Patterns of variability at the major histocompatibility class II alpha locus in Atlantic salmon contrast with those at the class I locus

In order to investigate the mechanisms creating and maintaining variability at the major histocompatibility (MH) class II alpha (DAA) locus we examined patterns of polymorphism in two isolated Atlantic salmon populations which share a common post-glacial origin. As expected from their common origin, but contrary to the observation at the MH class I locus, these populations shared the majority of DAA alleles: out of 17 sequences observed, 11 were common to both populations. Recombination seems to play a more important role in the origin of new alleles at the class II alpha locus than at the class I locus. A greater than expected proportion of sites inferred to be positively selected (potentially peptide binding residues, PBRs) were found to be involved in recombination events, suggesting a mechanism for increasing MH variability through an interaction between recombination and natural selection. Thus it appears that although selection and recombination are important mechanisms for the evolution of both class II alpha and class I loci in the Atlantic salmon, the pattern of variability differs markedly between these classes of MH loci.     

Evidence for co‐dominant allelic expression of a phosphoglucomutase regulatory locus polymorphism in the Atlantic salmon

Starch gel electrophoresis of the phosphoglucomutase isozyme, PGM‐1, in liver tissue in Atlantic salmon Salmo salar shows three phenotypes – full, partial and no expression. The genetic basis of the variation has not been established. Studies of the inheritance of the variation and of liver enzyme levels carried out indicate that the variation, as in rainbow trout Onchorynchus mykiss , reflects co‐dominant allelic variation at a cis ‐acting regulatory locus, designated PGM‐1r *, with one allele promoting and the other suppressing expression.     

Multiplication of the class I alcohol dehydrogenase locus in mammalian evolution

Chromosomal DNA samples derived from various primates and other mammals (horse, sheep, rabbit, and mouse) were digested with restriction endonuclease and hybridized with a probe of the sixth exon of the human ADH gene, which is highly conserved in the class I alcohol dehydrogenase of these mammalian species. The copy number of the class I ADH gene in each species was estimated from the number of hybridized bands. Primate DNA samples showed three distinct bands in the blots of PstI digest and DraI digest. Moreover, most of the bands from primate DNA showed a similarity in size so as to allow us to assign the ADH1, ADH2, and ADH3 homologues in each species. In contrast, mouse has only one gene, and rabbit, sheep, and horse seem to have only two genes, for the class I ADH, which showed divergent hybridization bands. These results are consistent with the view that the human class I ADH gene cluster has been generated through gene multiplication events which occurred before the Catarrhini branch point in the course of primate evolution.     

Two ancient allelic lineages at the single classical class I locus in the Xenopus MHC

Unlike all other vertebrates examined to date, there is only one detectable class I locus in the Xenopus MHC. On the bases of a nearly ubiquitous and high tissue expression, extensive polymorphism, and MHC linkage, this gene is of the classical or class Ia type. Sequencing analysis of class Ia cDNAs encoded by eight defined MHC haplotypes reveals two very old allelic lineages that perhaps emerged when humans and mice diverged from a common ancestor up to 100 million years ago. The unprecedented age of these lineages suggests that different class Ia genes from ancestors of the laboratory model Xenopus laevis are now expressed as alleles in this species. The lineages are best defined by their cytoplasmic and alpha2 peptide-binding domains, and there are highly diverse alleles (defined by the alpha1 peptide-binding domain) in each lineage. Surprisingly, the alpha3 domains are homogenized in both lineages, suggesting that interallelic gene conversion/recombination maintains the high sequence similarity.     

Repeated evolution of vertebrate pollination syndromes in a recently diverged Andean plant clade

Although specialized interactions, including those involving plants and their pollinators, are often invoked to explain high species diversity, they are rarely explored at macroevolutionary scales. We investigate the dynamic evolution of hummingbird and bat pollination syndromes in the centropogonid clade (Lobelioideae: Campanulaceae), an Andean‐centered group of ∼550 angiosperm species. We demonstrate that flowers hypothesized to be adapted to different pollinators based on flower color fall into distinct regions of morphospace, and this is validated by morphology of species with known pollinators. This supports the existence of pollination syndromes in the centropogonids, an idea corroborated by ecological studies. We further demonstrate that hummingbird pollination is ancestral, and that bat pollination has evolved ∼13 times independently, with ∼11 reversals. This convergence is associated with correlated evolution of floral traits within selective regimes corresponding to pollination syndrome. Collectively, our results suggest that floral morphological diversity is extremely labile, likely resulting from selection imposed by pollinators. Finally, even though this clade's rapid diversification is partially attributed to their association with vertebrate pollinators, we detect no difference in diversification rates between hummingbird‐ and bat‐pollinated lineages. Our study demonstrates the utility of pollination syndromes as a proxy for ecological relationships in macroevolutionary studies of certain species‐rich clades.     

