** The title, authors, and abstract for this completion report are provided below.  For a copy of the completion report, please contact the author at [email protected] or via telephone at 204-474-8831. Questions? Contact the GLFC via e-mail or via telephone at 734-662-3209 **

Gene expression differences between feeding types in the paired lampreys Ichthyomyzon unicuspis and I. fossor

Margaret Docker

Department of Biological Sciences, University of Manitoba

Winnipeg, MB, R3T 2N2, Canada

February 2013

Abstract

There is considerable debate regarding the species status (and hence eligibility for conservation) of the silver (Ichthyomyzon unicuspis) and northern brook (I. fossor) lampreys. The lampricide TFM that is used for controlling the invasive sea lamprey (Petromyzon marinus) is also toxic to these native lamprey species. Silver lampreys seem to have been particularly affected by sea lamprey control since their in-stream distribution is very similar to that of the sea lamprey; populations of non-migratory northern brook lampreys also occur in upstream reaches of streams. It may therefore become necessary to reduce TFM applications to streams that contain silver lamprey.

However, many lines of evidence from my laboratory are showing a lack of genetic differentiation between parasitic silver and nonparasitic northern brook lampreys where they co-occur in the Great Lakes, suggesting that they may be a single species with two morphotypes. The larvae of both species are both filter feeders and are morphologically indistinguishable, but they diverge at metamorphosis: the silver lamprey delays sexual maturation until after it completes a one-year adult feeding phase whereas the northern brook lamprey immediately begins sexual maturation and spawns and dies within 6–8 months. An ongoing GLFC-sponsored research project by F. Neave and collaborators (including myself) is investigating the potential environmental triggers that may determine feeding type in these lampreys; the current project investigated the genetic basis of feeding type.

Rather than continue to search for species- or feeding-type-specific differences at the level of the genome (i.e., encoded within the organism’s DNA), this project explored the hypothesis that alternate feeding types are achieved through differences in the timing or level of gene expression when the developmental trajectories of the two feeding types diverge during metamorphosis. Rather than attempt to individually quantify differences in gene expression in a small number of genes that might (based on their purported functions in other organisms) be involved in feeding type differences, we proposed to search for genome-wide differences in expression between these two species during early and late metamorphosis (i.e., prior to morphological differentiation) to identify which genes are differentially expressed in parasitic versus nonparasitic lampreys. This project took advantage of next-generation sequencing (NGS) technologies, specifically transcriptome sequencing (so-called RNA-Seq).

Transcriptomes were assembled de novo for northern brook and chestnut lampreys; we used parasitic chestnut lamprey (Ichthyomyzon castaneus) as a proxy for silver lamprey since metamorphosing silver lamprey were not found in Manitoba (even though feeding-phase adults are frequently encountered). For chestnut lamprey, 112 million sequences were assembled into 33,813 contigs and 147 million northern brook lamprey sequences were assembled into 42,295 contigs. Gene matches were assigned to each contig and annotated with gene ontology terms. Gene expression patterns were compared between early (stage 2) and late (stage 5) metamorphosing chestnut lamprey; 494 and 506 genes, respectively, were overexpressed in each stage relative to the other. Differential gene expression analysis performed between stage 4 northern brook lamprey and stage 5 chestnut lamprey showed that 1496 and 504 genes, respectively, were overexpressed in each species relative to the other. Of particular interest were genes related to developmental processes overexpressed in the parasitic chestnut lamprey and protein metabolism genes overexpressed in the nonparasitic northern brook lamprey; these warrant further investigation. With a more complete sea lamprey genome sequence and assembly now available, our alignments and annotations will likely be improved. Conversely, our study can also provide some assistance in annotating the sea lamprey genome by identifying regions with gene products. This research has not led to any direct applications for Great Lakes fishery management, but will add to the genomic resources now available for lampreys.