The joint policy of the U.S. Fish and Wildlife Service and the National Marine Fisheries Service provides guidelines for defining distinct population segments below the taxonomic level of species (USFWS-NMFS 1996). The first of two elements to be considered is the discreteness of a population segment with respect to the rest of the populations within the species. Discreteness may result from physical factors that isolate the population segment and may be reflected as life-history differences in physiology, ecology, or behavior between the DPS and other populations. Genetic or morphological differences between the population segment being considered, and other populations, may also be used to evaluate discreteness. The policy also states that international boundaries within the geographical range of the species may be used to delimit a distinct population segment in the United States. This criterion is applicable if differences in the control of exploitation of the species, the management of the species' habitat, the conservation status of the species, or regulatory mechanisms differ between countries that would influence the conservation status of the population segment in the United States. In past assessments of evolutionarily significant units (ESUs) in Pacific salmon, however, NMFS has placed the emphasis on biological information in defining DPSs and has considered political boundaries only at the implementation of ESA listings.
A second element in defining distinct population segments is that the segment must be biologically or ecologically significant. Significance is evaluated in terms of the importance of the population segment to the overall welfare of the species and may be considered in the light of, but not limited to, the following factors. The population segment may be considered significant if it persists in an unusual or unique ecological setting for the species. A population segment may also be considered significant, if its loss would result in a significant gap in the geographical range of the species. Such a gap may disrupt the normal connectivity between populations. A segment also meets the significance guideline, if it represents the only surviving natural occurrence of the species that may be more abundant elsewhere as an introduced population outside its historical range. Another guideline is that the population segment differs markedly in its genetic characteristics from other populations of the species. Genetic differences may be detected by molecular genetic methods or may be reflected in unique adaptations to habitats not found in other parts of the species' geographical range. Other classes of information may also bear on the biological or ecological importance of a distinct population segment.
We considered several kinds of information in this status review to attempt to delineate DPSs of Pacific hake, Pacific cod, and walleye pollock in Puget Sound. The first kind of information was habitat characteristics that might indicate the population segment occupies an unusual or unique ecological setting for the species as a whole. The second kind of information was to consider geographical variability in phenotypic and life-history traits that may reflect local adaptation. Such traits may have an underlying genetic basis, but are often strongly influenced by environmental factors from one locality to another. The third kind of information consisted of mark-recapture studies, which give insight into the physical movement of individuals between areas. The fourth kind of information consisted of traits that are inherited in a predictable way and remain unchanged throughout the life of an individual. Differences among populations in the frequencies of these genetically-determined traits may reflect isolation between the populations.
In order to properly evaluate data and information on fish populations
in Puget Sound, similar information was gathered for each biological species,
as a whole, to provide a context in which to determine whether Puget Sound
populations are distinct. Therefore,
reference data were gathered on Pacific hake, Pacific cod, and walleye pollock
from throughout their ranges. Not
all types of biological data were available for each of the three species
under review but an effort was made to utilize the best scientific information
available at the time. The kinds
of information analyzed and the relative usefulness of this information in
delineating DPSs for marine fish in general are discussed briefly in the
following sections.
The analysis of habitat characteristics may indicate that a population segment occupies an unusual or distinctive habitat, relative to the biological species as a whole. The persistence of a discrete population segment in an ecological setting unusual or unique for the taxon is one factor identified in the joint DPS policy (USFWS-NMFS 1996) that may provide evidence of the population's significance. However, Waples (1991a, p. 15) cautioned against "drawing inferences based on physical characteristics of the habitat without supporting biological information linking the habitat differences to adaptations."
Conversely, the continuous distribution of a population segment within a region possessing similar habitat and ecological characteristics makes it less likely that unique adaptations have arisen in local populations. Without associated compelling phenetic or genetic evidence for a finer population structure, marine fish continuously distributed throughout similar habitat and lacking physical or behavioral barriers to migration are not likely to be composed of multiple DPSs.
Isolation between populations may be reflected in several life history variables, including differences in behavior (e.g., spawning timing, migration) and demography (e.g., growth rate, fecundity, age structure), among others. Although some of these traits may have a broad genetic basis and may reflect local adaptations of evolutionary importance, they are usually strongly influenced by environmental factors over the life time of an individual or over a few generations. Differences can arise among populations in response to environmental variability among areas and they can sometimes be used to infer the degree of independence among populations. However, differences in phenotypic and life-history traits among populations do not provide definitive information on reproductive isolation between populations, because the genetic basis of many phenotypic and life-history traits is weak or unknown. Likewise, elemental profiles present in otoliths, and other structures, reflect local environmental conditions or diets and although they may indicate that different areas or environments are occupied, they also provide little definitive information on the degree of reproductive isolation between populations.
