Hi-C: Zika mosquito genome assembled via breakthrough method

yee Mike Yee

As recently reported in Nature News, the genome of the Zika mosquito was mapped and published in Science, which details the breakthrough method used to assemble known sequences. Zika infection in pregnant women can lead to major birth defects for babies. As outlined by the CDC, these defects include microcephaly, decreased brain tissue, and defects involving the eyes, joints, and muscles. The Zika virus was first isolated in 1947, and its entire genome was sequenced in 2007. The virus, passed by a bite mainly from the mosquito species Aedes aegypti (Ae. aegypti) and Aedes albopictus, has spread at an alarming rate, and was headed to the United States. To fight the ongoing spread, scientists have been working to form a more complete map of the genome for the Aedes aegypti mosquito. Parts of the genome were already known and published as a draft in 2007, but researchers were having difficulties fully piecing it together.

Ten years after that initial publication of the draft sequences, the aforementioned Science article by Dudchenko et al. details how they used a technique called Hi-C to map the genome of Ae. aegypti. Using this method, they were able to put together Hi-C data with assemblies already known to generate chromosome-length scaffolds. This was validated by assembling the human genome with 99-percent accuracy. Then they created genome assemblies for the mosquito species Ae. aegypti and Culex quinquefasciatus. Culex quinquefasciatus is the mosquito species notorious for spreading West Nile virus, so this research impacts not only the fight against Zika, but West Nile disease as well. Each assembly contained three scaffolds—one for each of the three chromosomes per species. Dudchenko et al. note that genomic rearrangements occurred within chromosome arms instead of between chromosomal arms. Furthermore, the price was relatively low, with each genome analysis costing less than $10,000.

Hi-C works via an analysis of chromosome folding. These folds were mapped, and frequency of physical interactions between parts of the genome were statistically analyzed. Then, using this Hi-C data, Dudchenko et al were able to piece together the location of genome fragments. The principles behind this technique are described in this 2013 Nature article. The key principle is proximity ligation, where DNA that is near each other is preferentially ligated together upon collection from cells. Researchers leveraged this by crosslinking and digesting the DNA with a restriction enzyme. Then DNA fragments were biotinylated and ligated, and the fragments were sheared, purified, and put through pair-end sequencing. Researchers were then able to order and orient contigs to generate de novo assemblies and perform haplotype analyses. Specifically, LACHESIS was used to combine Hi-C data with sequencing data to generate the de novo assemblies. Here, HaploSeq was used to determine connectivity between linked blocks that likely match the Hi-C data, and this was done to determine chromosome-scale haplotypes.

With 94 percent of the genomes now mapped, scientists can continue conducting further research into understanding how the genome of the mosquito species influences specific phenotypes. Then, further down the line, researchers can better understand how to target the mosquito directly to test ways of stopping the spread of deadly viruses such as Zika and West Nile. As mentioned in this recent Nature News article, the University of Notre Dame’s David Severson says the efforts by the research team are “phenomenal,” but much work still needs to be done, with millions of base pairs still missing and “ghost genes” still yet to be researched.


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mikecyee

mikecyee

Mike has a Ph.D. in Biomedical Sciences from the University of California, Riverside, a M.S. in Cell and Molecular Biology from San Francisco State University, and a B.A. in English from the University of California, Berkeley.