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PacBio assembly of a Plasmodium knowlesi genome sequence with Hi-C correction and manual annotation of the SICAvar gene family

  • S. A. LAPP (a1), J. A. GERALDO (a2) (a3), J.-T. CHIEN (a1) (a4), F. AY (a5), S. B. PAKALA (a6) (a7), G. BATUGEDARA (a8), J. HUMPHREY (a6) (a7), the MaHPIC consortium (a9), J. D. DeBARRY (a6) (a7), K. G. Le ROCH (a8), M. R. GALINSKI (a1) (a4) (a10) and J. C. KISSINGER (a6) (a7) (a11)...


Plasmodium knowlesi has risen in importance as a zoonotic parasite that has been causing regular episodes of malaria throughout South East Asia. The P. knowlesi genome sequence generated in 2008 highlighted and confirmed many similarities and differences in Plasmodium species, including a global view of several multigene families, such as the large SICAvar multigene family encoding the variant antigens known as the schizont-infected cell agglutination proteins. However, repetitive DNA sequences are the bane of any genome project, and this and other Plasmodium genome projects have not been immune to the gaps, rearrangements and other pitfalls created by these genomic features. Today, long-read PacBio and chromatin conformation technologies are overcoming such obstacles. Here, based on the use of these technologies, we present a highly refined de novo P. knowlesi genome sequence of the Pk1(A+) clone. This sequence and annotation, referred to as the ‘MaHPIC Pk genome sequence’, includes manual annotation of the SICAvar gene family with 136 full-length members categorized as type I or II. This sequence provides a framework that will permit a better understanding of the SICAvar repertoire, selective pressures acting on this gene family and mechanisms of antigenic variation in this species and other pathogens.

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*Corresponding author: Coverdell Center, University of Georgia, 500 D.W. Brooks Drive, Suite 107, Athens, GA 30602, USA. E-mail:


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Ahmed, M. A. and Cox-Singh, J. (2015). Plasmodium knowlesi – an emerging pathogen. ISBT Science Series 10, 134140.
al-Khedery, B., Barnwell, J. W. and Galinski, M. R. (1999). Antigenic variation in malaria: a 3′ genomic alteration associated with the expression of a P. knowlesi variant antigen. Molecular Cell 3, 131141.
Assefa, S., Lim, C., Preston, M. D., Duffy, C. W., Nair, M. B., Adroub, S. A., Kadir, K. A., Goldberg, J. M., Neafsey, D. E., Divis, P., Clark, T. G., Duraisingh, M. T., Conway, D. J., Pain, A. and Singh, B. (2015). Population genomic structure and adaptation in the zoonotic malaria parasite Plasmodium knowlesi . Proceedings of the National Academy of Sciences of the USA 112, 1302713032.
Aurrecoechea, C., Barreto, A., Basenko, E. Y., Brestelli, J., Brunk, B. P., Cade, S., Crouch, K., Doherty, R., Falke, D., Fischer, S., Gajria, B., Harb, O. S., Heiges, M., Hertz-Fowler, C., Hu, S., Iodice, J., Kissinger, J. C., Lawrence, C., Li, W., Pinney, D. F., Pulman, J. A., Roos, D. S., Shanmugasundram, A., Silva-Franco, F., Steinbiss, S., Stoeckert, C. J. Jr., Spruill, D., Wang, H., Warrenfeltz, S. and Zheng, J. (2017). EuPathDB: the eukaryotic pathogen genomics database resource. Nucleic Acids Research 45(D1), D581D591.
Ay, F., Bailey, T. L. and Noble, W. S. (2014a). Statistical confidence estimation for Hi-C data reveals regulatory chromatin contacts. Genome Res 24(6), 9991011. doi: 10.1101/gr.160374.113.
Ay, F., Bunnik, E. M., Varoquaux, N., Bol, S. M., Prudhomme, J., Vert, J. P., Noble, WS. and Le Roch, K. G. (2014b). Three-dimensional modeling of the P. falciparum genome during the erythrocytic cycle reveals a strong connection between genome architecture and gene expression. Genome Res 24, 974–88.
