Home > Major Projects > Evolution of Thermoacidophilic Algae

  The Phylogeny and Evolution of the
Thermoacidophilic Cyanidiales		  

  SUMMARY:
The objective of this project is to isolate different plastid-encoded genes from cultured and environmental samples of Cyanidiales. The main goal of these experiments is to discover new taxa and analyze cultured strains in this primitive order of red algae to address the biodiversity of eukaryotic thermoacidophiles. We are also studying poorly understood mesophilic “cave” Cyanidiales to assess their relationship to the different extremophilic genera. The molecular phylogenetic framework will form the platform for addressing Cyanidiales morphological evolution through detailed TEM analyses and their physiological ecology through growth analysis in different culture conditions. The work is done in collaboration with the lab of Gabriele Pinto at the Universita Federico II, Dipartimento di Biologia vegetale, Via Foria 223, 80139 Napoli, Italy. Post-docs Claudia Ciniglia from Napoli and Hwan Su Yoon in Iowa City have taken charge of this project and work in both labs. Presently, we are focusing on environmental samples from the Phlegrean Fields near Napoli and mesophilic samples from the Sybilla Cave and from Monte Rotaro in Ischia. We will, in the future, analyze environmental samples of Cyanidiales from Yellowstone National Park and other extreme environments. The initial analyses of the Napoli material have identified a plethora of new genera and species that are currently being isolated and analyzed using TEM. Our prediction is that the present understanding of Cyanidiales biodiversity and distribution has been limited by the availability of only a handful of cultured strains. The environmental samples promise to radically revise our understanding of Cyanidiales evolution and lead to a major revision of their taxonomy. In the long-term, our work will form the basis for using the thermoacidophilic Cyanidiales as a biotechnological resource and for understanding the nature and ramifications of thermo-acidoadaptation in eukaryotes. This research is partially funded by a grant to D. Bhattacharya from the Systematics Program at the National Science Foundation (DEB 01-07754).		  

  SPECIFIC AIMS OF PROJECT:  
1. Isolate psaA, psbA, and rbcL coding regions from different environmental samples of Cyanidiales from Italy and from existing cultures. Characterize up to 50 clones from each environmental site.  
2. Develop plastid gene RFLP markers for all genera of Cyanidiales, with concentration in the Phlegrean Fields samples, to allow rapid assessment of species representation from different field sites.  
3. Create a robust phylogeny of the plastid sequences.  
4. Study in detail the species distribution at 5 environmentally divergent sites (e.g., acid fumes, cryptoendolithic, moderate temperature but low pH, mesophilic) in Pisciarelli (Napoli) to address the biodiversity at each site.  
5. Erect a robust taxonomy of the Cyanidiales.  
6. Do TEM analyses of all existing and new species/genera of Cyanidiales to address the utility of morphological markers in a taxonomic setting and generally to understand phenotypic plasticity in these taxa.  
7. Study the physiology of all taxa by culturing them in different in vitro conditions.  
8. Isolate individual cells from each Pisciarelli site to do population genetic analyses of selected species to understand gene flow and extent of sexual reproduction in these taxa.  

  Previous work has resulted in the following paper that is in press at Cryptogamie Algologie.  
  Comparative Approaches to the Taxonomy of the Genus Galdieria Merola (Cyanidiales, Rhodophyta).
Gabriele Pinto, Patrizia Albertano, Claudia Ciniglia, Salvatore Cozzolino, Antonino Pollio, Hwan Su Yoon, Debashish Bhattacharya

 

  
 
TEM micrographs of cells of Galdieria. (1) sporangium of G. sulphuraria from Napoli. (2) young cell of G. sulphuraria from Napoli; (3) mature cell of G. sulphuraria from California; arrowhead showing the protrusions of the innermost layer of the cell wall (transfer-cell like structure); (4) mature cell of G. sulphuraria from Yellowstone National Park; arrow indicates osmiophilic globules in the plastid stroma. (5) detailed view of a section of a mature cell of G. sulphuraria from Yellowstone National Park. Arrows indicates dyctiosomes. (6) mature cells of G. sulphuraria from Java, with coalescent vacuoles. Abbreviations: m = mitochondrion; n = nucleus; v = vacuole; vs = vesicles.

