Using CoGe for the analysis of Plasmodium spp

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About this guide

This 'cookbook' style document is meant to provide an introduction to many of our tools and services and is structured around a case study of investigating genome evolution of the malaria-causing Plasmodium spp. The small size and unique features of this pathogen's genome make it ideal for beginning to understand how our tools can be used to conduct comparative genomic analyses and uncover meaningful discoveries.

Through a number of example analyses, this guide will teach users about the following tools:

  • LoadGenome: Add a new genome to CoGe.
  • LoadAnnotation: Add structural and/or functional annotations to a genome.
  • GenomeInfo: Get information about a genome.
  • GenomeList: Get information about several genomes in a table.
  • CoGeBLAST: BLAST against any set of genomes.
  • GEvo: Microsynteny analysis.
  • SynMap: Whole genome syntenic analysis.
- SynMap#Calculating and displaying synonymous/non-synonymous (Ks, Kn), data Kn/Ks Analysis: Characterize the evolution of populations of genes.
- SPA tool: Syntenic Path Assembly to assist in genome analysis.
  • SynFind: Identify syntenic genes across multiple genomes.
  • CodeOn: Characterize patterns of codon and amino acid evolution in coding sequence.

A brief introduction to Plasmodium genome evolution

The study of parasitic genomes via comparative genomics offers many unique challenges. Parasite genomes are characterized by a combination of gene loss and the acquisition of species- or lineage-specific genes; in particular, many specialized genes mediate host–parasite interaction [1]. The dynamic nature of parasitic genomes is particularly evident within the genus Plasmodium. The genus emerged ~40 million years ago and harbors roughly 200 species of parasitic protozoa better known as malaria parasites. All Plasmodium species have a complex life cycle involving some kind of vertebrate host and a mosquito vector of the genus Anopheles (mammals) or Culex (birds). In addition, Plasmodium species share similar life cycle characteristics, albeit with a few exceptions (e.g. hypnozoites). However, host and vector preferences differ among Plasmodium species [2].

Plasmodium genomes are tiny (between 17-28Mb) in comparison to those of their vertebrate (1Gb for birds; 2-3Gb for mammals) and mosquito (230–284Mbp) hosts [3]. All Plasmodium genomes consist of fourteen chromosomes (nuclear genome), as well as a mitochondrial and apicoplast genome. Despite these shared genomic characteristics, the structural organization, gene content, and sequence of Plasmodium genomes is highly variably within the genus [4]. The exact origins and mechanisms of these differences remain largely unexplored, however, they are generally hypothesized to stem from host shift events [5][6].

An increase in funding devoted to malaria research has coincided with a dramatic increase in publicly available genomic information for Plasmodium [7]. The most prominent repository is found at NCBI/Genbank [8]; while additional and unique sequences can also be found on other databases: PlasmoDB [9], GeneDB [10], and MalAvi [11]. This wealth of genomic data facilitates detailed comparative genomic approaches, opening the possibility to:

  • Infer origins of certain traits, specialized phenotypes, and genomic features.
  • Track the maintenance of conserved genes across the genus, as well as the gain or loss of genes unique to a single species or a group of closely related species.
  • Identify the potential historical interactions that might have lead to the development of genomic adaptations.

Through a case study on Plasmodium evolution, we will illustrate how CoGe can be used for the analysis of multigene families, local synteny, and whole genome comparisons (genome composition, rearrangement events, and gene order conservation).

Finding and integrating Plasmodium genomes in CoGe

You can find the details of Plasmodium genome integration in the following link: Finding and intregating Plasmodium genomes to CoGe

Using CoGe tools to perform comparative analyses

Workflows

The following links direct to specific tools for the comparative analysis of Plasmodium genomes:

Plasmodium analysis workflow 1: Tools that evaluate genomic properties and amino acid usage

Plasmodium analysis workflow 2: Tools for the syntenic analysis of whole genomes and microsyntenic regions

Plasmodium analysis workflow 3: Tools useful on the study of multigene families

Additional tools for genome analysis with CoGe

You can learn about the SPA usage on Plasmodium genomes in the following link: Plasmodium genome analysis using Syntenic Path Assembly

Overall conclusions

The number of available Plasmodium genomes has increased considerably during recent years. This wealth of genomic information creates an unprecedented opportunity to study the unique genomic qualities of this genus using comparative genomics.

There have been tremendous achievements in malaria treatment and control strategies. Thanks to worldwide efforts, there has been a significant reduction in the number of malaria cases and malaria-related deaths between 2000 and 2015. By 2015, it was estimated that the number of malaria cases decreased from 262 million to 214 million, and the number of malaria-related deaths from 839,000 to 438,000 [12]. However, there are still numerous aspects of malaria research that need to be further addressed.

