GCalignR: An R package for aligning gas-chromatography data for ecological and evolutionary studies.

Chemical cues are arguably the most fundamental means of animal communication and play an important role in mate choice and kin recognition. Consequently, there is growing interest in the use of gas chromatography (GC) to investigate the chemical basis of eco-evolutionary interactions. Both GC-MS (mass spectrometry) and FID (flame ionization detection) are commonly used to characterise the chemical composition of biological samples such as skin swabs. The resulting chromatograms comprise peaks that are separated according to their retention times and which represent different substances. Across chromatograms of different samples, homologous substances are expected to elute at similar retention times. However, random and often unavoidable experimental variation introduces noise, making the alignment of homologous peaks challenging, particularly with GC-FID data where mass spectral data are lacking. Here we present GCalignR, a user-friendly R package for aligning GC-FID data based on retention times. The package was developed specifically for ecological and evolutionary studies that seek to investigate similarity patterns across multiple and often highly variable biological samples, for example representing different sexes, age classes or reproductive stages. The package also implements dynamic visualisations to facilitate inspection and fine-tuning of the resulting alignments and can be integrated within a broader workflow in R to facilitate downstream multivariate analyses. We demonstrate an example workflow using empirical data from Antarctic fur seals and explore the impact of user-defined parameter values by calculating alignment error rates for multiple datasets. The resulting alignments had low error rates for most of the explored parameter space and we could also show that GCalignR performed equally well or better than other available software. We hope that GCalignR will help to simplify the processing of chemical datasets and improve the standardization and reproducibility of chemical analyses in studies of animal chemical communication and related fields

Evidence for frequent incest in a cooperatively breeding mammal.

As breeding between relatives often results in inbreeding depression, inbreeding avoidance is widespread in the animal kingdom. However, inbreeding avoidance may entail fitness costs. For example, dispersal away from relatives may reduce survival. How these conflicting selection pressures are resolved is challenging to investigate, but theoretical models predict that inbreeding should occur frequently in some systems. Despite this, few studies have found evidence of regular incest in mammals, even in social species where relatives are spatio-temporally clustered and opportunities for inbreeding frequently arise. We used genetic parentage assignments together with relatedness data to quantify inbreeding rates in a wild population of banded mongooses, a cooperatively breeding carnivore. We show that females regularly conceive to close relatives, including fathers and brothers. We suggest that the costs of inbreeding avoidance may sometimes outweigh the benefits, even in cooperatively breeding species where strong within-group incest avoidance is considered to be the norm

inbreedR: An R package for the analysis of inbreeding based on genetic markers

1. Heterozygosity fitness correlations (HFCs) have been used extensively to explore the impact of inbreeding on individual fitness. Initially, most studies used small panels of microsatellites, but more recently with the advent of next generation sequencing, large SNP datasets are becoming increasingly available and these provide greater power and precision to quantify the impact of inbreeding on fitness. 2. Despite the popularity of HFC studies, effect sizes tend to be rather small. One reason for this may be a low variation in inbreeding level across individuals. Using genetic markers, it is possible to measure variance in inbreeding through the strength of correlation in heterozygosity across marker loci, termed identity disequilibrium (ID). 3. ID can be quantified using the measure g2 which is also a central parameter in HFC theory that can be used within a wider framework to estimate the direct impact of inbreeding on both marker heterozygosity and fitness. However, no software exists to calculate g2 for large SNP datasets nor to implement this framework. 4. inbreedR is an R package that provides functions to calculate g2 based on microsatellite and SNP markers with associated p-values and confidence intervals. Within the framework of HFC theory, inbreedR also estimates the impact of inbreeding on marker heterozygosity and fitness. Moreover, we implemented easy-to-use simulations to explore the precision and magnitude of estimates based on different numbers of genetic markers. We hope this package will facilitate good practice in the analysis of HFCs and help to deepen our understanding of inbreeding effects in natural populations

Banded mongooses avoid inbreeding when mating with members of the same natal group

