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Researchers Design Salmon Louse SNP Chip, ID Genetic Regions Associated with Pesticide Resistance

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NEW YORK (GenomeWeb) – The average sea louse is only about 2 cm in size, but the marine ectoparasites have been causing enormous losses both for salmon farms as well as wild salmon.

According to the Sea Lice Research Center at the University of Bergen in Norway, sea lice each year cause about $377 million in damages to the salmon industry worldwide. And while the introduction of pesticides – especially emamectin benzoate, marketed as SLICE – in 1999 helped salmon farmers to control sea lice infestation for some time, in recent years there has been a resurgence of sea lice that are resistant to SLICE.

To better understand the relationship between sea lice populations at salmon farms across the North Atlantic, as well as to identify genomic regions that could potentially shed light on pesticide resistance, researchers at the Institute for Marine Research in Bergen, the Sea Louse Research Center at the University of Bergen, and the Center for Integrative Genomics (CIGENE) at the Norwegian University of Life Sciences recently designed a 6,000-SNP genotyping microarray focused on Lepeophtheirus salmonis, the salmon louse.

As described in a new BMC Genomics paper, the researchers genotyped nearly 600 sea lice using the array, discovering that sea lice populations from sites in Canada, Ireland, and Norway are largely homogeneous, and also identifying heritable blocks of their genomes that may be playing a role in their resistance to SLICE.

"Sea lice have always been around, but their populations have increased many fold since the salmon industry has expanded," Kevin Glover, population genetics group leader at IMR and the corresponding author on the paper, told BioArray News this week. According to Glover, sea lice are now the "largest challenge to sustainable aquaculture of salmonids in sea cages," and they are also causing high mortality in wild salmon.

Indeed, according to a 2013 Journal of Fish Diseases article, about a third of wild salmon perish each year due to infestation with sea lice.

However, obtaining information on sea lice has been limited to date, with most researchers performing microsatellite genotyping to understand population diversity, Glover said. To create a new genomic tool for sea louse research, the group of investigators recently developed a SNP array with genome-wide coverage.

"We wanted to produce a genomic resource to be able to answer population genetics questions as well as to look at resistance and conduct linkage mapping," Glover said.

To start, Glover and colleagues pooled lice samples from five populations with geographically diverse origins – Canada, the Shetland Islands, Ireland, Northern Norway, and Western Norway – and resequenced those samples several years ago using the Illumina HiSeq platform, identifying about 500,000 SNPs. From that master set of half a million SNPs, they eventually selected 6,000 that "provided good coverage on all of the chromosomes."

The chip was designed in collaboration with CIGENE in Oslo, and the center performed all subsequent genotyping on the array, Glover said. The chip has been manufactured by Affymetrix.

Once the chip's design was finalized, the researchers collected and genotyped 576 samples from 12 sites around the North Atlantic – two in Canada, two in Ireland, two in the Shetland Island, two in the Faroe Islands, and four in Norway – where there had been reports of salmon lice densensitized to SLICE.

They found no relationship between genetic distance and geographic distance. Instead, most of the samples across the site appeared to be under strong selection for certain selective sweeps of the genome – genes and flanking regions passed on from generation to generation – on chromosomes 1, 5, and 14.

"Across the whole Atlantic, lice are being selected strongly because of these selective sweeps," said Glover. And since the main pressure on these lice that could cause such a population bottleneck is the use of pesticides, SLICE in particular, he and the other researchers hypothesized in their BMC Genomics paper that the selective sweeps are related to pesticide resistance, and that lice with these selective sweeps have essentially recolonized the North Atlantic.

Glover noted that SLICE was introduced first in 1999, and seemed to serve as a "wonder drug" up until about 2005, when salmon farmers first began to report the emergence of pesticide-resistant lice. He speculated that it was around this time that sea lice began to be selected strongly for the sweeps on chromosomes 1, 5, and 14 that have led to the emergence of a resistant population.

The ability to design an array for a marine parasite and use it to understand the mechanism underlying resistance, the scale of those genetic mechanisms, and what has caused them in one study is "unprecedented," in Glover's words, and could be replicated in other studies related to other pesticides used to combat sea lice in the industrial setting. Furthermore, the newly gained information on selective sweeps related to pesticide resistance could impact aquaculture practices, Glover stated, including the development of new pesticides.

"This will create a management challenge for aquaculture across the Atlantic," said Glover.

Because the IRM array was funded through the Norwegian Research Council, the array design and data are all public and can be used by researchers anywhere. Several arrays for salmon research already exist, but a number of them are private designs and used by specific companies to improve the breeding of salmon resistant to, among other things, sea lice.

Last year, for example, the Norwegian breeding company AquaGen developed a 923,000-SNP array for use in its internal programs. The chip was developed in collaboration with CIGENE and Affymetrix.

At the same time, it is unlikely that Glover and colleagues will continue to use the 6K sea louse array, as researchers in Canada have designed a higher-density sea louse array with hundreds of thousands of SNPs.

Led by Ben Koop's group at the University of Victoria, the Canadian researchers have developed a 200,000-marker SNP chip for salmon louse. Koop told BioArray News in an email this week that the array is being used to identify population structure in the Atlantic and Pacific Oceans, the impact of infection of salmon farms on wild
salmon, the impact of therapeutics on sea louse control, and the development of a high-density genetic map in order to identify genes related to beneficial phenotypes.

The initial tests of the chip were presented at the 2014 International Sea Lice Conference last summer in Portland, Maine, Koop said. He noted that his lab developed the array with partners at Novartis, the University of Prince Edward Island, Guelph University, and CIGENE.

"That's modern science," said IRM's Glover. "By the time you describe a tool in a publication, it's already obsolete," he said. "We have nevertheless used our newly developed tool to provide unique insights into the population genetic structure and dispersal of genes within the salmon louse in the entire North Atlantic – and were really happy with that."

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