Time-calibrated phylogeny and full mitogenome sequence of the Galapagos sea lion (Zalophus wollebaeki) from scat DNA.

Background: The Galapagos sea lion, Zalophus wollebaeki, is an endemic and endangered otariid, which is considered as a sentinel species of ecosystem dynamics in the Galapagos archipelago. Mitochondrial DNA is an important tool in phylogenetic and population genetic inference. In this work we use Illumina sequencing to complement the mitogenomic resources for Zalophus genus-the other two species employed Sanger sequencing-by a complete mitochondrial genome and a molecular clock of this species, which is not present in any case. Materials and Methods: We used DNA obtained from a fresh scat sample of a Galapagos sea lion and shotgun-sequenced it on the Illumina NextSeq platform. The obtained raw reads were processed using the GetOrganelle software to filter the mitochondrial Zalophus DNA reads (∼16% survive the filtration), assemble them, and set up a molecular clock. Results: From the obtained 3,511,116 raw reads, we were able to assemble a full mitogenome of a length of 16,676 bp, consisting of 13 protein-coding genes (PCGs), 22 transfer RNAs (tRNA), and two ribosomal RNAs (rRNA). A time-calibrated phylogeny confirmed the phylogenetic position of Z. wollebaeki in a clade with Z. californianus, and Z. japonicus, and sister to Z. californianus; as well as establishing the divergence time for Z. wollebaeki 0.65 million years ago. Our study illustrates the possibility of seamlessly sequencing full mitochondrial genomes from fresh scat samples of marine mammals.

Epigenetic effects of parasites and pesticides on captive and wild nestling birds.

Anthropogenic changes to the environment challenge animal populations to adapt to new conditions and unique threats. While the study of adaptation has focused on genetic variation, epigenetic mechanisms may also be important. DNA methylation is sensitive to environmental stressors, such as parasites and pesticides, which may affect gene expression and phenotype. We studied the effects of an invasive ectoparasite, Philornis downsi, on DNA methylation of Galápagos mockingbirds (Mimus parvulus). We used the insecticide permethrin to manipulate P. downsi presence in nests of free-living mockingbirds and tested for effects of parasitism on nestling mockingbirds using epiGBS, a reduced-representation bisulfite sequencing (RRBS) approach. To distinguish the confounding effects of insecticide exposure, we conducted a matching experiment exposing captive nestling zebra finches (Taeniopygia guttata) to permethrin. We used zebra finches because they were the closest model organism to mockingbirds that we could breed in controlled conditions. We identified a limited number of differentially methylated cytosines (DMCs) in parasitized versus nonparasitized mockingbirds, but the number was not more than expected by chance. In contrast, we saw clear effects of permethrin on methylation in captive zebra finches. DMCs in zebra finches paralleled documented effects of permethrin exposure on vertebrate cellular signaling and endocrine function. Our results from captive birds indicate a role for epigenetic processes in mediating sublethal nontarget effects of pyrethroid exposure in vertebrates. Environmental conditions in the field were more variable than the laboratory, which may have made effects of both parasitism and permethrin harder to detect in mockingbirds. RRBS approaches such as epiGBS may be a cost-effective way to characterize genome-wide methylation profiles. However, our results indicate that ecological epigenetic studies in natural populations should consider the number of cytosines interrogated and the depth of sequencing in order to have adequate power to detect small and variable effects.

Relieving efforts in palm-tree tissue sampling for population genetics analyses.

The young leaves are the main source of nucleic acids for population genetic studies in palm-trees; however, the access to this tissue may be limited by specific features of each species. Using root tissues as an alternative source of nucleic acids could facilitate the sampling in large populations.This study tests root tissue viability as an alternative nucleic acid source (root versus. leaf) and explores different protocols (tissue storage and DNA extraction methods) to obtain high-quality DNA samples.The results showed no significant differences in DNA concentration (603.7 vs. 599.1 ng/µl) and quality ratios (A260/280:2.1 vs. 1.9, and A260/230:2.1 vs. 2.0) for the comparisons of tissue source (leaf vs. root) and DNA extraction method (manual vs. kit). For tissue storage method, DNA concentration was significantly higher for root tissues stored in 70% and 90% alcohol solutions (692.8 and 822.6 ng/µl, respectively) versus those obtained from leaf tissue (603.7 ng/µl); however, for the quality parameters, no differences were found.Results showed the effective potential of using root tissue as an alternative source for nucleic acids, which could facilitate population sampling of palm-tree species for future studies, and this methodological alternative could be applied to other plant systems with similar sampling challenges. ​.