Inbreeding and inbreeding avoidance are key factors in the evolution of animal societies, influencing dispersal and reproductive strategies which can affect relatedness structure and helping behaviours. In cooperative breeding systems, individuals typically avoid inbreeding through reproductive restraint and/or dispersing to breed outside their natal group. However, where groups contain multiple potential mates of varying relatedness, strategies of kin recognition and mate choice may be favoured. Here, we investigate male mate choice and female control of paternity in the banded mongoose (Mungos mungo), a cooperatively-breeding mammal where both sexes are often philopatric and mating between relatives is known to occur. We find evidence suggestive of inbreeding depression in banded mongooses, indicative of a benefit to avoiding breeding with relatives. Successfully breeding pairs were less related than expected under random mating, which appeared to be driven by both male choice and female control of paternity. Male banded mongooses actively guard females to gain access to mating opportunities, and this guarding behaviour is preferentially directed towards less closely related females. Guard-female relatedness did not affect the guard’s probability of gaining reproductive success. However, where mate-guards are unsuccessful they lose paternity to males that are less related to the females than themselves. Together our results suggest that both sexes of banded mongoose use kin discrimination to avoid inbreeding. Although this strategy appears to be rare among cooperative breeders, it may be more prominent in species where relatedness to potential mates is variable, and/or where opportunities for dispersal and mating outside of the group are limited

Heterogeneous Genomic Divergence Landscape in Two Commercially Important European Scallop Species

Two commercially important scallop species of the genus Pecten are found in Europe: the north Atlantic Pecten maximus and the Mediterranean Pecten jacobaeus whose distributions abut at the Almeria–Orán front. Whilst previous studies have quantified genetic divergence between these species, the pattern of differentiation along the Pecten genome is unknown. Here, we mapped RADseq data from 235 P. maximus and 27 P. jacobaeus to a chromosome-level reference genome, finding a heterogeneous landscape of genomic differentiation. Highly divergent genomic regions were identified across 14 chromosomes, while the remaining five showed little differentiation. Demographic and comparative genomics analyses suggest that this pattern resulted from an initial extended period of isolation, which promoted divergence, followed by differential gene flow across the genome during secondary contact. Single nucleotide polymorphisms present within highly divergent genomic regions were located in areas of low recombination and contrasting patterns of LD decay were found between the two species, hinting at the presence of chromosomal inversions in P. jacobaeus. Functional annotations revealed that highly differentiated regions were enriched for immune-related processes and mRNA modification. While future work is necessary to characterize structural differences, this study provides new insights into the speciation genomics of P. maximus and P. jacobaeus

Unexpected Fine-Scale Population Structure in a Broadcast-Spawning Antarctic Marine Mollusc

Several recent empirical studies have challenged the prevailing dogma that broadcast-spawning species exhibit little or no population genetic structure by documenting genetic discontinuities associated with large-scale oceanographic features. However, relatively few studies have explored patterns of genetic differentiation over fine spatial scales. Consequently, we used a hierarchical sampling design to investigate the basis of a weak but significant genetic difference previously reported between Antarctic limpets (Nacella concinna) sampled from Adelaide and Galindez Islands near the base of the Antarctic Peninsula. Three sites within Ryder Bay, Adelaide Island (Rothera Point, Leonie and Anchorage Islands) were each sub-sampled three times, yielding a total of 405 samples that were genotyped at 155 informative Amplified Fragment Length Polymorphisms (AFLPs). Contrary to our initial expectations, limpets from Anchorage Island were found to be subtly, but significantly distinct from those sampled from the other sites. This suggests that local processes may play an important role in generating fine-scale population structure even in species with excellent dispersal capabilities, and highlights the importance of sampling at multiple spatial scales in population genetic surveys

A Novel Approach for Mining Polymorphic Microsatellite Markers In Silico

An important emerging application of high-throughput 454 sequencing is the isolation of molecular markers such as microsatellites from genomic DNA. However, few studies have developed microsatellites from cDNA despite the added potential for targeting candidate genes. Moreover, to develop microsatellites usually requires the evaluation of numerous primer pairs for polymorphism in the focal species. This can be time-consuming and wasteful, particularly for taxa with low genetic diversity where the majority of primers often yield monomorphic polymerase chain reaction (PCR) products. Transcriptome assemblies provide a convenient solution, functional annotation of transcripts allowing markers to be targeted towards candidate genes, while high sequence coverage in principle permits the assessment of variability in silico. Consequently, we evaluated fifty primer pairs designed to amplify microsatellites, primarily residing within transcripts related to immunity and growth, identified from an Antarctic fur seal (Arctocephalus gazella) transcriptome assembly. In silico visualization was used to classify each microsatellite as being either polymorphic or monomorphic and to quantify the number of distinct length variants, each taken to represent a different allele. The majority of loci (n = 36, 76.0%) yielded interpretable PCR products, 23 of which were polymorphic in a sample of 24 fur seal individuals. Loci that appeared variable in silico were significantly more likely to yield polymorphic PCR products, even after controlling for microsatellite length measured in silico. We also found a significant positive relationship between inferred and observed allele number. This study not only demonstrates the feasibility of generating modest panels of microsatellites targeted towards specific classes of gene, but also suggests that in silico microsatellite variability may provide a useful proxy for PCR product polymorphism

