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Protistology ■ 67
School of Medicine, The University of Tokyo 3 - School of Tropical Medicine and Global Health, Nagasaki University [email protected]
Plastids in apicomplexan parasites are highly degenerated. The organelle is nevertheless essential for completion of the parasite life cycle. Interestingly, an oyster parasite Perkinsus marinus, which is sister to dinoflagellates and close to Apicomplexa, also has a DNA-lacking, extremely degenerated plastid. Functional analysis of the cryptic organelle is attracting and required to understand the relationship between the organelle degeneration and parasitism. The transgenic technique is a convincing approach for the analyses ofproteins ofinterest and is practicable in P. marinus. However, each transfected cell must be isolated from untransfected cells by hand labor using a micromanipulator multiple times to obtain any transfected cell lines. This is because drug selection system has not been established. Here, we identified two drugs that are available for selection of transfected P. marinus cells. Firstly, we screened antibiotics shown utility in apicomplexan parasites and determined that blasticidin S, bleo-mycin and puromycin effectively inhibited the parasite growth. Then, their resistance genes were fused downstream of gfp or mCherry gene, and each construct was transfected to the parasite. After two months, the fluorescent signals were observed in almost all cells cultured with bleomycin or puromycin. Furthermore, dual transfected cells were selected by using the two drugs, which enables us to examine colocalization of plastid proteins. We believe that this system provides new opportunities for functional analyses ofthe plastids in the parasite.
A DRAFT GENOME OF THE ANAEROBIC FLAGELLATE CARPEDIEMONAS MEMBRA-NIFERA, A FREE-LIVING RELATIVE OF METAMONAD PARASITES Salas Leiva D.E.1, Kolisko M.2, Curtis B.1, Eme L.1, Kamikawa R.3, Roger A.1
1 - Centre for Comparative Genomics and Evolutionary Bioinformatics (CGEB), Department ofBiochemistry and Molecular Biology, Dalhousie University, Halifax, NS, Canada, B3H4R2
2 - Beatty Biodiversity Centre, Dept. Botany, University ofBritish Columbia
3 - Graduate School of Human and Environmental Studies, Kyoto University, Yoshida-Nihonmatsu cho, Kyoto 606-8501, Japan [email protected]
Carpediemonas membranifera is a free living flagellated metamonad related to diplomonad parasites such as Giardia intestinalis and Spironucleus
salmonicida. We are interested in elucidating the evolutionary transitions to anaerobiosis and parasitism within metamonada, and sequenced the genome of C. membranifera. The genome assembly is 22.4 Mb long with 11328 predicted protein-coding genes, 41% of those have introns. Automatic annotation was carried out by searching against the Interpro, PFAM, Prosite, TIGR databases to identify domains, assign putative functions and predict metabolic pathways. Expert manual annotation is underway for genes encoding proteins functioning in DNA repair, mRNA degradation, mitochondrion-related organelles, cell surface or external cellular processes involved in host tissue adhesion, immune evasion, pathogenicity, nutrient acquisition, metabolite transport and environmental sensing, among others. We have completed analyses ofthe DNA repair pathways. Those can drive sexual/ parasexual pathways, antigen diversification and copy number variation, and are of great importance for adaptive evolution. C. membranifera possesses a complete system for excision repair, and the double strand break repair machinery including 1) a homologous recombination pathway and 2) microhomology-mediated end joining and singlestrand annealing. Also, it has several gene family expansions, as well as, a complete repertoire of cell cycle checkpoints and sex-related proteins. G. intestinalis and S. salmonicida have minimalistic and slightly different versions ofthe pathways found in C. membranifera suggesting that there have been some secondary losses and modifications in diplomonads as a result of their parasitic lifestyle.
THE GENOMIC COST OF BECOMING A RED ALGAL FREELOADER Salomaki E.D., Lane C.E.
DepartmentofBiologicalSciences, University ofRho-de Island
An abundance of genomic and transcriptomic data have been gathered over the past decade providing a wealth of knowledge about what it takes to be a successful parasite. Genomes of highly derived eukaryotic parasites have been sequenced including those from formerly photosynthetic lineages including apicomplexans. These data have revealed fascinating innovations that evolved over hundreds of millions of years, enabling parasites to infect and evade their hosts. Unlike highly derived lineages of eukaryotic parasites, red algae appear to be fertile ground for adopting a parasitic life strategy as seen by numerous recent and independent evolutions of parasitic taxa. Red algal parasites provide a great system to investigate the early stages of genome