Barber, B. E., Rajahram, G. S., Grigg, M. J., William, T. and Anstey, N. M. (2017). World Malaria Report: time to acknowledge Plasmodium knowlesi malaria. Malaria Journal 16, 135.
Barnwell, J. W., Howard, R. J. and Miller, L. H. (1982). Altered expression of Plasmodium knowlesi variant antigen on the erythrocyte membrane in splenectomized rhesus monkeys. Journal of Immunology 128, 224226.
Barnwell, J. W., Howard, R. J., Coon, H. G. and Miller, L. H. (1983). Splenic requirement for antigenic variation and expression of the variant antigen on the erythrocyte membrane in cloned Plasmodium knowlesi malaria. Infection and Immunity 40, 985994.
Benson, G. (1999). Tandem repeats finder: a program to analyze DNA sequences. Nucleic Acids Research 27, 573580.
Bickhart, D. M., Rosen, B. D., Koren, S., Sayre, B. L., Hastie, A. R., Chan, S., Lee, J., Lam, E. T., Liachko, I., Sullivan, S. T., Burton, J. N., Huson, H. J., Nystrom, J. C., Kelley, C. M., Hutchison, J. L., Zhou, Y., Sun, J., Crisa, A., Ponce de Leon, F. A., Schwartz, J. C., Hammond, J. A., Waldbieser, G. C., Schroeder, S. G., Liu, G. E., Dunham, M. J., Shendure, J., Sonstegard, T. S., Phillippy, A. M., Van Tassell, C. P. and Smith, T. P. (2017). Single-molecule sequencing and chromatin conformation capture enable de novo reference assembly of the domestic goat genome. Nature Genetics 49, 643650.
Brown, K. N. and Brown, I. N. (1965). Immunity to malaria: antigenic variation in chronic infections of Plasmodium knowlesi . Nature 208, 12861288.
Chien, J. T., Pakala, S. B., Geraldo, J. A., Lapp, S. A., Humphrey, J. C., Barnwell, J. W., Kissinger, J. C. and Galinski, M. R. (2016). High-quality genome assembly and annotation for Plasmodium coatneyi, generated using single-molecule real-time PacBio technology. Genome Announcements 4(5), e00883–16.
Chin, W., Contacos, P. G., Coatney, G. R. and Kimball, H. R. (1965). A naturally acquired quotidian-type malaria in man transferable to monkeys. Science 149, 865.
Corredor, V., Meyer, E. V., Lapp, S., Corredor-Medina, C., Huber, C. S., Evans, A. G., Barnwell, J. W. and Galinski, M. R. (2004). A SICAvar switching event in Plasmodium knowlesi is associated with the DNA rearrangement of conserved 3′ non-coding sequences. Molecular and Biochemical Parasitology 138, 3749.
Cox-Singh, J. and Culleton, R. (2015). Plasmodium knowlesi: from severe zoonosis to animal model. Trends in Parasitology 31, 232238.
Duan, Z., Andronescu, M., Schutz, K., McIlwain, S., Kim, Y. J., Lee, C., Shendure, J., Fields, S. and Blau, C. A., Noble, W. S. (2010). A three-dimensional model of the yeast genome. Nature 465(7286), 363–7. doi: 10.1038/nature08973.
Dudchenko, O., Batra, S. S., Omer, A. D., Nyquist, S. K., Hoeger, M., Durand, N. C., Shamim, M. S., Machol, I., Lander, E. S., Aiden, A. P. and Aiden, E. L. (2017). De novo assembly of the Aedes aegypti genome using Hi-C yields chromosome-length scaffolds. Science, doi: 10.1126/science.aal3327.
Emms, D. M. and Kelly, S. (2015). OrthoFinder: solving fundamental biases in whole genome comparisons dramatically improves orthogroup inference accuracy. Genome Biology 16, 157.
Flot, J. F., Marie-Nelly, H. and Koszul, R. (2015). Contact genomics: scaffolding and phasing (meta)genomes using chromosome 3D physical signatures. FEBS Letters 589, 29662974.
Galinski, M. R., Lapp, S. A., Peterson, M. S., Ay, F., Joyner, C. J., Le Roch, K. G., Fonseca, L. L., Voit, E. O. and Consortium, T. M. (2017). Plasmodium knowlesi: a superb in vivo nonhuman primate model of antigenic variation in malaria. Parasitology, doi: 10.1017/S0031182017001135.
