Reduction of bitter taste receptor gene family in folivorous colobine primates relative to omnivorous cercopithecine primates.

Bitter taste perception is important in preventing animals from ingesting potentially toxic compounds. Whole-genome assembly (WGA) data have revealed that bitter taste receptor genes (TAS2Rs) comprise a multigene family with dozens of intact and disrupted genes in primates. However, publicly available WGA data are often incomplete, especially for multigene families. In this study, we employed a targeted capture (TC) approach specifically probing TAS2Rs for ten species of cercopithecid primates with diverse diets, including eight omnivorous cercopithecine species and two folivorous colobine species. We designed RNA probes for all TAS2Rs that we modeled to be intact in the common ancestor of cercopithecids ("ancestral-cercopithecid TAS2R gene set"). The TC was followed by short-read and high-depth massive-parallel sequencing. TC retrieved more intact TAS2R genes than found in WGA databases. We confirmed a large number of gene "births" at the common ancestor of cercopithecids and found that the colobine common ancestor and the cercopithecine common ancestor had contrasting trajectories: four gene "deaths" and three gene births, respectively. The number of intact TAS2R genes was markedly reduced in colobines (25-28 detected via TC and 20-26 detected via WGA analysis) as compared with cercopithecines (27-36 via TC and 19-30 via WGA). Birth or death events occurred at almost every phylogenetic-tree branch, making the composition of intact genes variable among species. These results show that evolutionary change in intact TAS2R genes is a complex process, refute a simple general prediction that herbivory favors more TAS2R genes, and have implications for understanding dietary adaptations and the evolution of detoxification abilities.

Targeted enrichment of whole-genome SNPs from highly burned skeletal remains.

Genetic assessment of highly incinerated and/or degraded human skeletal material is a persistent challenge in forensic DNA analysis, including identifying victims of mass disasters. Few studies have investigated the impact of thermal degradation on whole-genome single-nucleotide polymorphism (SNP) quality and quantity using next-generation sequencing (NGS). We present whole-genome SNP data obtained from the bones and teeth of 27 fire victims using two DNA extraction techniques. Extracts were converted to double-stranded DNA libraries then enriched for whole-genome SNPs using unpublished biotinylated RNA baits and sequenced on an Illumina NextSeq 550 platform. Raw reads were processed using the EAGER (Efficient Ancient Genome Reconstruction) pipeline, and the SNPs filtered and called using FreeBayes and GATK (v. 3.8). Mixed-effects modeling of the data suggest that SNP variability and preservation is predominantly determined by skeletal element and burn category, and not by extraction type. Whole-genome SNP data suggest that selecting long bones, hand and foot bones, and teeth subjected to temperatures <350°C are the most likely sources for higher genomic DNA yields. Furthermore, we observed an inverse correlation between the number of captured SNPs and the extent to which samples were burned, as well as a significant decrease in the total number of SNPs measured for samples subjected to temperatures >350°C. Our data complement previous analyses of burned human remains that compare extraction methods for downstream forensic applications and support the idea of adopting a modified Dabney extraction technique when traditional forensic methods fail to produce DNA yields sufficient for genetic identification.

The effect of missing data on evolutionary analysis of sequence capture bycatch, with application to an agricultural pest.

Sequence capture is a genomic technique that selectively enriches target sequences before high throughput next-generation sequencing, to generate specific sequences of interest. Off-target or 'bycatch' data are often discarded from capture experiments, but can be leveraged to address evolutionary questions under some circumstances. Here, we investigated the effects of missing data on a variety of evolutionary analyses using bycatch from an exon capture experiment on the global pest moth, Helicoverpa armigera. We added > 200 new samples from across Australia in the form of mitogenomes obtained as bycatch from targeted sequence capture, and combined these into an additional larger dataset to total > 1000 mitochondrial cytochrome c oxidase subunit I (COI) sequences across the species' global distribution. Using discriminant analysis of principal components and Bayesian coalescent analyses, we showed that mitogenomes assembled from bycatch with up to 75% missing data were able to return evolutionary inferences consistent with higher coverage datasets and the broader literature surrounding H. armigera. For example, low-coverage sequences broadly supported the delineation of two H. armigera subspecies and also provided new insights into the potential for geographic turnover among these subspecies. However, we also identified key effects of dataset coverage and composition on our results. Thus, low-coverage bycatch data can offer valuable information for population genetic and phylodynamic analyses, but caution is required to ensure the reduced information does not introduce confounding factors, such as sampling biases, that drive inference. We encourage more researchers to consider maximizing the potential of the targeted sequence approach by examining evolutionary questions with their off-target bycatch where possible-especially in cases where no previous mitochondrial data exists-but recommend stratifying data at different genome coverage thresholds to separate sampling effects from genuine genomic signals, and to understand their implications for evolutionary research.

Long-insert sequence capture detects high copy numbers in a defence-related beta-glucosidase gene ßglu-1 with large variations in white spruce but not Norway spruce.

Conifers are long-lived and slow-evolving, thus requiring effective defences against their fast-evolving insect natural enemies. The copy number variation (CNV) of two key acetophenone biosynthesis genes Ugt5/Ugt5b and ßglu-1 may provide a plausible mechanism underlying the constitutively variable defence in white spruce (Picea glauca) against its primary defoliator, spruce budworm. This study develops a long-insert sequence capture probe set (Picea_hung_p1.0) for quantifying copy number of ßglu-1-like, Ugt5-like genes and single-copy genes on 38 Norway spruce (Picea abies) and 40 P. glauca individuals from eight and nine provenances across Europe and North America respectively. We developed local assemblies (Piabi_c1.0 and Pigla_c.1.0), full-length transcriptomes (PIAB_v1 and PIGL_v1), and gene models to characterise the diversity of ßglu-1 and Ugt5 genes. We observed very large copy numbers of ßglu-1, with up to 381 copies in a single P. glauca individual. We observed among-provenance CNV of ßglu-1 in P. glauca but not P. abies. Ugt5b was predominantly single-copy in both species. This study generates critical hypotheses for testing the emergence and mechanism of extreme CNV, the dosage effect on phenotype, and the varying copy number of genes with the same pathway. We demonstrate new approaches to overcome experimental challenges in genomic research in conifer defences.