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Background: Recent work suggests that gene duplications may play an important role in the evolution of immunity genes. Passerine birds, and in particular Sylvioidea warblers, have highly duplicated major histocompatibility complex (MHC) genes, which are key in immunity, compared to other vertebrates. However, reasons for this high MHC gene copy number are yet unclear. High-throughput sequencing (HTS) allows MHC genotyping even in individuals with extremely duplicated genes. This HTS data can reveal evidence of selection, which may help to unravel the putative functions of different gene copies, i.e. neofunctionalization. We performed exhaustive genotyping of MHC class I in a Sylvioidea warbler, the sedge warbler, Acrocephalus schoenobaenus, using the Illumina MiSeq technique on individuals from a wild study population. Results: The MHC diversity in 863 genotyped individuals by far exceeds that of any other bird species described to date. A single individual could carry up to 65 different alleles, a large proportion of which are expressed (transcribed). The MHC alleles were of three different lengths differing in evidence of selection, diversity and divergence within our study population. Alleles without any deletions and alleles containing a 6 bp deletion showed characteristics of classical MHC genes, with evidence of multiple sites subject to positive selection and high sequence divergence. In contrast, alleles containing a 3 bp deletion had no sites subject to positive selection and had low divergence. Conclusions: Our results suggest that sedge warbler MHC alleles that either have no deletion, or contain a 6 bp deletion, encode classical antigen presenting MHC molecules. In contrast, MHC alleles containing a 3 bp deletion may encode molecules with a different function. This study demonstrates that highly duplicated MHC genes can be characterised with HTS and that selection patterns can be useful for revealing neofunctionalization. Importantly, our results highlight the need to consider the putative function of different MHC genes in future studies of MHC in relation to disease resistance and fitness. [ABSTRACT FROM AUTHOR]

Background: The number of partners that individuals mate with over their lifetime is a defining feature of mating systems, and variation in mate number is thought to be a major driver of sexual evolution. Although previous research has investigated the evolutionary consequences of reductions in the number of mates, we know little about the costs and benefits of increased numbers of mates. Here, we use a genetic manipulation of mating frequency in Drosophila melanogaster to create a novel, highly promiscuous mating system. We generated D. melanogaster populations in which flies were deficient for the sex peptide receptor (SPR) gene - resulting in SPR- females that mated more frequently - and genetically-matched control populations, and allowed them to evolve for 55 generations. At several time-points during this experimental evolution, we assayed behavioural, morphological and transcriptional reproductive phenotypes expected to evolve in response to increased population mating frequencies. Results: We found that males from the high mating frequency SPR- populations evolved decreased ability to inhibit the receptivity of their mates and decreased copulation duration, in line with predictions of decreased per-mating investment with increased sperm competition. Unexpectedly, SPR- population males also evolved weakly increased sex peptide (SP) gene expression. Males from SPR- populations initially (i.e., before experimental evolution) exhibited more frequent courtship and faster time until mating relative to controls, but over evolutionary time these differences diminished or reversed. Conclusions: In response to experimentally increased mating frequency, SPR- males evolved behavioural responses consistent with decreased male post-copulatory investment at each mating and decreased overall pre-copulatory performance. The trend towards increased SP gene expression might plausibly relate to functional differences in the two domains of the SP protein. Our study highlights the utility of genetic manipulations of animal social and sexual environments coupled with experimental evolution. [ABSTRACT FROM AUTHOR]

Background Geographic barriers to gene flow and divergence among populations in sexual traits are two important causes of genetic isolation which may lead to speciation. Genetic isolation may be facilitated if these two mechanisms act synergistically. The guppy from the Cumaná region (within the Cariaco drainage) of eastern Venezuela has been previously described as a case of incipient speciation driven by sexual selection, significantly differentiated in sexual colouration and body shape from the common guppy, Poecilia reticulata. The latter occurs widely in northern Venezuela, including the south-eastern side of Cordillera de la Costa, where it inhabits streams belonging to the San Juan drainage. Here, we present molecular and morphological analyses of differentiation among guppy populations in the Cariaco and San Juan drainages. Our analyses are based on a 953 bp long mtDNA fragment, a set of 15 microsatellites (519 fish from 20 populations), and four phenotypic traits. Results Both microsatellite and mtDNA data showed that guppies inhabiting the two drainages are characterised by a significant genetic differentiation, but a higher proportion of the genetic variance was distributed among populations within regions. Most guppies in the Cariaco drainage had mtDNA from a distinct lineage, but we also found evidence for widespread introgression of mtDNA from the San Juan drainage into the Cariaco drainage. Phenotypically, populations in the two regions differed significantly only in the number of black crescents. Phenotypic clustering did not support existence of two distinct groupings, but indicated a degree of distinctiveness of Central Cumaná (CC) population. owever, CC population showed little differentiation at the neutral markers from the proximate populations within the Cariaco drainage. Conclusions Our findings are consistent with only partial genetic isolation between the two geographic regions and indicate that the geographic barrier of Cordillera de la Costa has not played an important role in strengthening the incomplete pre-zygotic reproductive barrier between Cumaná and common guppy. Significant phenotypic differentiation between genetically similar (in terms of neutral variation) populations suggests that mate choice can maintain divergence at sexually selected traits despite gene flow. However, neither genetic nor phenotypic clustering supported delineation of two species within the region. [ABSTRACT FROM AUTHOR]

