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The availability of genetic data from wild populations limits our understanding of primate evolution and conservation, particularly for small nocturnal species such as lorisiforms (galagos, lorises, angwantibos, and pottos). Emerging methods for recovering genomic DNA from historical museum specimens have been rarely used in primate studies. We aimed to optimize extraction and bioinformatics protocols to maximize the recovery of historical DNA to fill important geographic and taxonomic gaps, improve phylogenetic resolution, and inform conservation of Lorisiform primates. First, we compared the performance of two DNA extraction methods by using 238 specimens up to a hundred years old. We then selected 96 samples with the highest DNA yields for shotgun sequencing. To evaluate the impact of phylogenetic divergence in bioinformatic read mapping, we compared coverage depths when using human and three lorisiform reference mitogenomes. Based on whole genomic data, we performed metagenomics and microbial diversity analyses to assess the composition of potentially exogenous content. Lastly, based on the most geographically and taxonomically comprehensive sampling for the West African lorisiforms to date (19/32 currently recognized species), we performed phylogenetic inference using Maximum Likelihood. The results showed that older samples yield lower DNA concentration, with an optimized phenol-chloroform protocol outperforming a commercial kit. However, both extraction methods generated DNA in sufficient amount and quality for phylogenetic inference. Our reference bias comparisons showed that higher phylogenetic proximity between focal species and reference mitogenome increases coverage depth. The metagenomic analysis found human contamination in only one of 96 samples (1%), whereas ten of 96 (11%) samples showed nonnegligible levels of other exogenous contents, among which are certain blood parasites. We inferred low support for the monophyly of Asian and African Lorisids but confirmed the monophyly and previously suggested relationships among Galagid genera. Lastly, we found evidence of cryptic species diversity within the western dwarf galagos (genus Galagoides). Taken together, these results attest to the enormous potential of museomics to advance our understanding of galago evolution, ecology, and conservation, an approach that can be extended to other primate clades.
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Molecular Phylogenetic Relationships and Unveiling Novel Genetic Diversity among Slow and Pygmy Lorises, including Resurrection of Xanthonycticebus intermedius
Genetic analysis of historical museum collections presents an opportunity to clarify the evolutionary history of understudied primate groups, improve taxonomic inferences, and inform conservation efforts. Among the most understudied primate groups, slow and pygmy lorises (genera Nycticebus and Xanthonycticebus) are nocturnal strepsirrhines found in South and Southeast Asia. Previous molecular studies have supported five species, but studies using morphological data suggest the existence of at least nine species. We sequenced four mitochondrial loci, CO1, cytb, d-loop, and ND4, for a total of 3324 aligned characters per sample from 41 historical museum specimens for the most comprehensive geographic coverage to date for these genera. We then combined these sequences with a larger dataset composed of samples collected in Vietnam as well as previously published sequences (total sample size N = 62). We inferred phylogenetic relationships using Bayesian inference and maximum likelihood methods based on data from each locus and on concatenated sequences. We also inferred divergence dates for the most recent common ancestors of major lineages using a BEAST analysis. Consistent with previous studies, we found support for Xanthonycticebus pygmaeus as a basal taxon to the others in the group. We also confirmed the separation between lineages of X. pygmaeus from northern Vietnam/Laos/China and southern Vietnam/Cambodia and included a taxonomic revision recognizing a second taxon of pygmy loris, X. intermedius. Our results found support for multiple reciprocally monophyletic taxa within Borneo and possibly Java. The study will help inform conservation management of these trade-targeted animals as part of a genetic reference database for determining the taxonomic unit and provenance of slow and pygmy lorises confiscated from illegal wildlife trade activities.
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- PAR ID:
- 10421628
- Date Published:
- Journal Name:
- Genes
- Volume:
- 14
- Issue:
- 3
- ISSN:
- 2073-4425
- Page Range / eLocation ID:
- 643
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3390/genes14030643