High glucose up-regulates ENaC and SGK1 expression in HCD-cells

Background/Aim: Diabetic nephropathy is associated with progressive renal damage, leading to impaired function and end-stage renal failure. Secondary hypertension stems from a deranged ability of cells within the kidney to resolve and appropriately regulate sodium resorption in response to hyperglycaemia. However, the mechanisms by which glucose alters sodium re-uptake have not been fully characterised. Methods: Here we present RT-PCR, western blot and immunocytochemistry data confirming mRNA and protein expression of the serum and glucocorticoid inducible kinase (SGK1) and the a conducting subunit of the epithelial sodium channel (ENaC) in a model in vitro system of the human cortical collecting duct (HCD). We examined changes in expression of these elements in response to glucose challenge, designed to mimic hyperglycaemia associated with type 2 diabetes mellitus. Changes in Na+ concentration were assessed using single-cell microfluorimetry. Results: Incubation with glucose, the Ca2+-ionophore ionomycin and the cytokine TGF-beta 1 were all found to evoke significant and time-dependent increases in both SGK1 and alpha ENaC protein expression. These molecular changes were correlated to an increase in Na+-uptake at the single-cell level. Conclusion: Together these data offer a potential explanation for glucose-evoked Na+-resorption and a potential contributory role of SGK1 and ENaCs in development of secondary hypertension, commonly linked to diabetic nephropathy

The genetic legacy of extreme exploitation in a polar vertebrate

Understanding the effects of human exploitation on the genetic composition of wild populations is important for predicting species persistence and adaptive potential. We therefore investigated the genetic legacy of large-scale commercial harvesting by reconstructing, on a global scale, the recent demographic history of the Antarctic fur seal (Arctocephalus gazella), a species that was hunted to the brink of extinction by 18th and 19th century sealers. Molecular genetic data from over 2,000 individuals sampled from all eight major breeding locations across the species’ circumpolar geographic distribution, show that at least four relict populations around Antarctica survived commercial hunting. Coalescent simulations suggest that all of these populations experienced severe bottlenecks down to effective population sizes of around 150–200. Nevertheless, comparably high levels of neutral genetic variability were retained as these declines are unlikely to have been strong enough to deplete allelic richness by more than around 15%. These findings suggest that even dramatic short-term declines need not necessarily result in major losses of diversity, and explain the apparent contradiction between the high genetic diversity of this species and its extreme exploitation history

A preliminary study of genetic factors that influence susceptibility to bovine tuberculosis in the British cattle herd

Associations between specific host genes and susceptibility to Mycobacterial infections such as tuberculosis have been reported in several species. Bovine tuberculosis (bTB) impacts greatly the UK cattle industry, yet genetic predispositions have yet to be identified. We therefore used a candidate gene approach to study 384 cattle of which 160 had reacted positively to an antigenic skin test (‘reactors’). Our approach was unusual in that it used microsatellite markers, embraced high breed diversity and focused particularly on detecting genes showing heterozygote advantage, a mode of action often overlooked in SNP-based studies. A panel of neutral markers was used to control for population substructure and using a general linear model-based approach we were also able to control for age. We found that substructure was surprisingly weak and identified two genomic regions that were strongly associated with reactor status, identified by markers INRA111 and BMS2753. In general the strength of association detected tended to vary depending on whether age was included in the model. At INRA111 a single genotype appears strongly protective with an overall odds ratio of 2.2, the effect being consistent across nine diverse breeds. Our results suggest that breeding strategies could be devised that would appreciably increase genetic resistance of cattle to bTB (strictly, reduce the frequency of incidence of reactors) with implications for the current debate concerning badger-culling