Background: Major histocompatibility complex (MHC) proteins constitute an essential component of the vertebrate immune response, and are coded by the most polymorphic of the vertebrate genes. Here, we investigated sequence variation and evolution of MHC class I and class II DRB, DQA and DQB genes in the brown bear Ursus arctos to characterise the level of polymorphism, estimate the strength of positive selection acting on them, and assess the extent of gene orthology and trans-species polymorphism in Ursidae. Results: We found 37 MHC class I, 16 MHC class II DRB, four DQB and two DQA alleles. We confirmed the expression of several loci: three MHC class I, two DRB, two DQB and one DQA. MHC class I also contained two clusters of non-expressed sequences. MHC class I and DRB allele frequencies differed between northern and southern populations of the Scandinavian brown bear. The rate of nonsynonymous substitutions (dN) exceeded the rate of synonymous substitutions (dS) at putative antigen binding sites of DRB and DQB loci and, marginally significantly, at MHC class I loci. Models of codon evolution supported positive selection at DRB and MHC class I loci. Both MHC class I and MHC class II sequences showed orthology to gene clusters found in the giant panda Ailuropoda melanoleuca. Conclusions: Historical positive selection has acted on MHC class I, class II DRB and DQB, but not on the DQA locus. The signal of historical positive selection on the DRB locus was particularly strong, which may be a general feature of caniforms. The presence of MHC class I pseudogenes may indicate faster gene turnover in this class through the birth-and-death process. South–north population structure at MHC loci probably reflects origin of the populations from separate glacial refugia. [ABSTRACT FROM AUTHOR]

Background: Because of their functional significance, the Major Histocompatibility Complex (MHC) class I and II genes have been the subject of continuous interest in the fields of ecology, evolution and conservation. In some vertebrate groups MHC consists of multiple loci with similar alleles; therefore, the multiple loci must be genotyped simultaneously. In such complex systems, understanding of the evolutionary patterns and their causes has been limited due to challenges posed by genotyping. Results: Here we used 454 amplicon sequencing to characterize MHC class IIB exon 2 variation in the collared flycatcher, an important organism in evolutionary and immuno-ecological studies. On the basis of over 152,000 sequencing reads we identified 194 putative alleles in 237 individuals. We found an extreme complexity of the MHC class IIB in the collared flycatchers, with our estimates pointing to the presence of at least nine expressed loci and a large, though difficult to estimate precisely, number of pseudogene loci. Many similar alleles occurred in the pseudogenes indicating either a series of recent duplications or extensive concerted evolution. The expressed alleles showed unambiguous signals of historical selection and the occurrence of apparent interlocus exchange of alleles. Placing the collared flycatcher's MHC sequences in the context of passerine diversity revealed transspecific MHC class II evolution within the Muscicapidae family. Conclusions: 454 amplicon sequencing is an effective tool for advancing our understanding of the MHC class II structure and evolutionary patterns in Passeriformes. We found a highly dynamic pattern of evolution of MHC class IIB genes with strong signals of selection and pronounced sequence divergence in expressed genes, in contrast to the apparent sequence homogenization in pseudogenes. We show that next generation sequencing offers a universal, affordable method for the characterization and, in perspective, genotyping of MHC systems of virtually any complexity. [ABSTRACT FROM AUTHOR]

Background: The extreme polymorphism that is observed in major histocompatibility complex (MHC) genes, which code for proteins involved in recognition of non-self oligopeptides, is thought to result from a pressure exerted by parasites because parasite antigens are more likely to be recognized by MHC heterozygotes (heterozygote advantage) and/or by rare MHC alleles (negative frequency-dependent selection). The Ewens- Watterson test (EW) is often used to detect selection acting on MHC genes over the recent history of a population. EW is based on the expectation that allele frequencies under balancing selection should be more even than under neutrality. We used computer simulations to investigate whether this expectation holds for selection exerted by parasites on host MHC genes under conditions of heterozygote advantage and negative frequency-dependent selection acting either simultaneously or separately. Results: In agreement with simple models of symmetrical overdominance, we found that heterozygote advantage acting alone in populations does, indeed, result in more even allele frequency distributions than expected under neutrality, and this is easily detectable by EW. However, under negative frequency-dependent selection, or under the joint action of negative frequency-dependent selection and heterozygote advantage, distributions of allele frequencies were less predictable: the majority of distributions were indistinguishable from neutral expectations, while the remaining runs resulted in either more even or more skewed distributions than under neutrality. Conclusions: Our results indicate that, as long as negative frequency-dependent selection is an important force maintaining MHC variation, the EW test has limited utility in detecting selection acting on these genes. [ABSTRACT FROM AUTHOR]