Variation in reproductive behavior within a species of marine fish is an important factor to consider because it may provide the isolating mechanism required for differentiation. The presence of geographically-discrete and temporally-persistent spawning aggregations in a species indicates that reproductive isolation may be occurring. However, it is necessary to evaluate the degree of reproductive isolation by addressing the questions of migration rate, gene flow, and recolonization rate. These later considerations are dependent on the degree of homing ability and natal-site fidelity of adults.
Studies of parasite incidence can provide important information about the degree of intermingling of marine fish stocks, particularly when a parasite is present in one area and totally absent in an adjoining area. However, parasite studies have some inherent interpretation problems: 1) in most cases parasite incidences exhibit clinal trends with latitude, and the degree to which parasite occurrence is due to environmental differences, acting on the parasite, or to a lack of host stock intermingling, is unknown, 2) the lack of a parasite in an area may be due to a regional absence of an alternate host organism, independent of host distribution, and 3) parasites may not be permanent natural tags in that parasites may be lost during the lifetime of the host.
The analysis of applied or acquired tags can indicate the degree of migration between localities. These tags consist of physical tags that are attached to a fish and later recovered. These tags provide evidence of movement of individuals from one place to another, but not necessarily of population connectivity through gene flow. Since these kinds of population markers largely lack a genetic basis and are not inherited, they must be applied each generation or must arise naturally anew each generation.
The application and recovery of physical tags on adult marine fish on spawning grounds can answer the question of whether fish return to the same locality to spawn in subsequent years, but these studies lack the direct evidence of parent-offspring linkage. In other words, these studies do not provide direct evidence that fish return to their natal area for spawning; however, they may provide evidence of straying and thus, the potential for gene flow between spawning aggregations.
Two problems inherent in the use of morphometric and meristic characteristics to separate marine fish populations are that: 1) the characteristics are often under strong environmental influence and are not inherited in a simple Mendelian fashion, and 2) the characteristics are continuously variable and exhibit clinal trends and a high variance about the mean. A further drawback of using morphometric and meristic characteristics to detect population structure in fish is that few of these characteristics have been examined from a genetic standpoint. As shown by studies on several species, environmental parameters such as temperature, salinity, pH, and oxygen concentration can modify the expression of genes responsible for meristic characters (see references in Ihssen et al. 1981).
The BRT considered molecular genetic evidence that might be used to define reproductively isolated populations or groups of populations of Pacific hake, Pacific cod, and walleye pollock in Puget Sound, as well as throughout their respective ranges. Molecular genetic markers appear to be largely unaffected by natural selection, so that geographical differences in gene frequencies can be interpreted in terms of genetic flow and genetic drift. The analysis of the geographical distributions of these markers may reveal historical dispersals, equilibrium levels of migration (gene flow), and past isolation. Evidence for genetic population structure is based on the analysis of protein variants (allozymes), microsatellite loci (variable numbers of short tandem DNA repeats), and mitochondrial DNA (mtDNA).
Evidence of substantial genetic divergence between populations, as shown through analysis of these neutral molecular markers, is an important aspect of distinctiveness because even a small amount of interbreeding between populations will reduce the genetic differentiation between them. Although these molecular genetic methods "provide valuable insight into the process of genetic differentiation among populations" they offer "little direct information regarding the extent of adaptive genetic differences" (Waples 1995).
One widely used method of population analysis is sequence or RFLP (restriction fragment length polymorphism) analysis of mtDNA, which codes for several genes that are not found in the cell nucleus. Mitochondrial DNA differs from nuclear DNA (nDNA) in two important ways. One way is that recombination is lacking in mtDNA, so that gene combinations (haplotypes) are passed unaltered from one generation to the next, except for new mutations. A second way is that mtDNA is inherited from only the maternal parent in most fishes, so that gene phylogenies correspond to female lineages. A greater amount of random genetic drift among populations is expected for mtDNA genes, because the effective population size for mtDNA is about one-fourth of that for nuclear genes. These characteristics permit phylogeographical analyses of mtDNA haplotypes, which can potentially indicate dispersal pathways for females and the extent of gene flow between populations (Avise et al. 1987).