Inbreeding depression and low between-population heterosis in recently diverged experimental populations of a selfing species

In fragmented populations, genetic drift and selection reduce genetic diversity, which in turn results in a loss of fitness or in a loss of evolvability. Genetic rescue, that is, controlled input of diversity from distant populations, may restore evolutionary potential, whereas outbreeding depression might counteract the positive effect of this strategy. We carried out self-pollination and crosses within and between populations in an experimental subdivided population of a selfing species, Triticum aestivum L., to estimate the magnitude of these two phenomena. Surprisingly, for a self-fertilizing species, we found significant inbreeding depression within each population for four of the six traits studied, indicating that mildly deleterious mutations were still segregating in these populations. The progeny of within- and between-population crosses was very similar, indicating low between-population heterosis and little outbreeding depression. We conclude that relatively large population effective sizes prevented fixation of a high genetic load and that local adaptation was limited in these recently diverged populations. The kinship coefficient estimated between the parents using 20 neutral markers was a poor predictor of the progeny phenotypic values, indicating that there was a weak link between neutral diversity and genes controlling fitness-related traits. These results show that when assessing the viability of natural populations and the need for genetic rescue, the use of neutral markers should be complemented with information about the presence of local adaptation in the subdivided population.     

The major histocompatibility class I locus in Atlantic salmon (Salmo salar L.): polymorphism,linkage analysis and protein modelling

A cDNA library screening using the conserved exon 4 of Atlantic salmon Mhc class I as probe provided the basis for a study on Mhc class I polymorphism in a breeding population. Twelve different alleles were identified in the 82 dams and sires studied. No individual expressed more than two alleles, which corresponded to the diploid segregation patterns of the polymorphic marker residing within the 3'-untranslated tail. Close linkage between the Sasa-UBA and Sasa-TAP2B loci strengthens the claim that Sasa-UBA is the major Mhc class I locus in Atlantic salmon. We found no evidence for a second expressed classical or non-classical Mhc class I locus in Atlantic salmon. A phylogenetic analysis of salmonid Mhc class I sequences showed domains conserved between rainbow trout, brown trout and Atlantic salmon. Evidence for shuffling of the alpha(1) domain was identified and lineages of the remaining alpha(2) through the cytoplasmic tail gene segment can be defined. The coding sequence of one allele was found associated with two different markers, suggesting recombination within the 3'-tail dinucleotide repeat itself. Protein modelling of several Sasa-UBA alleles shows distinct differences in their peptide binding domains and enables a further understanding of the functionality of the high polymorphism.     

Reconstructing a missing link in the evolution of a recently diverged phosphotriesterase by active-site loop remodeling

Only decades after the introduction of organophosphate pesticides, bacterial phosphotriesterases (PTEs) have evolved to catalyze their degradation with remarkable efficiency. Their closest known relatives, lactonases, with promiscuous phosphotriasterase activity, dubbed PTE-like lactonases (PLLs), share only 30% sequence identity and also differ in the configuration of their active-site loops. PTE was therefore presumed to have evolved from a yet unknown PLL whose primary activity was the hydrolysis of quorum sensing homoserine lactones (HSLs) (Afriat et al. (2006) Biochemistry45, 13677-13686). However, how PTEs diverged from this presumed PLL remains a mystery. In this study we investigated loop remodeling as a means of reconstructing a homoserine lactonase ancestor that relates to PTE by few mutational steps. Although, in nature, loop remodeling is a common mechanism of divergence of enzymatic functions, reproducing this process in the laboratory is a challenge. Structural and phylogenetic analyses enabled us to remodel one of PTE's active-site loops into a PLL-like configuration. A deletion in loop 7, combined with an adjacent, highly epistatic, point mutation led to the emergence of an HSLase activity that is undetectable in PTE (k(cat)/K(M) values of up to 2 × 10(4)). The appearance of the HSLase activity was accompanied by only a minor decrease in PTE's paraoxonase activity. This specificity change demonstrates the potential role of bifunctional intermediates in the divergence of new enzymatic functions and highlights the critical contribution of loop remodeling to the rapid divergence of new enzyme functions.     

