Mikko Koskinen

Contemporary fisherian life-history evolution in small salmonid populations

The relative importance of natural selection and random drift in phenotypic evolution has been discussed since the introduction of the first population genetic models. The empirical evidence used to evaluate the evolutionary theories of Fisher and Wright remains obscure because formal tests for neutral divergence or sensitive attempts to separate the effects of selection and drift are scarce, subject to error, and have not been interpreted in the light of well-known population demography. We combined quantitative genetic and microsatellite DNA analyses to investigate the determinants of contemporary life-history evolution in isolated populations of grayling (Thymallus thymallus, Salmonidae) that originated from a common source 80–120 years ago. Here we show that natural selection was the dominant diversifying agent in the evolution of the quantitative traits. However, the populations were founded by a small number of individuals, exhibit very low microsatellite-based effective sizes and show genetic imprints of severe ‘bottlenecks’; which are conditions often suggested to constrain selection and favour drift. This study demonstrates a very clear case of fisherian evolution in small natural populations across a contemporary timescale.

The one that did not get away: individual assignment using microsatellite data detects a case of fishing competition fraud

Assignment of an individual to the population from which it most probably originated based on its multilocus genotype has been widely applied in recent years. In this study, individual assignment based on microsatellite data was used to identify a case of fishing competition fraud. Despite the fact that the true population of origin was most probably not among the reference populations, recent modifications of the assignment tests were used in confidently excluding (p < 0.0001) the possibility of a 5.5 kg salmon (Salmo salar) originating from the fishing competition location, Lake Saimaa (south-east Finland). In fact, the probability of the suspect salmon originating from one of the regions that supply most of Finlands fish markets was found to be over 600 times higher than it originating from Lake Saimaa. When presented with this evidence, the offender confessed to purchasing the salmon at a local fish shop and criminal charges were laid. This study emphasizes the potential practical application of the individual assignment procedure, in particular the usefulness of confidently excluding populations as the origin of an individual. A similar strategy could be also used, for example in suspected cases of illegal poaching, in order to assign or exclude individuals from originating from a claimed population.

Isolation by distance within a river system: genetic population structuring of Atlantic salmon, Salmo salar, in tributaries of the Varzuga River in northwest Russia

An important issue for designing any conservation programme aimed at preserving genetic diversity is estimation of the scale at which genetic structuring occurs. Additional relevant factors include distinguishing whether or not population structuring is expected to be stable as predicted by the member‐vagrant hypothesis, or alternatively, whether populations are more prone to local extinction–recolonization processes, as predicted by the metapopulation evolutionary model. In this study, the population genetic structure of Atlantic salmon from 11 locations within or nearby the Varzuga River tributary system was assessed using 17 microsatellites. Mantel tests and spatial autocorrelation analyses revealed a significant isolation‐by‐distance signal within the tributary system as well as a negative association between the level of genetic diversity and waterway distance from the river mouth, indicating that dispersal is less likely to occur to populations deep in the tributary system. Individual‐level spatial autocorrelation analyses indicated that the majority of migration occurred between populations situated within 20 km of each other. The relatively high level of genetic structuring and significant isolation‐by‐distance signal observed in the Varzuga tributaries are concordant with the predictions of the member‐vagrant evolutionary model. However, one subpopulation in particular revealed signs of instability which may be due to its location in the tidal zone, or due to the fact that it is more affected by human impacts. The results suggest that preservation of a number of spawning sites spaced throughout the tributary system is recommendable for ensuring sustainable fishing tourism in the river.

Specific DNA adducts induced by some mono-substitued epoxides in vitro and in vivo

Alkyl epoxides are important intermediates in the chemical industry. They are also formed in vivo during the detoxification of alkenes. Alkyl epoxides have shown genotoxicity in many toxicology assays which has been associated with their covalent binding to DNA. Here aspects of the formation and properties of DNA adducts, induced by some industrially important alkenes and mono-substituted epoxides are discussed. These include propylene oxide, epichlorohydrin, allyl glycidyl ether and the epoxy metabolites of styrene and butadiene. The major DNA adducts formed by epoxides are 7-substituted guanines, 1- and 3-substituted adenines and 3-substituted cytosines. In addition, styrene oxide and butadiene monoepoxide are able to modify exocyclic sites in the DNA bases, the sites being in the case of styrene oxide N(2)- and O(6)-positions of guanine, N(6)-adenine as well as N(4)-and O(2)-cytosine. In vivo the main adduct is the 7-substituted guanines. The 1-substituted adenines have also shown marked levels, and these adducts should also be targets in biomonitoring of human exposures. Due to its low mutagenicity, 7-substituted guanines are considered as a surrogate marker for other mutagenic lesions, e.g. those of 1-adenine or 3-uracil adducts.