Microsatellite DNA markers can potentially detect stock structure on finer spatial and temporal scales than can other DNA or protein markers, because of higher levels of polymorphism found in microsatellite DNA (reflecting a high mutation rate). When populations are at least partially isolated, genetic markers at loci with high mutation rates may accumulate more rapidly in some areas than in others.
Several standard statistical methods have been used to analyze molecular genetic data to detect reproductive isolation between populations. Comparisons of genotypic frequencies in a sample with frequencies expected under random mating (Hardy-Weinberg proportions) may be used to infer the breeding structure of a population or to detect population mixing (Wahlund's effect). Contingency-table comparisons of allozyme or microsatellite allele frequencies among population samples with chi-square or G (log-likelihood ratio) test statistics, or with randomization tests, can be used to detect significant differences between populations, which may be evidence of reproductive isolation.
A complementary way of assessing genetic isolation between populations is to analyze genetic distances based on allele-frequency estimates. Several genetic distance measures (e.g., Cavalli-Sforza and Edwards 1967; Nei 1972, 1978) have been used to study the population genetic structure of anadromous salmonids. It is unclear, however, which measure is most appropriate in a particular case or whether there is one measure that is always most appropriate. Discussions of the features of genetic distances appear in Nei (1978), Rogers (1991), and Hillis et al. (1996). Most of this discussion has focused on the merits of the various measures for phylogenetic reconstruction among species or higher taxa.
Sample sizes and heterozygosity may also influence the power of the genetic distance approach to resolving genetic population structure. When sample sizes used to estimate allelic frequencies are 50 individuals or more, the difference between Nei's genetic distance, D, (Nei 1972) and Nei's unbiased genetic distance (Nei 1978) is small in absolute terms, but still might be a substantial proportion of D, if D is small. When genetic distances between populations are also small, as they often are between populations of marine fishes, low but significant levels of genetic differentiation may not be detected by an unbiased distance measure because sample size corrections may reduce estimates of genetic distance to zero. These measures range from 0.0 (identity) to infinity (complete dissimilarity). In most cases, the different genetic-distance measures yield highly-correlated results.
The degree of reproductive isolation between populations can be inferred from an analysis of the pattern of genetic distances between populations. Clustering methods, such as the unweighted pair group method with averages (UPGMA, Sneath and Sokal 1973) and the neighbor-joining method (Saitou and Nei 1987), find hierarchical groupings of genetically similar populations. Multivariate methods, such as multidimensional scaling (MDS, Kruskal 1964) or principal components analysis (PCA), find groupings of genetically-similar populations in several dimensions, which are depicted here in two or three dimensions.
Various studies have estimated levels of genetic variability within populations, because the level of within-population variability may reflect evolutionary or historical differences in population size and in migration patterns between populations. Within-population gene diversity was measured by the expected proportion of heterozygous genotypes in a population of randomly mating individuals averaged over the number of loci examined (H). Estimates of H based on a small number of individuals are usually accurate, as long as a large number of loci (>30) are surveyed for variability (Nei 1978).
Genetic differentiation between populations at various hierarchical levels has been estimated in many studies with a gene diversity analysis (Nei 1973, Chakraborty 1980), which apportions allele-frequency variability among populations into its geographical or ecological components. For example, the proportion of the total genetic variability in a set of samples that is due to differences among populations may be estimated with FST or the multiallelic equivalent statistics, GST. These variables range from 0.0 (no difference among populations) to 1.0 (fixed allele-frequency differences). The range 0.05-0.15 for FST indicates moderate differentiation, and the range 0.15-0.25 indicates strong genetic differentiation among populations (Wright 1978). These statistics facilitate comparisons among groups of populations that may reveal regional differences in gene flow between populations.
The term "stock" has been used rather loosely in fisheries management and no single definition has been accepted by all fisheries biologists. Stock may be used to refer to groups of fish being harvested in a particular area, whether these fish are genetically related or not. However, in most cases, identification of a group of fish as a stock implies that these fish are in some way different or distinct from those in another stock, and generally implies some genetic relatedness among its members (Ihssen et al. 1981). Evidence of stock structure may be shown through differences in demographic population statistics (age composition, growth rate, fecundity, etc.), morphology (morphometrics and meristics), or genetics (differentiation at allozyme or DNA loci).