Differential evolution of the 13 Atlantic salmon Hox clusters

Hox cluster organization represents a valuable marker to study the effects of recent genome duplication in salmonid fish (25-100 Mya). Using polymerase chain reaction amplification of cDNAs, BAC library screening, and genome walking, we reconstructed 13 Hox clusters in the Atlantic salmon containing 118 Hox genes including 8 pseudogenes. Hox paralogs resulting from the genome duplication preceding the radiation of ray-finned fish have been much better preserved in salmon than in other model teleosts. The last genome duplication in the salmon lineage has been followed by the loss of 1 of the 4 HoxA clusters. Four rounds of genome duplication after the vertebrate ancestor salmon Hox clusters display the main organizational features of vertebrate Hox clusters, with Hox genes exclusively that are densely packed in the same orientation. Recently, duplicated Hox clusters have engaged a process of divergence, with several cases of pseudogenization or asymmetrical evolution of Hox gene duplicates, and a marked erosion of identity in noncoding sequences. Strikingly, the level of divergence attained strongly depends on the Hox cluster pairs rather than on the Hox genes within each cluster. It is particularly high between both HoxBb clusters and both HoxDa clusters, whereas both HoxBa clusters remained virtually identical. Positive selection on the Hox protein-coding sequences could not be detected.     

Characteristics of mature male parr in the northernmost Atlantic salmon populations

The incidence, relative abundance and biological characteristics of mature male parr of Atlantic salmon Salmo salar were studied in two subarctic (69–70 N°) river systems in the northernmost part of Scandinavia. The proportion of mature males was low (0–25%) among ≥1 year‐old parr, and clearly lower than reported earlier in more southern rivers. The ages of mature males ranged from 1+ to 6 + year, the most abundant age group was 3 + year. Both age structure and size of the mature male parr varied between rivers and years. Mature male parr in the main stem were younger and also smaller than the parr of the same age in the tributaries.     

Allelic polymorphism in MHC class II B in four populations of Atlantic salmon (Salmo salar)

We sequenced exon 2 of the MHC class II B gene in Atlantic salmon from the Baltic Sea and identified 17 different exon 2 alleles among 22 different restriction fragment length polymorphism haplotypes. The sequences differed at between 1 and 34 bases. Two different tests were used to estimate the importance of recombination in the generation of new alleles. Recombination events appear to have occurred between three and nine times. Only two pairs of sequences differed by less than five nucleotides, minimizing the importance of point mutations for generating new alleles. Phylogenetic analysis showed that sequences did not cluster according to populations, and genetic distances between populations were small compared to those obtained by allele frequency data. These results, together with the similarity found between exon 2 sequences from Baltic salmon and Norwegian salmon, indicate that all of the identified alleles were present in the ancient salmon population colonizing the Baltic rivers after the last glaciation.     

Natural selection acts on Atlantic salmon major histocompatibility (MH) variability in the wild

Pathogen-driven balancing selection is thought to maintain polymorphism in major histocompatibility (MH) genes. However, there have been few empirical demonstrations of selection acting on MH loci in natural populations. To determine whether natural selection on MH genes has fitness consequences for wild Atlantic salmon in natural conditions, we compared observed genotype frequencies of Atlantic salmon (Salmo salar) surviving in a river six months after their introduction as eggs with frequencies expected from parental crosses. We found significant differences between expected and observed genotype frequencies at the MH class II alpha locus, but not at a MH class I-linked microsatellite or at seven non-MH-linked microsatellite loci. We therefore conclude that selection at the MH class II alpha locus was a result of disease-mediated natural selection, rather than any demographic event. We also show that survival was associated with additive allelic effects at the MH class II alpha locus. Our results have implications for both the conservation of wild salmon stocks and the management of disease in hatchery fish. We conclude that natural or hatchery populations have the best chance of dealing with episodic and variable disease challenges if MH genetic variation is preserved both within and among populations.     

How specific MHC class I and class II combinations affect disease resistance against infectious salmon anaemia in Atlantic salmon (Salmo salar)

The aim was to evaluate the performance of selected individual MHC class I and class II alpha (A) alleles, and combinations of these on disease resistance against infectious salmon anaemia (ISA). The material consisting of 1966 fish from seven families, contained five MHC class I alleles and four MHC class II A alleles. Which representing given class II A and class II beta (B) haplotypes, totalling 19 MHC class I and class II A genotypes. The fish were challenged with infectious salmon anaemia virus (ISAV), the virus causing ISA. Dead fish were collected daily during the challenge experiment and the survivors were collected at termination. All fish were genotyped for MHC class I and class II A. The total mortality in the material was 85.14%. For MHC class I, UBA*0201 and UBA*0301 were significantly the most resistant alleles, while UBA*0601 for class I and DAA*0301 for class II A were the significantly most susceptible alleles. The analysis of combined MHC class I and class II A genotypes detected that fish with the genotype UBA*0201/*0301;DAA*0201/*0201 were the most resistant fish with a hazard ratio (HR) at 0.750, while the fish with the genotypes UBA*0601/*0801;DAA*0501/*0501 and UBA*0201/*0301;DAA*0301/*0501 were the most susceptible fish with HR of 1.334 and 1.425. In addition, Cox regression analysis within family detected combined MHC class I and class II A genotypes that contributed significantly to resistance and susceptibility. The study confirmed the expectation of performance of individual MHC class I and class II A alleles, and also detected an effect of MHC class I and class II A in combinations.     