Background: Understanding the genetic basis of adaptive changes has been a major goal of evolutionary biology. In complex organisms without sequenced genomes, de novo transcriptome assembly using a longer read sequencing technology followed by expression profiling using short reads is likely to provide comprehensive identification of adaptive variation at the expression level and sequence polymorphisms in coding regions. We performed sequencing and de novo assembly of the bank vole heart transcriptome in lines selected for high metabolism and unselected controls. Results: A single 454 Titanium run produced over million reads, which were assembled into 63,581 contigs. Searches against the SwissProt protein database and the ENSEMBL collection of mouse transcripts detected similarity to 11,181 and 14,051 genes, respectively. As judged by the representation of genes from the heart-related Gene Ontology categories and UniGenes detected in the mouse heart, our detection of the genes expressed in the heart was nearly complete (> 95% and almost 90% respectively). On average, 38.7% of the transcript length was covered by our sequences, with notably higher (45.0%) coverage of coding regions than of untranslated regions (24.5% of 5' and 32.7% of 3'UTRs). Lower sequence conservation between mouse and bank vole in untranslated regions was found to be partially responsible for poorer UTR representation. Our data might suggest a widespread transcription from noncoding genomic regions, a finding not reported in previous studies regarding transcriptomes in non-model organisms. We also identified over 19 thousand putative single nucleotide polymorphisms (SNPs). A much higher fraction of the SNPs than expected by chance exhibited variant frequency differences between selection regimes. Conclusion: Longer reads and higher sequence yield per run provided by the 454 Titanium technology in comparison to earlier generations of pyrosequencing proved beneficial for the quality of assembly. An almost full representation of genes known to be expressed in the mouse heart was identified. Usage of the extensive genomic resources available for the house mouse, a moderately (20-40 mln years) divergent relative of the voles, enabled a comprehensive assessment of the transcript completeness. Transcript sequences generated in the present study allowed the identification of candidate SNPs associated with divergence of selection lines and constitute a valuable permanent resource forming a foundation for RNAseq experiments aiming at detection of adaptive changes both at the level of gene expression and sequence variants, that would facilitate studies of the genetic basis of evolutionary divergence. [ABSTRACT FROM AUTHOR]

Background: Understanding the genetic basis of adaptive changes has been a major goal of evolutionary biology. In complex organisms without sequenced genomes, de novo transcriptome assembly using a longer read sequencing technology followed by expression profiling using short reads is likely to provide comprehensive identification of adaptive variation at the expression level and sequence polymorphisms in coding regions. We performed sequencing and de novo assembly of the bank vole heart transcriptome in lines selected for high metabolism and unselected controls., Results: A single 454 Titanium run produced over million reads, which were assembled into 63,581 contigs. Searches against the SwissProt protein database and the ENSEMBL collection of mouse transcripts detected similarity to 11,181 and 14,051 genes, respectively. As judged by the representation of genes from the heart-related Gene Ontology categories and UniGenes detected in the mouse heart, our detection of the genes expressed in the heart was nearly complete (> 95% and almost 90% respectively). On average, 38.7% of the transcript length was covered by our sequences, with notably higher (45.0%) coverage of coding regions than of untranslated regions (24.5% of 5' and 32.7% of 3'UTRs). Lower sequence conservation between mouse and bank vole in untranslated regions was found to be partially responsible for poorer UTR representation. Our data might suggest a widespread transcription from noncoding genomic regions, a finding not reported in previous studies regarding transcriptomes in non-model organisms. We also identified over 19 thousand putative single nucleotide polymorphisms (SNPs). A much higher fraction of the SNPs than expected by chance exhibited variant frequency differences between selection regimes., Conclusion: Longer reads and higher sequence yield per run provided by the 454 Titanium technology in comparison to earlier generations of pyrosequencing proved beneficial for the quality of assembly. An almost full representation of genes known to be expressed in the mouse heart was identified. Usage of the extensive genomic resources available for the house mouse, a moderately (20-40 mln years) divergent relative of the voles, enabled a comprehensive assessment of the transcript completeness. Transcript sequences generated in the present study allowed the identification of candidate SNPs associated with divergence of selection lines and constitute a valuable permanent resource forming a foundation for RNAseq experiments aiming at detection of adaptive changes both at the level of gene expression and sequence variants, that would facilitate studies of the genetic basis of evolutionary divergence.