Nuclear markers reveal unexpected genetic variation and a Congolese-Nilotic origin of the Lake Victoria cichlid species flock

Phylogenetic analyses based on mitochondrial (mt) DNA have indicated that the cichlid species flock of the Lake Victoria region is derived from a single ancestral species found in East African rivers, closely related to the ancestor of the Lake Malawi cichlid species flock. The Lake Victoria flock contains ten times less mtDNA variation than the Lake Malawi radiation, consistent with current estimates of the ages of the lakes. We present results of a phylogenetic investigation using nuclear (amplified fragment length polymorphism) markers and a wider coverage of riverine haplochromines. We demonstrate that the Lake Victoria–Edward flock is derived from the morphologically and ecologically diverse cichlid genus Thoracochromis from the Congo and Nile, rather than from the phenotypically conservative East African Astatotilapia. This implies that the ability to express much of the morphological diversity found in the species flock may by far pre–date the origin of the flock. Our data indicate that the nuclear diversity of the Lake Victoria–Edward species flock is similar to that of the Lake Malawi flock, indicating that the genetic diversity is considerably older than the 15 000 years that have passed since the lake began to refill. Most of this variation is manifested in trans–species polymorphisms, indicating very recent cladogenesis from a genetically very diverse founder stock. Our data do not confirm strict monophyly of either of the species flocks, but raise the possibility that these flocks have arisen from hybrid swarms.

Continental-Scale Footprint of Balancing and Positive Selection in a Small Rodent (Microtus arvalis)

Genetic adaptation to different environmental conditions is expected to lead to large differences between populations at selected loci, thus providing a signature of positive selection. Whereas balancing selection can maintain polymorphisms over long evolutionary periods and even geographic scale, thus leads to low levels of divergence between populations at selected loci. However, little is known about the relative importance of these two selective forces in shaping genomic diversity, partly due to difficulties in recognizing balancing selection in species showing low levels of differentiation. Here we address this problem by studying genomic diversity in the European common vole (Microtus arvalis) presenting high levels of differentiation between populations (average FST = 0.31). We studied 3,839 Amplified Fragment Length Polymorphism (AFLP) markers genotyped in 444 individuals from 21 populations distributed across the European continent and hence over different environmental conditions. Our statistical approach to detect markers under selection is based on a Bayesian method specifically developed for AFLP markers, which treats AFLPs as a nearly codominant marker system, and therefore has increased power to detect selection. The high number of screened populations allowed us to detect the signature of balancing selection across a large geographic area. We detected 33 markers potentially under balancing selection, hence strong evidence of stabilizing selection in 21 populations across Europe. However, our analyses identified four-times more markers (138) being under positive selection, and geographical patterns suggest that some of these markers are probably associated with alpine regions, which seem to have environmental conditions that favour adaptation. We conclude that despite favourable conditions in this study for the detection of balancing selection, this evolutionary force seems to play a relatively minor role in shaping the genomic diversity of the common vole, which is more influenced by positive selection and neutral processes like drift and demographic history.

Differential diagnosis of the honey bee trypanosomatids Crithidia mellificae and Lotmaria passim

Trypanosomatids infecting honey bees have been poorly studied with molecular methods until recently. After the description of Crithidia mellificae (Langridge and McGhee, 1967) it took about forty years until molecular data for honey bee trypanosomatids became available and were used to identify and describe a new trypanosomatid species from honey bees, Lotmaria passim (Evans and Schwarz, 2014). However, an easy method to distinguish them without sequencing is not yet available. Research on the related bumble bee parasites Crithidia bombi and Crithidia expoeki revealed a fragment length polymorphism in the internal transcribed spacer 1 (ITS1), which enabled species discrimination. In search of fragment length polymorphisms for differential diagnostics in honey bee trypanosomatids, we studied honey bee trypanosomatid cell cultures of C. mellificae and L. passim. This research resulted in the identification of fragment length polymorphisms in ITS1 and ITS1-2 markers, which enabled us to develop a diagnostic method to differentiate both honey bee trypanosomatid species without the need for sequencing. However, the amplification success of the ITS1 marker depends probably on the trypanosomatid infection level. Further investigation confirmed that L. passim is the dominant species in Belgium, Japan and Switzerland. We found C. mellificae only rarely in Belgian honey bee samples, but not in honey bee samples from other countries. C. mellificae was also detected in mason bees (Osmia bicornis and Osmia cornuta) besides in honey bees. Further, the characterization and comparison of additional markers from L. passim strain SF (published as C. mellificae strain SF) and a Belgian honey bee sample revealed very low divergence in the 18S rRNA, ITS1-2, 28S rRNA and cytochrome b sequences. Nevertheless, a variable stretch was observed in the gp63 virulence factor.