Genetic lineages and postglacial colonization of grayling (Thymallus thymallus, Salmonidae) in Europe, as revealed by mitochondrial DNA analyses

In stark contrast to other species within the Salmonidae family, phylogeographic information on European grayling, Thymallus thymallus, is virtually nonexistent. In this paper, we utilized mitochondrial DNA polymerase chain reaction–restriction fragment length polymorphism (mtDNA PCR–RFLP) and sequence variation to infer the postglacial dispersal routes of T. thymallus into and within northern Europe, and to locate geographically, potential evolutionarily distinct populations. Mitochondrial analyses revealed a total of 27 T. thymallus haplotypes which clustered into three distinct lineages. Average pairwise interlineage divergence was four and nine times higher than average intralineage divergence for RFLP and sequence data, respectively. Two European grayling individuals from the easternmost sample in Russia exhibited haplotypes more genetically diverged from any T. thymallus haplotype than T. arcticus haplotype, and suggested that hybridization/introgression zone of these two sister species may extend much further west than previously thought. Geographic division of the lineages was generally very clear with northern Europe comprising of two genetically differentiated areas: (i) Finland, Estonia and north‐western Russia; and (ii) central Germany, Poland and western Fennoscandia. Average interpopulation divergence in North European T. thymallus was 10 times higher than that observed in a recent mtDNA study of North American T. arcticus. We conclude that (i) North European T. thymallus populations have survived dramatic Pleistocene temperature oscillations and originate from ancient eastern and central European refugia; (ii) genetic divergence of population groups within northern Europe is substantial and geographically distinct; and (iii) the remainder of Europe harbours additional differentiated assemblages that likely descend from a Danubian refugium. These findings should provide useful information for developing appropriate conservation strategies for European grayling and exemplify a case with a clear need for multinational co‐operation for managing and conserving biodiversity.

Microsatellite data resolve phylogeographic patterns in European grayling, Thymallus thymallus, Salmonidae

The phylogeography of an endangered salmonid, European grayling (Thymallus thymallus), was studied based on analysis of 17 nuclear microsatellite DNA loci. In agreement with earlier mitochondrial DNA (mtDNA) studies, phylogenetic relationships of the populations suggested that northern Europe was colonized from two distinct Pleistocene refugia. Furthermore, microsatellites revealed highly supported grouping of mainland Swedish, Norwegian, Danish, German and Slovenian populations, suggesting that grayling from the northwestern and central Europe have descended from their southern conspecifics. The level of divergence between populations was substantial, even across short geographical distances. Although this was in part due to postglacial colonization patterns and contemporary barriers for gene flow, the high divergence estimates between hydrologically connected sampling locations implied efficient interpopulation reproductive isolation. Microsatellites revealed that the populations exhibited, on average, only 3.5 (±2.2) alleles per locus, indicating that T. thymallus has strikingly low levels of intrapopulation genetic diversity as compared with other freshwater fish species. Accordingly, as indicated by analysis of molecular variance (AMOVA), only 49.1–58.0% of the total grayling microsatellite diversity resided within populations. A latitudinal genetic diversity gradient, potentially resulting from glaciation-mediated founder events, was not evident. Alternatively, it is possible that grayling display limited dispersal behaviour/capability, leading to low long-term effective population sizes and, consequently, depauperate intrapopulation polymorphism. These findings have implications for conservation of T. thymallus. Importantly, they exemplify that microsatellites can be highly informative for intraspecific phylogeography studies dealing with substantial divergence scales.

Mitochondrial and nuclear DNA phylogeography of Thymallus spp. (grayling) provides evidence of ice-age mediated environmental perturbations in the world's oldest body of fresh water, Lake Baikal

Theories on the hydrological history of Lake Baikal, the worlds oldest and deepest body of freshwater, and its surrounding great rivers, are currently based solely on geological evidence and are conflicting. Baikal is inhabited by numerous zoogeographical enigmas but their high level of endemism has hindered phylogeographic inferences. We provide a biological perspective of the regions palaeo‐hydrological development based on the demographic and genealogical history of the widespread Thymallus spp. (grayling). Phylogenetic reconstruction reveals that old lineages of grayling (pre‐Pleistocene) currently inhabit the Enisey, Lena and Amur River basins. For Lake Baikal however, we conclude that a mid‐Pleistocene colonization (110 000–450 000 years ago) of an unoccupied niche has occurred. Population genetic inferences support an Enisey–Angara river route of colonization into Baikal, corresponding to the cataclysmic palaeo‐hydrological event that led to the formation of the lakes only contemporary outlet, and a subsequent range expansion several thousand kilometres into the uppermost reaches of the Selenga River basin. The evolutionary history of Lake Baikal grayling is congruent with the controversial hypothesis of repeated glaciation. However, considering the extraordinary levels of endemism and proposed Miocene or Oligocene coalescence of other faunal lineages, a less profound but equally consequential cycle of environmental perturbations may have taken place. Bi‐parentally inherited microsatellite DNA loci supported the phylogenetic relationships of Thymallus spp. and the geographical expansion of Baikal grayling strikingly well. A Markov Chain Monte Carlo modelling approach suggested severe contemporary population decline during the last century, possibly reflecting the influence of an uncontrolled fishery on this treasured ecosystem. These complementary pictures of the demographic history of grayling underscore the breadth of historical inquiry that can be entertained through the modelling of sufficient molecular data, and may significantly alter the zoogeographical and limnological perspectives of Baikals history.