The intricacies of parasite-host relations in Plasmodium infection might be more complex than previously considered [13]. Humans have recently been infected by Plasmodium species classically considered specific to non-human primates (e.g. a single infection with P. cynomolgi [14] and various infections with P. knowlesi [15]). In addition, african primates have been infected by unique P. falciparum strains (a parasite classically considered exclusive to humans) and are proposed to act as reservoirs for this parasite [16][17]. In bird Plasmodium, the putative evolutionary time of parasite-host associations has a significant role in the development of pathogenicity and in host mortality [18]. Finally, multiple host-switch events between largely divergent host types are thought to have occurred in bat Haemosporidia [19]. These cases highlight the complexity of the Plasmodium infection landscape. Insights into the unique patterns of Plasmodium biology, epidemiology, ecology, and genetics can be obtained from molecular and comparative genomic studies.

The rapid growth of genomic information makes implementing tools that facilitate assessing genome evolutionary trends an imperative task. The services and tools provided by the CoGe platform are of considerable use in advancing Plasmodium comparative genomics. Here, we showed how various CoGe tools could be used to assess evolutionary patterns unique to Plasmodium. We also showed how to use this platform to further characterize sequenced Plasmodium genomes. Overall, we have demonstrated that CoGe’s tools can be used to address evolutionary questions such as:

  • The evolutionary origins of Laveranian AT-rich genomes.
  • The location and nature of genome rearrangements between Plasmodium.
  • The evolutionary patterns of genes crucial in cell invasion.
  • The evolutionary trends of multigene families.

Useful links

Plasmodium Notebooks in CoGe

Link to Notebook for published Plasmodium genome data: https://genomevolution.org/coge/NotebookView.pl?lid=1753
Link to Notebook for published P. falciparum strains: https://genomevolution.org/coge/NotebookView.pl?lid=1758
Link to Notebook for published P. vivax strains: https://genomevolution.org/coge/NotebookView.pl?lid=1760
Link to Notebook for published Plasmodium apicoplast data: https://genomevolution.org/coge/NotebookView.pl?lid=1754
Link to Notebook for published Plasmodium mitochondrion data: https://genomevolution.org/coge/NotebookView.pl?lid=1756

Sample data

  • Gene sequences used on CoGeBLAST analysis (obtained from PlasmoDB):
PVX_113230.1 | Plasmodium vivax Sal-1 | variable surface protein Vir14-related (http://plasmodb.org/plasmo/app/record/gene/PVX_113230)
PVX_096004.1 | Plasmodium vivax Sal-1 | VIR protein (http://plasmodb.org/plasmo/app/record/gene/PVX_096004)
  • Gene sequence used on SynFind to inform GEvo analysis (obtained from PlasmoDB):
PVX_003830.1 | Plasmodium vivax Sal-1 | serine-repeat antigen 5 (SERA) (http://plasmodb.org/plasmo/app/record/gene/PVX_003830)
  • Gene sequences used on CoGeBLAST to inform GEvo analysis (obtained from PlasmoDB):
PF3D7_0424100.1 | Plasmodium falciparum 3D7 | reticulocyte binding protein homologue 5 (http://plasmodb.org/plasmo/app/record/gene/PF3D7_0424100)
PVX_096410.1 | Plasmodium vivax Sal-1 | cysteine repeat modular protein 2, putative (http://plasmodb.org/plasmo/app/record/gene/PVX_096410)


