keystone taxa will test and develop the model of Young et al. (1992) that heterococcolithophorid biomineralisation is characterised by a conserved mode of nucleation, with alternating sub-vertical and sub-radial c-axes. (RT6; NHM)
• 5. Determine photosynthetic pigment composition. Pigment composition variations are of importance for the ecological adaptation of taxa and for remote sensing detection. Present data on pigment composition is erratic (REF) (RT5, CSIC).
• 6. Measure plastid genome size. Preliminary evidence suggests that the haptophyte plastid genome is an order of magnitude larger than in any other algal group or higher plant with little repetitive DNA. Investigations will indicate whether this is a universal feature and probe its implications. (RT3A; AWI)
• 7. Sequence the 18S rRNA, Tuf A genes and non-coding regions. These are slow to fast evolving components of the genome ideal for investigating high level phylogeny and relationships with other algal groups. (RT3B; AWI)
• 8. Analyse the palaeontological record of ancestral lineages to determine divergence times. (RT15; NHM, ETHZ)

KEY SYNTHETIC OBJECTIVES ARE TO:

• Determine the major patterns of biodiversity in coccolithophorid life-cycles, biomarker composition, photosynthetic pigments, cytology and plastid genome.
• Re-evaluate the phylogeny of the coccolithophorids using separate and combined analyses of: molecular genetic (AWI), morphological (NHM) and biochemical (NIOZ, CSIC) data and compare this with the palaeontological record of coccolithophorid evolution (NHM, ETHZ). Calculate divergence times of groups and rates of evolution, including molecular clock calibrations (AWI, ETHZ).
• Reconstruct the sequence of major evolutionary steps in coccolithogenesis, lipid biochemistry, plastid evolution, and life cycle differentiation.