A second generation radiation hybrid map to aid the assembly of the bovine genome sequence

BACKGROUND: Several approaches can be used to determine the order of loci on chromosomes and hence develop maps of the genome. However, all mapping approaches are prone to errors either arising from technical deficiencies or lack of statistical support to distinguish between alternative orders of loci. The accuracy of the genome maps could be improved, in principle, if information from different sources was combined to produce integrated maps. The publicly available bovine genomic sequence assembly with 6x coverage (Btau_2.0) is based on whole genome shotgun sequence data and limited mapping data however, it is recognised that this assembly is a draft that contains errors. Correcting the sequence assembly requires extensive additional mapping information to improve the reliability of the ordering of sequence scaffolds on chromosomes. The radiation hybrid (RH) map described here has been contributed to the international sequencing project to aid this process. RESULTS: An RH map for the 30 bovine chromosomes is presented. The map was built using the Roslin 3000-rad RH panel (BovGen RH map) and contains 3966 markers including 2473 new loci in addition to 262 amplified fragment-length polymorphisms (AFLP) and 1231 markers previously published with the first generation RH map. Sequences of the mapped loci were aligned with published bovine genome maps to identify inconsistencies. In addition to differences in the order of loci, several cases were observed where the chromosomal assignment of loci differed between maps. All the chromosome maps were aligned with the current 6x bovine assembly (Btau_2.0) and 2898 loci were unambiguously located in the bovine sequence. The order of loci on the RH map for BTA 5, 7, 16, 22, 25 and 29 differed substantially from the assembled bovine sequence. From the 2898 loci unambiguously identified in the bovine sequence assembly, 131 mapped to different chromosomes in the BovGen RH map. CONCLUSION: Alignment of the BovGen RH map with other published RH and genetic maps showed higher consistency in marker order and chromosome assignment than with the current 6x sequence assembly. This suggests that the bovine sequence assembly could be significantly improved by incorporating additional independent mapping information.

Histone-specific RNA 3' processing in nuclear extracts from mammalian cells.

The mature 3' ends of histone mRNAs are formed by endonucleolytic cleavage of longer precursor transcripts. This process occurs in the nucleus and can be regarded as the equivalent of the polyadenylation reaction involved in 3′-end-generation of all other mRNAs. A sea urchin H3 gene that failed to be properly processed in the Xenopus oocyte system proved particularly useful, because it allowed the identification of a processing component from sea urchins by a complementation assay. Nuclear extracts prepared from cells under various growth conditions have helped to reveal proliferation-dependent changes in the efficiency of histone RNA 3′ processing. RNA substrates for in vitro processing are best prepared by runoff transcription of specific DNA templates with bacterial or phage RNA polymerases. For this purpose, a restriction fragment containing the 3′-terminal region of a histone gene and including the conserved palindrome and spacer motifs is cloned into a polylinker sequence downstream of a strong promoter.

Mutations within the FGF5 gene are associated with hair length in cats

Hereditary hair length variability in mice and dogs is caused by mutations within the fibroblast growth factor 5 (FGF5) gene. The aim of this study was to evaluate the feline FGF5 orthologue as a functional candidate gene for the long hair phenotype in cats, which is recessive to short hair. We amplified the feline FGF5 cDNA and characterised two alternatively spliced transcripts by RT-PCR. Comparative cDNA and genomic DNA sequencing of long- and short-haired cats revealed four non-synonymous polymorphisms in the FGF5 coding sequence. A missense mutation (AM412646:c.194C>A) was found in the homozygous state in 25 long-haired Somali, Persian, Maine Coon, Ragdoll and crossbred cats. Fifty-five short-haired cats had zero or one copy of this allele. Additionally, we found perfect co-segregation of the c.194C>A mutation within two independent pedigrees segregating for hair length. A second FGF5 exon 1 missense mutation (AM412646:c.182T>A) was found exclusively in long-haired Norwegian Forest cats. The c.182T>A mutation probably represents a second FGF5 mutation responsible for long hair in cats. In addition to the c.194C>A mutation, a frameshift mutation (AM412646:c.474delT) was found with a high frequency in the long-haired Maine Coon breed. Finally, a missense mutation (AM412646:c.475A>C) was also associated with the long-haired phenotype in some breeds. However, as one short-haired cat was homozygous for this polymorphism, it is unlikely that it has a functional role in the determination of hair length.

Influence of growth hormone (GH) receptor deletion of exon 3 and full-length isoforms on GH response and final height in patients with severe GH deficiency

CONTEXT: A polymorphism of the GH receptor (GHR) gene resulting in genomic deletion of exon 3 (GHR-d3) has been associated with responsiveness to GH therapy. However, the data reported so far do vary according to the underlying condition, replacement dose, and duration of the treatment. OBJECTIVE, DESIGN: The aim of this study was to analyze the impact of the GHR genotypes in terms of the initial height velocity (HV) resulting from treatment and the impact upon adult height in patients suffering from severe isolated GH deficiency. CONTROLS, PATIENTS, SETTING: A total of 181 subjects (peak stimulated GHlength/full-length genotype (P<0.05). A GHR-d3 allele dose-dependent effect was found for both HV SDS (r=0.72) and height gain (r=0.77). However, there was no significant difference in final adult height and height SDS according to the exon-3 genotypes. CONCLUSIONS: Our results indicate that in patients with severe isolated GH deficiency, although the GHR genotype might play a role in GH responsiveness, at least at the beginning of treatment, there is no effect on final height.