Ricker (1972, p. 28) defined a salmon stock as "the fish spawning in a particular lake or stream (or portion of it) at a particular season, which fish to a substantial degree do not interbreed with any group spawning in a different place, or in the same place at a different season." Larkin (1972) defined a stock as "a population of organisms, which, sharing a common environment and participating in a common gene pool, is sufficiently discrete to warrant consideration as a self-perpetuating system which can be managed." Booke (1981) provided a general definition of a stock as "a species group, or population, of fish that maintains and sustains itself over time in a definable area." Ihssen et al. (1981) defined a stock as "an intraspecific group of randomly mating individuals with temporal or spatial integrity." In none of these definitions is it implied that a fish stock is ecologically or biologically significant in relation to the biological species as a whole.
By contrast, not only must a marine fish DPS be "markedly separated from other populations of the same taxon," it must also exhibit ecological or biological significance in comparison to other population segments of the biological species. Thus, following the guidance supplied by the joint policy statement (USFWS-NMFS 1996), a distinct population segment of marine fish may be viewed as a group of related stocks (or in some cases, if the evidence warrants, a single stock) that form(s) a discrete population and are(is) significant to the biological species as a whole. As stated previously, considerations that can be used to determine a discrete population's significance to the taxon as a whole include: 1) persistence of the population segment in an ecological setting unusual or unique for the taxon, 2) evidence that loss of the population segment would result in a significant gap in the range of the taxon, 3) evidence that the population segment represents the only surviving natural occurrence of a taxon that may be more abundant elsewhere as an introduced population outside its historic range, and 4) evidence that the population segment differs markedly from other populations of the species in its genetic characteristics.
The ESA (Section 3) defines "endangered species" as "any species which is in danger of extinction throughout all or a significant portion of its range." "Threatened species" is defined as "any species which is likely to become an endangered species within the foreseeable future throughout all or a significant portion of its range." NMFS considers a variety of information in evaluating the level of risk faced by a DPS, including: 1) absolute numbers of fish and their spatial and temporal distributions, 2) current abundance in relation to historical abundance and carrying capacity of the habitat, 3) trends in abundance, based on indices such as catch statistics, CPUE, and spawner-recruit ratios, 4) natural and human-influenced factors that cause variability in survival and abundance, 5) possible threats to genetic integrity (e.g., selective fisheries and interactions between cultured and natural populations), and 6) recent events (e.g., climate change and changes in management) that have predictable short-term consequences for the abundance of a DPS. Additional risk factors, such as disease prevalence or changes in life-history traits, also may be considered in the evaluation of risk to a population.
The determination of whether a species is threatened or endangered, according to the ESA, should be based on the best scientific information available, after taking into consideration conservation measures that are proposed or in place. The BRT did not evaluate likely or possible effects of conservation measures. Therefore, they did not make recommendations on whether DPSs should be listed as threatened or endangered species, because that determination requires evaluation of factors not considered by the BRT. However, the BRT did draw scientific conclusions about the risk of extinction faced by DPSs, under the assumption that present conditions will continue, and recognizing that natural demographical and environmental variability is an inherent feature of present conditions. Conservation measures will be taken into account by the NMFS's Northwest Regional Office in making listing recommendations. The following sections summarize the kinds of information the BRT considered in evaluating the potential effects of risk factors on the each of the DPSs identified by the BRT.
The absolute number of individuals in a population is important in assessing two aspects of extinction risk. First, population sizes of small populations that are stable or increasing can be an indicator of whether the population can sustain itself in the face of environmental fluctuations and small-population stochasticity. This conclusion follows from the theory of minimum viable populations (see Gilpin and Soulé 1986, Thompson 1991). Second, present abundance in a declining population is an indicator of the time expected until the population reaches critically low numbers. This follows from the idea of "driven extinction" (Caughley 1994). In addition to absolute numbers, the spatial and temporal distributions of adults are important in assessing risk to a DPS. Spatial distribution is important, both at the scale of the spawning population and the metapopulation.
Assessments of marine fish populations have focused on the biomass or numbers of adults harvested by commercial and sports fishing. Catch records, CPUE, and biomass estimates from research cruises constitute most of the data available to estimate abundance trends. However, the numbers of reproductive adults is the most important measure of abundance in assessing the status of a population. Data on other life-history stages can be used as a supplemental indicator of abundance.