References

  1. Jackson AP. 2015. Preface. The evolution of parasite genomes and the origins of parasitism. Parasitology. 142 Suppl 1:S1-5. https://www.ncbi.nlm.nih.gov/pubmed/25656359
  2. Sinka ME, Bangs MJ, Manguin S, Rubio-Palis Y, Chareonviriyaphap T, Coetzee M, Mbogo CM, Hemingway J, Patil AP, Temperley WH, Gething PW, Kabaria CW, Burkot TR, Harbach RE, Hay SI. 2012. A global map of dominant malaria vectors. Parasit Vectors. 5:69. https://www.ncbi.nlm.nih.gov/pubmed/22475528
  3. DeBarry JD, Kissinger JC. 2011. Jumbled Genomes: Missing Apicomplexan Synteny. Mol Biol Evol. 2011 Oct; 28(10): 2855–2871. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3176833/
  4. Carlton JM, Perkins SL, Deitsch KW. 2013. Malaria Parasites. Caister Academic Press
  5. Prugnolle F, Durand P, Ollomo B, Duval L, Ariey F, Arnathau C, Gonzalez JP, Leroy E, Renaud F. 2011. A Fresh Look at the Origin of Plasmodium falciparum, the Most Malignant Malaria Agent. PLoS Pathog. 7: e1001283. http://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1001283
  6. Prugnolle F, Rougeron V, Becquart P, Berry A, Makanga B, Rahola N, Arnathau C, Ngoubangoye B, Menard S, Willaume E, Ayala FJ, Fontenille D, Ollomo B, Durand P, Paupy C, Renaud F. 2013. Diversity, host switching and evolution of Plasmodium vivax infecting African great apes. Proc Natl Acad Sci U S A. 110:8123-8. https://www.ncbi.nlm.nih.gov/pubmed/23637341
  7. Buscaglia CA, Kissinger JC, Agüero F. 2015. Neglected Tropical Diseases in the Post-Genomic Era. Trends Genet. 31:539-55. https://www.ncbi.nlm.nih.gov/pubmed/26450337
  8. Clark K, Karsch-Mizrachi I, Lipman DJ, Ostell J, Sayers EW. 2016. GenBank. Nucleic Acids Res. 44: D67–D72. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4702903/
  9. Aurrecoechea C, Brestelli J, Brunk BP, Dommer J, Fischer S, Gajria B, Gao X, Gingle A, Grant G, Harb OS, Heiges M, Innamorato F, Iodice J, Kissinger JC, Kraemer E, Li W, Miller JA, Nayak V, Pennington C, Pinney DF, Roos DS, Ross C, Stoeckert CJ Jr, Treatman C, Wang H. 2009. PlasmoDB: a functional genomic database for malaria parasites. Nucleic Acids Res. 37:D539-43. https://www.ncbi.nlm.nih.gov/pubmed/18957442
  10. Logan-Klumpler FJ, De Silva N, Boehme U, Rogers MB, Velarde G, McQuillan JA, Carver T, Aslett M, Olsen C, Subramanian S, Phan I, Farris C, Mitra S, Ramasamy G, Wang H, Tivey A, Jackson A, Houston R, Parkhill J, Holden M, Harb OS, Brunk BP, Myler PJ, Roos D, Carrington M, Smith DF, Hertz-Fowler C, Berriman M. 2012. GeneDB--an annotation database for pathogens. Nucleic Acids Res. 40:D98-108. https://www.ncbi.nlm.nih.gov/pubmed/22116062
  11. Bensch S, Hellgren O, Pérez-Tris J. 2009. MalAvi: a public database of malaria parasites and related haemosporidian in avian hosts based on mitochondrial cytochrome b lineages. Mol Ecol Resour. 9:1353-8. https://www.ncbi.nlm.nih.gov/pubmed/21564906
  12. World Health Organization. (2015). World Malaria Report 2015. Retrieved from http://www.who.int/malaria/publications/world-malaria-report-2015/report/en/
  13. Garamszegi LZ. 2009. Patterns of co-speciation and host switching in primate malaria parasites. Malar J. 110. doi: 10.1186/1475-2875-8-110. https://www.ncbi.nlm.nih.gov/pubmed/19463162
  14. Ta TH, Hisam S, Lanza M, Jiram AI, Ismail N, Rubio JM. 2014. First case of a naturally acquired human infection with Plasmodium cynomolgi. Malar J. 13: 68. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3937822/
  15. Singh B, Daneshvar C. 2013. Human infections and detection of Plasmodium knowlesi. Clin Microbiol Rev. 26:165-84. https://www.ncbi.nlm.nih.gov/pubmed/23554413
  16. Prugnolle F, Durand P, Neel C, Ollomo B, Ayala FJ, Arnathau C, Etienne L, Mpoudi-Ngole E, Nkoghe D, Leroy E, Delaporte E, Peeters M, Renaud F. 2010. African great apes are natural hosts of multiple related malaria species, including Plasmodium falciparum. Proc Natl Acad Sci U S A. 107:1458-63. https://www.ncbi.nlm.nih.gov/pubmed/20133889
  17. Duval L, Fourment M, Nerrienet E, Rousset D, Sadeuh SA, Goodman SM, Andriaholinirina NV, Randrianarivelojosia M, Paul RE, Robert V, Ayala FJ, Ariey F. 2010. African apes as reservoirs of Plasmodium falciparum and the origin and diversification of the Laverania subgenus. Proc Natl Acad Sci U S A. 107:10561-6. https://www.ncbi.nlm.nih.gov/pubmed/20498054
  18. Krizanauskiene A, Hellgren O, Kosarev V, Sokolov L, Bensch S, Valkiunas G. 2006. Variation in host specificity between species of avian haemosporidian parasites: evidence from parasite morphology and cytochrome B gene sequences. J Parasitol. 6:1319-24. https://www.ncbi.nlm.nih.gov/pubmed/17304814
  19. Duval L, Robert V, Csorba G, Hassanin A, Randrianarivelojosia M, Walston J, Nhim T, Goodman SM, Ariey F. 2007. Multiple host-switching of Haemosporidia parasites in bats. Malar J. 6:157. https://www.ncbi.nlm.nih.gov/pubmed/18045505