Genomic Skimming and Nanopore Sequencing Uncover Cryptic Hybridization in One of World’s Most Threatened Primates

Malukiewicz, Joanna; Cartwright, Reed A.; Dergam, Jorge Abdala; Igayara, Claudia S; Nicola, Patrícia Avello; Pereira, Luiz M C; Ruiz‐Miranda, Carlos R.; Stone, Anne C.; Silva, Daniel L.; Silva, Fernanda de Fátima Rodrigues da; Varsani, Arvind; Walter, Lutz; Wilson, Melissa A.; Zinner, Dietmar; Roos, Christian

doi:10.1101/2021.04.16.440058

Cited by 3 publications

(3 citation statements)

References 33 publications

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“…Recently, the mitogenome of the related Brazilian buffy-tufted-ear marmoset (Callithrix aurita) was assembled by genomic skimming with a nanopore sequencer [27]. Genomic skimming uses low coverage sequencing and leverages naturally high copy number sequences (e.g.…”

Section: Discussionmentioning

confidence: 99%

The mitochondrial genome and Epigenome of the Golden lion Tamarin from fecal DNA using Nanopore adaptive sequencing

et al. 2021

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Background The golden lion tamarin (Leontopithecus rosalia) is an endangered Platyrrhine primate endemic to the Atlantic coastal forests of Brazil. Despite ongoing conservation efforts, genetic data on this species remains scarce. Complicating factors include limitations on sample collection and a lack of high-quality reference sequences. Here, we used nanopore adaptive sampling to resequence the L. rosalia mitogenome from feces, a sample which can be collected non-invasively. Results Adaptive sampling doubled the fraction of both host-derived and mitochondrial sequences compared to sequencing without enrichment. 258x coverage of the L. rosalia mitogenome was achieved in a single flow cell by targeting the unfinished genome of the distantly related emperor tamarin (Saguinus imperator) and the mitogenome of the closely related black lion tamarin (Leontopithecus chrysopygus). The L. rosalia mitogenome has a length of 16,597 bp, sharing 99.68% sequence identity with the L. chrysopygus mitogenome. A total of 38 SNPs between them were identified, with the majority being found in the non-coding D-loop region. DNA methylation and hydroxymethylation were directly detected using a neural network model applied to the raw signal from the MinION sequencer. In contrast to prior reports, DNA methylation was negligible in mitochondria in both CpG and non-CpG contexts. Surprisingly, a quarter of the 642 CpG sites exhibited DNA hydroxymethylation greater than 1% and 44 sites were above 5%, with concentration in the 3′ side of several coding regions. Conclusions Overall, we report a robust new mitogenome assembly for L. rosalia and direct detection of cytosine base modifications in all contexts.

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Section: Discussionmentioning

confidence: 99%

The mitochondrial genome and Epigenome of the Golden lion Tamarin from fecal DNA using Nanopore adaptive sequencing

et al. 2021

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“…A future direction could also involve the development of a machine-learning phone app to help biological and clinical workers easily identify marmosets. Ideally, phenotypic data should be combined with mitochondrial and nuclear genome data in identification and classification of marmosets, as phenotypic data is not fully reliable to this end as cryptic hybridization does occur in marmosets [25,65].…”

Section: Implications Of Understanding Marmoset Hybrid Pelage and Mor...mentioning

confidence: 99%

Pelage Variation and Morphometrics of Closely RelatedCallithrixMarmoset Species and Their Hybrids

