1 FASMOP is a national programme, funded by the Scottish Government through RAFTS (Rivers and Fisheries Trusts of Scotland) and
some fisheries trusts, including The Tweed Foundation, to work out the structuring
of Scottish Salmon stocks into distinct breeding populations. This has two
parts, firstly taking samples for DNA analysis from Salmon juveniles at a range
of locations within a catchment as, obviously, those found in any particular
area represent the Salmon that spawned there. Secondly, when such populations
have been identified and are found to be sufficiently distinct, adult Salmon
can be genetically tested to see to which they belong and therefore what sort
of fish (Spring Salmon, Autumn Grilse etc.) derive from each population.
2 The original aim was to use microsatellite DNA
markers (see below), which are highly genetically variable (i.e. have variant
forms) and which give information on ancestry and genetic relatedness, but not
on adaptive genetic differences, as they are chosen to be in what is called
“non-coding DNA”; variation in which is not expressed in the shape or
physiology of an animal and so is not subject to direct natural selection and
evolution. Over time a random evolutionary process called “genetic drift”
causes some of the variants of each microsatellite to be lost at random from
populations causing each population to have unique variant types and
frequencies that give each population a distinct genetic character. This allows
the study of the origins of populations and can also be used to detect the
population of origin of individual fish.
3 Genetic
drift has its greatest effects where populations are small, have existed for a
long time, or where there is little or no mixing of spawning populations. This
is most likely in undisturbed rivers, smaller tributaries where numbers of
breeders are small, and where rivers are broken up by physical obstacles such
as waterfalls or lochs so that their spawning areas are also fragmented. When
microsatellites were used to study the genetics of Salmon across the whole of
Ireland, they successfully showed up population structuring in many areas,
finding differences both between and within rivers. However, they were not able
to show any such structuring for a
group of five rivers in the south-east of Ireland though this was known to
exist in that area as shown by spawning migrations and findings from other
types of genetic markers such as protein coding genes.
4 The
FASMOP programme is therefore exploring a new range of molecular markers which
is likely to include variation in DNA that affects fish form and physiology and
so be subject to natural selection and evolution. These sections of DNA are
called “SNPs” and the Tweed, along with other Scottish rivers is, through
FASMOP, joining in with an existing Marine Scotland collaborative programme with
the Norwegian research group CIGENE, to study the potential for using a set of
SNPs that have been developed to show population structuring where microsatellites
appear not to be able to do so.
5 Over
the summers of 2008 and 2009, 39 of the 44 locations within the Tweed catchment
at which fry samples were planned to be taken have been sampled. Genetic
samples were taken from fifty fry at each location. As each fin clip costs £20
to analyse, a whole location costs £1,000 to process. The locations have been
limited in the first instance and more may be tested and selected dependent on
the initial results obtained. There is no great problem or expense in taking
samples from fry through electro-fishing, but the costs of analyses mean that
the samples processed have to be chosen to be as informative as possible.
6 The
second part of this programme, the taking of tissue samples from adults,
started mid-way through 2008 and so is now in its second full season of
collection. Boatmen at four fisheries on the lower River have been, and are,
collecting tissue samples from fish of different sizes in each month of the
season – the aim is to have samples from all the different 5lb size classes
coming into the River in each month. The reason that samples are being
collected only from the lower River is to ensure that the fish are fresh in
and, ideally, each fish sampled should have Sea-lice or have only recently lost
them. Scales are taken from each fish sampled, so the type of fish (Spring
Salmon, Summer Salmon, etc.) can be determined. There has also been special
permission from the Government for the boatmen at one of these beats to fish in
December and January in both 2008 & 2009 in order to get samples from runs
of fish that take place during the close season.
7 There
has also been – and is - similar structured sampling at one of the netting
stations in the estuary. This sampling of both rod and net fisheries is giving
a comprehensive range of tissue samples from all the different sizes of fish
entering the River throughout the year - and not just in the fishing season.
Each sample again costs £20 to analyse and how many of the samples taken to
be processed will, as with the fry samples, depend on the sort of results
obtained.
8 DNA
from scales is also usable for this sort of genetic analysis and some
laboratories prefer this material to tissue samples – e.g. for the genetics
work on Sea-trout that we are undertaking as part of the Living North Sea programme, the Technical University of Denmark, the partner performing the
analyses, has asked for scales to be taken from the adults sampled. Our
collection of scales from over 20,000 Tweed Salmon, taken since 1992, could
therefore be utilised for this present work as well, if found useful to do so.
Additionally, the Marine Scotland River Laboratory at Montrose has been
obtaining scales from a Tweed netting station one day a month during the
netting season since the 1960’s, providing a further source which could well
give information on changes in population structure over the last 50
years.
DNA is the molecule that carries
inherited characteristics and provides the “plan”, passed on from parents to
offspring, for the development of an individual Salmon egg into a Salmon, just
as in humans. DNA is built up of
four smaller molecules (called “Bases”) designated A, T, C and G, all linked
together. In the Atlantic Salmon of the Tweed, there are 28 pairs of DNA
molecules composed of more than 7 billion of these smaller molecules. In most cases the sequence of these
molecules is the same in all individuals but every so often at a location in
the DNA individuals will differ.
These differences can take different forms.
A MICROSATELLITE is a stretch of DNA where a certain sequence of 2 to 8 sets of bases (e.g. ATCC) are repeated (e.g. ATCC.ATCC.ATCC). This varies both between individuals and between the DNA inherited by an individual from its mother and father in respect of the number of these repeats. So, for example, in an individual the DNA inherited from its mother might have 4 repeats of a microsatellite while in that inherited from its father there might be 7. In another individual, the numbers might be 3 and 5, and in another 4 and 8. When an individual breeds, it will pass one of these variants on to each offspring, which one being a matter of chance. There are tens of thousands of such variable microsatellites in Salmon DNA and this means that each individual will end up having a unique combination of variants that allows its relatedness to other individuals to be assessed and tell whether a group of individuals belongs to a separate breeding population. In practise, however, it is only cost-effective and technically possible at present to study variation in relatively small numbers of microsatellites; the FASMOP programme uses 17. In many situations, this number would be sufficient to assess relatedness and show population structure but, as is clear, in other situations it is not. This situation could in part be addressed by increasing the number of microsatellites screened, but in practise this is not practical or cost-effective and other types of DNA variation have the potential to provide a more practical and informative solution.
A "SNP" (Single-Nucleotide Polymorphism) is a plce on a DNA molecule where there can be different bases (A, T, C, G). As every individual inherits one of its DNA strands from its mother and the other from its father these can also differ from each other as a SNP within the same individual, giving further variation between individuals. As with microsatellite variants therefore, the frequencies and types of these variants can also differ between individuals from different breeding populations. These differences may arise both by genetic drift or due to active selection as some base differences at some locations in the DNA may affect a fishes performance and thus its likelihood of survival and its success at leaving offspring. Where breeding populations live in different environments, some variants may be more successful than others, leading to diiferentiation. By both using larger numbers of SNPs (many tens or hundreds) and including some that show adaptive differences among populations, a higher level of population resolution is expected to be achieved than by a small suite of 17 microsatellites. With the recent advances in DNA technology, it is now possible to explore the potential for using SNPs to study population structuring in Atlantic Salmon.