Epstein-Barr virus (EBV) persists in human B-cells by maintaining its chromatinized episomes within the nucleus. We have previously shown that cellular factor Poly [ADP-ribose] polymerase 1 (PARP1) binds the EBV genome, stabilizes CTCF binding at specific loci, and that PARP1 enzymatic activity correlates with maintaining a transcriptionally active latency program. To better understand PARP1’s role in regulating EBV latency, here we functionally characterize the effect of PARP enzymatic inhibition on episomal structure through in situ HiC mapping, generating a complete 3D structure of the EBV genome. We also map intragenomic contact changes after PARP inhibition to global binding of chromatin looping factors CTCF and cohesin across the EBV genome. We find that PARP inhibition leads to fewer total unique intragenomic interactions within the EBV episome, yet new chromatin loops distinct from the untreated episome are also formed. This study also illustrates that PARP inhibition alters gene expression at the regions where chromatin looping is most effected. We observe that PARP1 inhibition does not alter cohesin binding sites but does increase its frequency of binding at those sites. Taken together, these findings demonstrate that PARP has an essential role in regulating global EBV chromatin structure and latent gene expression.

Formalin fixation, albeit an outstanding method for morphological and molecular preservation, induces DNA damage and cross-linking, which can hinder nucleic acid screening. This is of particular concern in the detection of low-abundance targets, such as persistent DNA viruses. In the present study, we evaluated the analytical sensitivity of viral detection in lung, liver, and kidney specimens from four deceased individuals. The samples were either frozen or incubated in formalin (±paraffin embedding) for up to 10 days. We tested two DNA extraction protocols for the control of efficient yields and viral detections. We used short-amplicon qPCRs (63–159 nucleotides) to detect 11 DNA viruses, as well as hybridization capture of these plus 27 additional ones, followed by deep sequencing. We observed marginally higher ratios of amplifiable DNA and scantly higher viral genoprevalences in the samples extracted with the FFPE dedicated protocol. Based on the findings in the frozen samples, most viruses were detected regardless of the extended fixation times. False-negative calls, particularly by qPCR, correlated with low levels of viral DNA (<250 copies/million cells) and longer PCR amplicons (>150 base pairs). Our data suggest that low-copy viral DNAs can be satisfactorily investigated from FFPE specimens, and encourages further examination of historical materials.

Genomic-scale data for non-model taxa are providing new insights into landscape genomic structuring and species limits, leading to more informed conservation decisions, particularly in taxa with extremely restricted microhabitat preferences and small geographic distributions. This study applied sequence capture of ultraconserved elements (UCEs) to gather genomic-scale data for two federally endangered Texella harvester species distributed in Edwards Formation cave and karst habitats of central Texas, near Austin. We gathered UCE data for 51 T. reyesi specimens from 46 different caves, seven T. reddelli specimens from five caves, and from relevant outgroup species. For these UCE data we applied a combination of phylogenomic, multispecies coalescent phylogenetic, and single-nucleotide polymorphism machine-learning analyses. We found that samples of T. reddelli and T. reyesi together form a single clade in phylogenetic analyses, but that T. reddelli samples are not recovered as monophyletic. Instead, T. reddelli samples from three northern caves are embedded within a larger T. reyesi genetic clade. Significantly, the genetic structuring of all samples closely follows geologic barriers defined for the region and formalized as karst fauna regions (KFRs). One exception is the Jollyville Plateau KFR, which includes two divergent, non-sister genetic lineages. Levels of troglomorphy, here assessed by a simple scoring of corneal and retinal development, also closely follows clade (and geographic) boundaries, implying that divergent genetic lineages might also have distinct ecologies. Overall, our study has important taxonomic implications, is the first to explore (and validate) regional KFR boundaries using intraspecific genetic data, and provides essential data for future management decisions involving these federally endangered species.

The major transmission route for SARS-CoV-2 is airborne. However, previous studies could not elucidate the contribution between large droplets and aerosol transmission of SARS-CoV-2 and its variants. Here, we designed and validated an optimized transmission caging setup, which allows for the assessment of aerosol transmission efficiency at various distances. At a distance of 2 m, only particles of <5 μm traversed between cages. Using this setup, we investigated the relative efficiency of aerosol transmission between the SARS-CoV-2 Alpha variant (B.1.1.7) and lineage A in Syrian hamsters. Aerosol transmission of both variants was confirmed in all sentinels after 24 h of exposure as demonstrated by respiratory virus shedding and seroconversion. Productive transmission also occurred after 1 h of exposure, highlighting the efficiency of this transmission route. Interestingly, after donors were infected with a mix of both variants, the Alpha variant outcompeted the lineage A variant in an airborne transmission chain. Overall, these data indicate that a lower infectious dose of the Alpha variant, compared to lineage A, could be sufficient for successful transmission. This highlights the continuous need to assess emerging variants and the development for pre-emptive transmission mitigation strategies.