Holt, C. and Yandell, M. (2011). MAKER2: an annotation pipeline and genome-database management tool for second-generation genome projects. BMC Bioinformatics 12, 491.
Howard, R. J., Barnwell, J. W. and Kao, V. (1983). Antigenic variation of Plasmodium knowlesi malaria: identification of the variant antigen on infected erythrocytes. Proceedings of the National Academy of Sciences of the USA 80, 41294133.
Hunt, M., Newbold, C., Berriman, M. and Otto, T. D. (2014). A comprehensive evaluation of assembly scaffolding tools. Genome Biology 15, R42.
Imakaev, M., Fudenberg, G., McCord, R. P., Naumova, N., Goloborodko, A., Lajoie, B. R., Dekker, J. and Mirny, L. A. (2012). Iterative correction of Hi-C data reveals hallmarks of chromosome organization. Nature Methods 9, 9991003.
Jones, P., Binns, D., Chang, H. Y., Fraser, M., Li, W., McAnulla, C., McWilliam, H., Maslen, J., Mitchell, A., Nuka, G., Pesseat, S., Quinn, A. F., Sangrador-Vegas, A., Scheremetjew, M., Yong, S. Y., Lopez, R. and Hunter, S. (2014). InterProScan 5: genome-scale protein function classification. Bioinformatics 30, 12361240.
Kaplan, N. and Dekker, J. (2013). High-throughput genome scaffolding from in vivo DNA interaction frequency. Nature Biotechnology 31, 11431147.
Kearse, M., Moir, R., Wilson, A., Stones-Havas, S., Cheung, M., Sturrock, S., Buxton, S., Cooper, A., Markowitz, S., Duran, C., Thierer, T., Ashton, B., Meintjes, P. and Drummond, A. (2012). Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics 28, 16471649.
Korbel, J. O. and Lee, C. (2013). Genome assembly and haplotyping with Hi-C. Nature Biotechnology 31, 10991101.
Korf, I. (2004). Gene finding in novel genomes. BMC Bioinformatics 5, 59.
Kurtz, S., Phillippy, A., Delcher, A. L., Smoot, M., Shumway, M., Antonescu, C. and Salzberg, S. L. (2004). Versatile and open software for comparing large genomes. Genome Biology 5, R12.
Langmead, B. and Salzberg, S. L. (2012). Fast gapped-read alignment with Bowtie 2. Nature Methods 9, 357359.
Lapp, S. A., Korir, C. C. and Galinski, M. R. (2009). Redefining the expressed prototype SICAvar gene involved in Plasmodium knowlesi antigenic variation. Malaria Journal 8, 181.
Lapp, S. A., Korir-Morrison, C., Jiang, J., Bai, Y., Corredor, V. and Galinski, M. R. (2013). Spleen-dependent regulation of antigenic variation in malaria parasites: Plasmodium knowlesi SICAvar expression profiles in splenic and asplenic hosts. PLoS ONE 8, e78014.
Lapp, S. A., Mok, S., Zhu, L., Wu, H., Preiser, P. R., Bozdech, Z. and Galinski, M. R. (2015). Plasmodium knowlesi gene expression differs in ex vivo compared to in vitro blood-stage cultures. Malaria Journal 14, 110.
Logan-Klumpler, F. J., De Silva, N., Boehme, U., Rogers, M. B., Velarde, G., McQuillan, J. A., 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, O. S., Brunk, B. P., Myler, P. J., Roos, D., Carrington, M., Smith, D. F., Hertz-Fowler, C. and Berriman, M. (2012). GeneDB – an annotation database for pathogens. Nucleic Acids Research 40, D98108.
Millar, S. B. and Cox-Singh, J. (2015). Human infections with Plasmodium knowlesi – zoonotic malaria. Clinical Microbiology and Infection 21, 640648.