 


A Recent publication in Molecular Ecology  
  The hidden biodiversity of extremophilic eukaryotes
Claudia Ciniglia1,3, Hwan Su Yoon2,3 , Antonino Pollio1, Gabriele Pinto1, Debashish Bhattacharya2,4		  
  1Dipartimento di Biologia vegetale, Università “Federico II”, via Foria 223, 80139 Napoli, Italy
2Department of Biological Sciences and Center for Comparative Genomics, University of Iowa, 210 Biology Building, Iowa City, Iowa 52242, United States
3These authors contributed equally to this work
* Corresponding author

  
  Abstract: The Cyanidiales is a group of unicellular red algae that thrive in acidic and high temperature conditions around hot springs. These taxa have a relatively simple morphology and are classified in three genera, Cyanidium, Cyanidioschyzon, and Galdieria. Little is known about the biodiversity of Cyanidiales, their population structure, and their phylogenetic relationships. Here we used a taxonomically broadly sampled 3-gene data set of plastid sequences to infer a robust phylogenetic framework for the Cyanidiales. The phylogenetic analyses support the existence of at least four distinct Cyanidiales lineages: the Galdieria spp. lineage, the Cyanidium caldarium lineage, a novel monophyletic lineage composed of mesophilic Cyanidium spp., and the Cyanidioschyzon merolae plus G. maxima lineage. Our analyses do not support the notion of a mesophilic ancestry of the Cyanidiales, rather they suggest the ancestral condition to have been thermo-acidotolerance. We also used environmental PCR with the rbcL gene to sample Cyanidiales biodiversity at five ecologically distinct sites at Pisciarelli in the Phlegrean Fields in Italy (see Fig. 1). This analysis showed a high level of sequence divergence among Cyanidiales species and the partitioning of taxa based on environmental conditions. Our research revealed an unexpected level of genetic diversity among Cyanidiales that revises current thinking about the phylogeny and biodiversity of this group (see Fig. 2). We predict that future environmental PCR studies will significantly augment the biodiversity that we have discovered and demonstrate the Cyanidiales to be a species-rich branch of red algal evolution.  


Figure1
  

Fig. 1. The Cyanidiales red algae. A) TEM micrograph of a crypto-endolithic strain of Galdieria sulphuraria. The abbreviations denote the following: m = mitochondrion, n = nucleus, p = plastid, v = vacuole. Scale bars = 1 µM. B-F) The environmental sites A-E used to collect Cyanidiales at Pisciarelli in the Phlegrean Fields, Italy (see text for details).

Fig. 2. (see below). Phylogeny of the Cyanidiales inferred from minimum evolution (ME) analysis using the LogDet transformation of the combined plastid DNA sequences of psaA, psbA, and rbcL. Results of a ME-LogDet bootstrap analysis are shown above the branches, whereas the bootstrap values from a protein maximum likelihood analysis using the JTT evolutionary model are shown below the branches. Only bootstrap values > 60% are shown. The thick nodes represent > 95% Bayesian posterior probability for clades using the site-specific GTR model. The broken lines show tree rearrangements that were tested with the one-sided Kishino-Hasegawa test (Ciniglia et al. submitted).
     

 


Figure 2

  

The w ork done by Claudia Ciniglia and Hwan Su Yoon is focusing on a molecular ecological analysis of Cyanidiales at thermoacidophilic sites in Toscana, Italy. We wish to test the idea that the high biodiversity that was uncovered at the Phlegrean Fields is also found in Toscana. The preliminary data suggest surprisingly that the Toscana sites are home to a few dominant strains/species of Cyanidiales in contrast to what was found in Napoli. We are also now using analysis of nuclear intron sequences to determine the level of genetic divergence in monospecific populations at cryptoendolithic sites in Napoli and Toscana.

Evolution of Thermoacidophilic
Algae in the area of Naples, Italy:
Study sites HabitatsImages of NapoliOther Cyanidiales sites