The relationship of present abundance to present carrying capacity is important for evaluating the health of a population, but a population with abundance near the carrying capacity of the habitat it occupies does not necessarily indicate that the population is healthy. Populations abundances near carrying capacity imply that the effectiveness of short-term management actions is limited in increasing population abundance. The relationship between current abundance and habitat capacity to the historical relationship between these variables is an important consideration in evaluating risk. An understanding of historical conditions provides a perspective of the conditions under which present populations evolved. Estimates of historical abundances also provide the basis for establishing long-term abundance trends. Comparisons of past and present habitat capacity can also indicate long-term population trends and potential problems stemming from population fragmentation.
>Short- and long-term trends in abundance are primary indicators of risk in natural populations. Trends may be calculated with a variety of quantitative data, including catch, CPUE, and survey data. Trend analyses for the three species considered in this review is greatly limited by the lack of long time series of abundances in Puget Sound. The times series of abundance estimates that are available are limited in their usefulness by the lack of regular sampling, by use of different survey methods for a species, and, for harvest data, by the imposition of harvest regulations. The influence of environmental variability on population abundances also limits the use of short-term trends, because the climate changes in the late 1970s and 1980s coincided with apparent declines in population abundances for each of the three species being considered in this review.
Several natural and anthropogenic factors influence the degrees of risk facing populations of marine fish in Puget Sound. Recent changes in these factors may influence the degree of risk of a population without apparent changes in abundance, because of time lags between the events and the effects on the population. Thus, a consideration of these effects extends beyond the examination of recent trends in abundance. The BRT considered documented physical and climatic changes, but did not consider possible effects of recent or proposed conservation measures. Population variability in itself may not be an indication of risk, because populations in Puget Sound for two of the species considered in this review are at the margins of the geographical distribution of the species, and thus have evolved in a variable environment. Habitat degradation and harvest have most likely weakened the resilience of populations in Puget Sound to climate variability. However, these effects are not easily quantified.
Artificial propagation and enhancement of populations in Puget Sound does not presently appear to be a risk factor for the species considered here. However, mariculture of some species is under development, and the effects of hatchery releases on natural populations may be important in the future. The interbreeding of cultured and natural fish can potentially lead to a loss in fitness of naturally-spawning populations. The genetic effects of artificially propagated releases of species with high fecundities, as is common for many marine fishes, could be substantial. Ryman and Lairkre (1991), Waples and Do (1994), and Ryman et al. (1995) discussed possible risks associated with enhancement of marine populations, but these risks are difficult to quantify and to incorporate into risk analysis. The chief concern is that the release of propagated fish, which may be inadvertently modified by breeding practices and novel rearing environments, may lead to the erosion of genetic diversity and fitness in natural populations.
Human activities other than population enhancement can also influence the genetic characteristics of natural populations. These include size-selective harvest methods (Nelson and Soulé 1987), introductions of non-native species, and alterations of marine habitats by shoreline development, by increased siltation in river runoff, and by pollution. At the present time, empirical information documenting the genetic effects of these kinds of changes is largely lacking.
Coupled changes in
climate and ocean conditions have occurred on several different time scales
and have influenced the geographical distributions, and hence local
abundances, of marine fishes. On
time scales of hundreds of millennia, periodic cooling produced several
glaciations in the Pleistocene Epoch (Imbrie et al. 1984, Bond et al. 1993). The central part of Puget Sound was covered with ice about 1 km thick
during the last glacial maximum about 14,000 years ago (Thorson 1980). Since the end of this major period of cooling, several population
oscillations of pelagic fishes, such as anchovies and sardines, have been
noted on the west coast of North America (Baumgartner et al. 1992). These oscillations, with periods of about 100 years, have presumably
occurred in response to climatic variability. On decadal time scales, climatic variability in the North Pacific and
North Atlantic Oceans has influenced the abundances and distributions of
widespread species, including several species of Pacific salmon (Francis et
al. 1998, Mantua et al. 1997) in the North Pacific, and Atlantic herring (Alheit
and Hagen 1997) and Atlantic cod (Swain 1999) in the North Atlantic. Recent declines in marine fish populations in Puget Sound may reflect
recent climatic shifts. However,
we do not know whether these climatic shifts represent long-term changes or
short-term fluctuations that may reverse in the near future. Although recent climatic conditions appear to be within the range of
historical conditions, the risks associated with climatic changes may be
exacerbated by human activities (Lawson 1993).