DNA adducts, strand breaks and micronuclei in mice exposed to styrene by inhalation

Genotoxic and clastogenic effects of styrene were studied in mice. Male NMRI mice were exposed by inhalation to styrene in concentrations of 750 and 1500 mg/m3 for 21, 7, 3 and 1 days (6 h/day, 7 days/week). Followed parameters included styrene in blood, specific styrene oxide (SO) induced DNA adducts, DNA strand breaks and micronuclei. The formation of SO induced 7-SO-guanines and 1-SO-adenines in DNA was analysed from lung tissues by two versions of the 32P-postlabeling technique. In lungs after 21 days of exposure to 1500 mg/m3 the level of 7-SO-guanine was 23.0+/-11.9 adducts/10(8) normal nucleotides, while 1-SO-adenine was detected at the levels of 0.6+/-0.2 adducts/10(8) normal nucleotides. Both 7-SO-guanines and 1-SO-adenines strongly correlated with exposure parameters, particularly with styrene concentration in blood (r=0.875, P=0.0002 and r=0.793, P=0.002, respectively). DNA breaks were measured in peripheral lymphocytes, bone marrow cells and liver cells using comet assay. To discern oxidative damage and abasic sites, endonuclease III was used. In bone marrow of exposed mice slight increase of strand breaks can be detected after 7 days of inhalation. A significant increase was revealed in the endonuclease III-sensitive sites after 21 days of inhalation in bone marrow. In the liver cells inhalation exposure to both concentrations of styrene did not virtually affect either levels of DNA single-strand breaks or endonuclease III-sensitive sites. The inhalation of 1500 mg/m3 of styrene induced significant increase of micronuclei after 7 days of exposure (10.4+/-2.5/1000 cells, i.e. twice higher micronuclei frequency than in controls). After 21 days of inhalation no significant difference between the control group and the two exposed groups was observed. Whether the decrease of micronuclei after 21 days of inhalation was due to the inhibition of cell proliferation caused by styrene or due to the natural elimination of chromatide fragments, remains to be clarified. An interesting link has been found between DNA single-strand breaks in bone marrow and frequencies of micronuclei (r=0.721, P=0.028).

Interpopulation genetic divergence in European grayling (Thymallus thymallus, Salmonidae) at a microgeographic scale: implications for conservation

In order to provide guidelines for conservationand management of the severely declining LakeSaimaa (eastern Finland) European grayling(Thymallus thymallus, Salmonidae), weinvestigated the microgeographic geneticstructure of three populations from the watersystem using 17 microsatellite loci andmitochondrial DNA polymerase chainreaction-restriction fragment lengthpolymorphism analysis. Microsatellites revealedlow levels of intrapopulation genetic diversityand substantial divergence between populationssampled from spawning sites separated by aslittle as 55 kilometers. Mitochondrial analysisindicated the occurrence of two compositehaplotypes within Lake Saimaa. The nucleotidesubstitution estimates between the haplotypessuggested their separation to markedly predatethe late Pleistocene period. The populationsexhibited marked frequency differences for thetwo mitochondrial haplotypes, reinforcing theview of limited interpopulation gene flowwithin Lake Saimaa. An individual basedmicrosatellite Neighbor-Joining dendrogramdemonstrated clustering of all the specimensaccording to their sampling origin. Individualassignment tests revealed 100% assignmentsuccess of individuals into their population oforigin and 100% exclusion (p <0.05) from all alternative referencepopulations. These findings exemplify that T. thymallus populations may be genetically highly structured over small geographical scales and provide a good starting point for the development of appropriate conservation strategies for Lake Saimaa grayling.

Spectrum of styrene-induced DNA adducts: the relationship to other biomarkers and prospects in human biomonitoring.

Styrene is an important industrial chemical that has shown genotoxicity in many toxicology assays. This is believed to be related to the DNA-binding properties of styrene-7,8-oxide (SO), a major metabolite of styrene. In this review, we have summarized knowledge on various aspects of styrene genotoxicity, especially in order to understand the formation and removal of primary DNA lesions, and the usefulness of biomarkers for risk assessment. Biological significances of specific DNA adducts and their role in the cascade of genotoxic events are discussed. Links between markers of external and internal exposure are evaluated, as well as metabolic aspects leading to the formation of DNA adducts and influencing biomarkers of biological effect. Finally, we suggest a design of a population study, which may contribute to our understanding genotoxic events in the exposure either to single xenobiotic or complex mixture.