Testing mate choice and overdominance at MH in natural families of Atlantic salmon Salmo salar

This study aimed to test mate choice and selection during early life stages on major histocompatibility (MH) genotype in natural families of Atlantic salmon Salmo salar spawners and juveniles, using nine microsatellites to reconstruct families, one microsatellite linked to an MH class I gene and one minisatellite linked to an MH class II gene. MH‐based mate choice was only detected for the class I locus on the first year, with lower expected heterozygosity in the offspring of actually mated pairs than predicted under random mating. The genotype frequencies of MH‐linked loci observed in the juveniles were compared with frequencies expected from Mendelian inheritance of parental alleles to detect selection during early life stages. No selection was detected on the locus linked to class I gene. For the locus linked to class II gene, observed heterozygosity was higher than expected in the first year and lower in the second year, suggesting overdominance and underdominance, respectively. Within family, juveniles' body size was linked to heterozygosity at the same locus, with longer heterozygotes in the first year and longer homozygotes in the second year. Selection therefore seems to differ from one locus to the other and from year to year.     

Genetic variation at enzyme loci in the southernmost European populations of Atlantic salmon

Genetic variation was studied at 8 allozyme loci in six severely endangered wild Spanish populations of Salmo salar located at the southernmost geographical limit of European Atlantic salmon. Low levels of variation were detected and no significant deviations from Hardy–Weinberg equilibrium were found.     

Zygosity at the major histocompatibility class IIB locus predicts susceptibility to Renibacterium salmoninarum in Atlantic salmon (Salmo salar L.)

Major histocompatibility (MH) class II genes play an important role in the vertebrate immune response. Here, we investigate the relationship between Atlantic salmon (Salmo salar) MH class IIB zygosity and susceptibility to Renibacterium salmoninarum, the causal agent of bacterial kidney disease. By combining DNA sequences from the salmon MH class IIB gene with quantitative ELISA data on R. salmoninarum antigen levels, we found that MH class IIB homozygotes were significantly more susceptible to R. salmoninarum than heterozygotes. These findings are discussed in the context of current evolutionary theory.     

Differences in morphology, performance and behaviour between recently diverged populations of Podarcis sicula mirror differences in predation pressure

We investigated the possible role of variation in predation pressure in the phenotypic divergence of two island populations of the Italian wall lizard, Podarcis sicula . In 1971, ten adult specimens from the island of Pod Kopište (Adriatic Sea, Croatia) were transported to the island of Pod Mrčaru, 3.5 km east, where they founded a new population. Although the two islands resemble each other in general physiognomy (size, elevation, microclimate) and in the absence of terrestrial predators, lizards from the newly established population are now on average larger and have shorter hind limbs. They also exhibit lower maximal sprint speed as measured on a racetrack, and fatigue faster when chased in a torus track. In the field, lizards from the original population of Pod Kopište respond to a simulated predatory attack by fleeing at larger approach distances and by running further from the predator than lizards from Pod Mrčaru. These changes in morphology, behaviour and performance may result from the relaxed predation intensity on the latter island. Our analysis of the structural features of the microhabitats suggests that the vegetation on Pod Mrčaru offers more protection to lizards. Also, plasticine models of lizards, laid out on the islands, less often exhibited signs of being attacked by birds on Pod Mrčaru than on Pod Kopište. Our findings provide an example of how changes in (possibly a single) environmental factor may simultaneously produce responses in behaviour, morphology and whole-animal physiology, and this on a surprisingly small spatial and temporal scale.     

Assessing footprints of selection in commercial Atlantic salmon populations using microsatellite data

Relatively large rates of response to traits of economic importance have been observed in different selection experiments in salmon. Several QTL have been mapped in the salmon genome, explaining unprecedented levels of phenotypic variation. Owing to the relatively large selection intensity, individual loci may be indirectly selected, leaving molecular footprints of selection, together with increased inbreeding, as its likely relatives will share the selected loci. We used population differentiation and levels of linkage disequilibrium in chromosomes known to be harbouring QTL for body weight, infectious pancreatic necrosis resistance and infectious salmon anaemia resistance to assess the recent selection history at the genomic level in Atlantic salmon. The results clearly suggest that the marker SSA0343BSFU on chromosome 3 (body weight QTL) showed strong evidence of directional selection. It is intriguing that this marker is physically mapped to a region near the coding sequence of DVL2 , making it an ideal candidate gene to explain the rapid evolutionary response of this chromosome to selection for growth in Salmo salar. Weak evidence of diversifying selection was observed in the QTL associated with infectious pancreatic necrosis and infectious salmon anaemia resistance. Overall, this study showed that artificial selection has produced important changes in the Atlantic salmon genome, validating QTL in commercial salmon populations used for production purposes according to the recent selection history.