Malukiewicz

Warren

Boere

et al. 2023

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Background: Hybrids are expected to show greater phenotypic variation than their parental species, yet how hybrid phenotype expression varies with genetic distances in closely-related parental species remains surprisingly understudied. Here we study pelage and morphometric trait variation in anthropogenic hybrids between four species of BrazilianCallithrixmarmosets, a relatively recent primate radiation. Marmoset species are distinguishable by pelage phenotype and level of morphological specialization for eating tree exudates. Here, we (1) describe qualitative phenotypic pelage differences between parental species and hybrids; (2) test whether significant quantitative differences exist between parental and hybrid morphometric phenotypes; and (3) determine which hybrid morphometic traits show heterosis, dysgenesis, trangression, or intermediacy relative to the parental trait. For morphometric traits, we investigated both cranial and post-cranial traits, particularly as most hybrid morphological studies focus on the former instead of the latter. Finally, we estimate mitogenomic distances between marmoset species from previously published data. Results: Hybrid facial and overall body pelage variation reflected coloration and patterns seen in parental species. In morphometric traits,C. jacchusandC. penicillatawere the most similar to each other, whileC. auritawas the most distinct, andC. geoffroyitrait measures fell between these other species. Most traits inC. jacchusxC. penicillatahybrids showed either heterosis or were intermediate relative to the parental trait values. We observed heterosis and dygenesis in traits ofC. penicillataxC. geoffroyihybrids. Trangressive segregation was observed in hybrids ofC. auritaand the other species. These hybrids were alsoC. aurita-like for a number of traits. Genetic distance was closest betweenC. jacchusandC. penicillataand farthest betweenC. auritaand the other species. Conclusion: We attributed significant phenotypic differences between marmoset species to differences in morphological exudivory specialization in these species. Our results suggest that intermediate hybrid traits relative to the parental trait values are more likely in crosses between species with relatively lesser genetic distance. More extreme phenotypic variation is more likely in parental species with greater genetic distance, with transgressive traits appearing in hybrids of the most genetically distant parental species. We further suggest that that less developmental disturbances can be expected in hybrids of more recently diverged parental species.

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“…This is the first report of nuclear DNA methylation assessment using genome skimming. Nanopore sequencing is revitalizing genome skimming in ecology with detection of cryptic hybridization of threatened primates and in non-invasive methods to assemble mitogenomes 16,17 . Extending its utility by adding DNA methylation capability will aid the use of genome skimming in epidemiology, conservation, and comparative genomics.…”

Section: Introductionmentioning

confidence: 99%

Genome Skimming with Nanopore Sequencing Precisely Determines Global and Transposon DNA Methylation in Vertebrates

Faulk

2023

Preprint

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Genome skimming is defined as low-pass sequencing below 0.05X coverage and is typically used for mitochondrial genome recovery and species identification. Long read nanopore sequencers enable simultaneous reading of both DNA sequence and methylation and can multiplex samples for low-cost genome skimming. Here I present nanopore sequencing as a highly precise platform for global DNA methylation and transposon assessment. At coverage of just 0.001X, or 30 Mb of reads, accuracy is sub-1%. Biological and technical replicates validate high precision. Skimming 40 vertebrate species reveals conserved patterns of global methylation consistent with whole genome bisulfite sequencing and an average mapping rate above 97%. Genome size directly correlates to global DNA methylation, explaining 44% of its variance. Accurate SINE and LINE transposon methylation in both mouse and primates can be obtained with just 0.0001X coverage, or 3 Mb of reads. Sample multiplexing, field portability, and the low price of this instrument combine to make genome skimming for DNA methylation an accessible method for epigenetic assessment from ecology to epidemiology, and by low resource groups.

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Genomic Skimming and Nanopore Sequencing Uncover Cryptic Hybridization in One of World’s Most Threatened Primates

Cited by 3 publications

References 33 publications

The mitochondrial genome and Epigenome of the Golden lion Tamarin from fecal DNA using Nanopore adaptive sequencing

The mitochondrial genome and Epigenome of the Golden lion Tamarin from fecal DNA using Nanopore adaptive sequencing

Pelage Variation and Morphometrics of Closely RelatedCallithrixMarmoset Species and Their Hybrids

Genome Skimming with Nanopore Sequencing Precisely Determines Global and Transposon DNA Methylation in Vertebrates

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