Moyes, C. L., Henry, A. J., Golding, N., Huang, Z., Singh, B., Baird, J. K., Newton, P. N., Huffman, M., Duda, K. A., Drakeley, C. J., Elyazar, I. R., Anstey, N. M., Chen, Q., Zommers, Z., Bhatt, S., Gething, P. W. and Hay, S. I. (2014). Defining the geographical range of the Plasmodium knowlesi reservoir. PLoS Neglected Tropical Diseases 8, e2780.
Muller, M. and Schlagenhauf, P. (2014). Plasmodium knowlesi in travellers, update 2014. International Journal of Infectious Diseases 22, 5564.
Pain, A., Bohme, U., Berry, A. E., Mungall, K., Finn, R. D., Jackson, A. P., Mourier, T., Mistry, J., Pasini, E. M., Aslett, M. A., Balasubrammaniam, S., Borgwardt, K., Brooks, K., Carret, C., Carver, T. J., Cherevach, I., Chillingworth, T., Clark, T. G., Galinski, M. R., Hall, N., Harper, D., Harris, D., Hauser, H., Ivens, A., Janssen, C. S., Keane, T., Larke, N., Lapp, S., Marti, M., Moule, S., et al. (2008). The genome of the simian and human malaria parasite Plasmodium knowlesi . Nature 455, 799803.
Pasini, E. M., Zeeman, A.-M., Voorberg-Van der Wel, A. and Kocken, C. H. (2017). Parasitology.
Rao, S. S., Huntley, M. H., Durand, N. C., Stamenova, E. K., Bochkov, I. D., Robinson, J. T., Sanborn, A. L., Machol, I., Omer, A. D., Lander, E. S. and Aiden, E. L. (2014). A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping. Cell 159, 16651680.
Rutherford, K., Parkhill, J., Crook, J., Horsnell, T., Rice, P., Rajandream, M. A. and Barrell, B. (2000). Artemis: sequence visualization and annotation. Bioinformatics 16, 944945.
Servant, N., Varoquaux, N., Lajoie, B. R., Viara, E., Chen, C. J., Vert, J. P., Heard, E., Dekker, J. and Barillot, E. (2015). HiC-Pro: an optimized and flexible pipeline for Hi-C data processing. Genome Biology 16, 259.
Shearer, F. M., Huang, Z., Weiss, D. J., Wiebe, A., Gibson, H. S., Battle, K. E., Pigott, D. M., Brady, O. J., Putaporntip, C., Jongwutiwes, S., Lau, Y. L., Manske, M., Amato, R., Elyazar, I. R., Vythilingam, I., Bhatt, S., Gething, P. W., Singh, B., Golding, N., Hay, S. I. and Moyes, C. L. (2016). Estimating geographical variation in the risk of zoonotic Plasmodium knowlesi infection in countries eliminating malaria. PLoS Neglected Tropical Diseases 10, e0004915.
Singh, B. and Daneshvar, C. (2013). Human infections and detection of Plasmodium knowlesi . Clinical Microbiology Review 26, 165184.
Singh, B., Kim Sung, L., Matusop, A., Radhakrishnan, A., Shamsul, S. S., Cox-Singh, J., Thomas, A. and Conway, D. J. (2004). A large focus of naturally acquired Plasmodium knowlesi infections in human beings. Lancet 363, 10171024.
Soderlund, C., Bomhoff, M. and Nelson, W. M. (2011). SyMAP v3.4: a turnkey synteny system with application to plant genomes. Nucleic Acids Research 39, e68.
Standage, D. S. and Brendel, V. P. (2012). ParsEval: parallel comparison and analysis of gene structure annotations. BMC Bioinformatics 13, 187.
Stanke, M., Diekhans, M., Baertsch, R. and Haussler, D. (2008). Using native and syntenically mapped cDNA alignments to improve de novo gene finding. Bioinformatics 24, 637644.
Steinbiss, S., Silva-Franco, F., Brunk, B., Foth, B., Hertz-Fowler, C., Berriman, M. and Otto, T. D. (2016). Companion: a web server for annotation and analysis of parasite genomes. Nucleic Acids Research 44, W29W34.
Wesolowski, R., Wozniak, A., Mila-Kierzenkowska, C. and Szewczyk-Golec, K. (2015). Plasmodium knowlesi as a Threat to Global Public Health. Korean Journal of Parasitology 53, 575581.
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