Resolution of rapid evolutionary radiations requires harvesting maximal signal from phylogenomic datasets. However, studies of non-model clades often target conserved loci that are characterized by reduced information content, which can negatively affect gene tree precision and species tree accuracy. Single nucleotide polymorphism (SNP)-based methods are an underutilized but potentially valuable tool for estimating phylogeny and divergence times because they do not rely on resolved gene trees, allowing information from many or all variant loci to be leveraged in species tree reconstruction. We evaluated the utility of SNP-based methods in resolving phylogeny of Holarctic ground squirrels (Urocitellus), a radiation that has been difficult to disentangle, even in prior phylogenomic studies. We inferred phylogeny from a dataset of >3,000 ultraconserved element loci (UCEs) using two methods (SNAPP, SVDquartets) and compared our results with a new mitogenome phylogeny. We also systematically evaluated how phasing of UCEs improves per-locus information content, and inference of topology and other parameters within each of these SNP-based methods. Phasing improved topological resolution and branch length estimation at shallow levels (within species complexes), but less so at deeper levels, likely reflecting true uncertainty due to ancestral polymorphisms segregating in these rapidly diverging lineages. We resolved several key clades in Urocitellus and present targeted opportunities for future phylogenomic inquiry. Our results extend the roadmap for use of SNPs to address vertebrate radiations and support comparative analyses at multiple temporal scales.

Traditionally, paleontologists have relied on the morphological features of bones and teeth to reconstruct the evolutionary relationships of extinct animals.1 In recent decades, the analysis of ancient DNA recovered from macrofossils has provided a powerful means to evaluate these hypotheses and develop novel phylogenetic models.2 Although a great deal of life history data can be extracted from bones, their scarcity and associated biases limit their information potential. The paleontological record of Beringia3—the unglaciated areas and former land bridge between northeast Eurasia and northwest North America—is relatively robust thanks to its perennially frozen ground favoring fossil preservation.4,5 However, even here, the macrofossil record is significantly lacking in small-bodied fauna (e.g., rodents and birds), whereas questions related to migration and extirpation, even among well-studied taxa, remain crudely resolved. The growing sophistication of ancient environmental DNA (eDNA) methods have allowed for the identification of species within terrestrial/aquatic ecosystems,6, 7, 8, 9, 10, 11, 12 in paleodietary reconstructions,13, 14, 15, 16, 17, 18, 19 and facilitated genomic reconstructions from cave contexts.8,20, 21, 22 Murchie et al.6,23 used a capture enrichment approach to sequence a diverse range of faunal and floral DNA from permafrost silts deposited during the Pleistocene-Holocene transition.24 Here, we expand on their work with the mitogenomic assembly and phylogenetic placement of Equus caballus (caballine horse), Bison priscus (steppe bison), Mammuthus primigenius (woolly mammoth), and Lagopus lagopus (willow ptarmigan) eDNA from multiple permafrost cores spanning the last 40,000 years. We identify a diverse metagenomic spectra of Pleistocene fauna and identify the eDNA co-occurrence of distinct Eurasian and American mitogenomic lineages.

Abstract Island biogeography is one of the most powerful subdisciplines of ecology: its mathematical predictions that island size and distance to mainland determine diversity have withstood the test of time. A key question is whether these predictions follow at a population-genomic level. Using rigorous ancient-DNA protocols, we retrieved approximately 1,000 genomic markers from approximately 100 historic specimens of two Southeast Asian songbird complexes from across the Sunda Shelf archipelago collected 1893–1957. We show that the genetic affinities of populations on small shelf islands defy the predictions of geographic distance and appear governed by Earth-historic factors including the position of terrestrial barriers (paleo-rivers) and persistence of corridors (Quaternary land bridges). Our analyses suggest that classic island-biogeographic predictors may not hold well for population-genomic dynamics on the thousands of shelf islands across the globe, which are exposed to dynamic changes in land distribution during Quaternary climate change.

Background: Somatic mutations, copy-number variations, and genome instability of mitochondrial DNA (mtDNA) have been reported in different types of cancers and are suggested to play important roles in cancer development and metastasis. However, there is scarce information about pheochromocytomas and paragangliomas (PCCs/PGLs) formation. Material: To determine the potential roles of mtDNA alterations in sporadic PCCs/PGLs, we analyzed a panel of 26 nuclear susceptibility genes and the entire mtDNA sequence of seventy-seven human tumors, using next-generation sequencing, and compared the results with normal adrenal medulla tissues. We also performed an analysis of copy-number alterations, large mtDNA deletion, and gene and protein expression. Results: Our results revealed that 53.2% of the tumors harbor a mutation in at least one of the targeted susceptibility genes, and 16.9% harbor complementary mitochondrial mutations. More than 50% of the mitochondrial mutations were novel and predicted pathogenic, affecting mitochondrial oxidative phosphorylation. Large deletions were found in 26% of tumors, and depletion of mtDNA occurred in more than 87% of PCCs/PGLs. The reduction of the mitochondrial number was accompanied by a reduced expression of the regulators that promote mitochondrial biogenesis (PCG1α, NRF1, and TFAM). Further, P62 and LC3a gene expression suggested increased mitophagy, which is linked to mitochondrial dysfunction. Conclusion: The pathogenic role of these finding remains to be shown, but we suggest a complementarity and a potential contributing role in PCCs/PGLs tumorigenesis.

Although wheat (Triticum aestivum L.) is the main staple crop in the world and a major source of carbohydrates and proteins, functional genomics and allele mining are still big challenges. Given the advances in next-generation sequencing (NGS) technologies, the identification of causal variants associated with a target phenotype has become feasible. For these reasons, here, by combining sequence capture and target-enrichment methods with high-throughput NGS re-sequencing, we were able to scan at exome-wide level 46 randomly selected bread wheat individuals from a recombinant inbred line population and to identify and classify a large number of single nucleotide polymorphisms (SNPs). For technical validation of results, eight randomly selected SNPs were converted into Kompetitive Allele-Specific PCR (KASP) markers. This resource was established as an accessible and reusable molecular toolkit for allele data mining. The dataset we are making available could be exploited for novel studies on bread wheat genetics and as a foundation for starting breeding programs aimed at improving different key agronomic traits.