Original Genome Research Articles

Defective HIV proviruses: possible involvement in the HIV infection pathogenesis

This review article analyzes information obtained from a literature search on defective HIV genomes (HIV-1, Human Immunodeficiency Virus, Lentivirus, Orthoretrovirinae, Retroviridae). It discusses the origins of defective HIV genomes, their potential for transcription and translation, and the role of defective RNA and proteins in stimulating both innate and adaptive immunity. The article also explores their contribution to HIV pathogenesis, immune system hyperactivation despite successful antiretroviral therapy (ART), and the evolutionary processes in HIV proviral populations under ART. Additionally, it addresses challenges in reservoir elimination and HIV eradication that arise from the existence of defective HIV viruses.

Whole-chromosome oligo-painting in licorice unveils interspecific chromosomal evolutionary relationships and possible origin of triploid genome species.

Licorice is one of the most extensively studied medicinal plants in the world, whose roots and rhizomes have long been used as both a sweetener and an essential component in numerous herbal preparations. However, the genus Glycyrrhiza has a complex composition, and the interspecies chromosomal relationships, origin, and evolution are still largely unclear. Here, we develop a set of whole-chromosome painting probes that allowed identification of all eight chromosomes of licorice on same metaphase chromosomes. Comparative chromosome painting analyses in seven different Glycyrrhiza species revealed that the genus Glycyrrhiza maintained extraordinarily conserved chromosomal synteny after about 3-12 million years of divergence. No cytologically visible inter-chromosomal rearrangements were identified in any species. By comparative chromosomal karyotype analyses, we revealed interspecific chromosome evolutionary relationships and dramatic variable chromosomal karyotype after independent divergence and demonstrated that G. prostrate was the most closely related to the ancestral type among the seven Glycyrrhiza species. Furthermore, we also discovered a G. glandulosa seed with distinct triploid-genome for the first time in China, suggesting the existence of a polyploid evolutionary pathway in the genus Glycyrrhiza, which challenges the previous notion that only diploids of licorice existed in nature. This study expands our knowledge of the chromosome evolution of licorice and will lay an important foundation for the genome origin and evolution studies in the genus Glycyrrhiza.

Phylogenomic Analysis and Functional Characterization of the APETALA2/Ethylene-Responsive Factor Transcription Factor Across Solanaceae.

The AP2/ERF family constitutes one of the largest groups of transcription factors in the Solanaceae. AP2/ERF contributes to various plant biological processes, including growth, development, and responses to various stresses. The origins and functional diversification of AP2/ERF within the Solanaceae family remain poorly understood, primarily because of the complex interactions between whole-genome duplications (WGDs) and tandem duplications. In this study, a total of 1282 AP2/ERF proteins are identified from 7 Solanaceae genomes. The amplification of AP2/ERF genes was driven not only by WGDs but also by the presence of clusters of tandem duplicated genes. The conservation of synteny across different chromosomes provides compelling evidence for the impact of the WGD event on the distribution pattern of AP2/ERF genes. Distinct expression patterns suggest that the multiple copies of AP2/ERF genes evolved in different functional directions, catalyzing the diversification of roles among the duplicated genes, which was of great significance for the adaptability of Solanaceae. Gene silencing and overexpression assays suggest that ERF-1 members' role in regulating the timing of floral initiation in C. annuum. Our findings provide insights into the genomic origins, duplication events, and function divergence of the Solanaceae AP2/ERF.

P1.03B.01 Multi-Omics Profiling Reveals the Genomic Origin and Adeno-Squamous Transdifferentiation Mechanism in Adenosquamous Carcinoma

P1.03B.01 Multi-Omics Profiling Reveals the Genomic Origin and Adeno-Squamous Transdifferentiation Mechanism in Adenosquamous Carcinoma

Novel centromeric repetitive DNA elements reveal karyotype dynamics in polyploid sainfoin (Onobrychis viciifolia).

Polyploidy is a common feature in eukaryotes with one of paramount consequences leading to better environmental adaptation. Heterochromatin is often located at telomeres and centromeres and contains repetitive DNA sequences. Sainfoin (Onobrychis viciifolia) is an important perennial forage legume for sustainable agriculture. However, there are only a few studies on the sainfoin genome and chromosomes. In this study, novel tandem repetitive DNA sequences of the sainfoin genome (OnVi180, OnVi169, OnVi176 and OnVidimer) were characterized using bioinformatics, molecular and cytogenetic approaches. The OnVi180 and OnVi169 elements colocalized within functional centromeres. The OnVi176 and OnVidimer elements were localized in centromeric, subtelomeric and interstitial regions. We constructed a sainfoin karyotype that distinguishes the seven basic chromosome groups. Our study provides the first detailed description of heterochromatin and chromosome structure of sainfoin and proposes an origin of heterozygous ancestral genomes, possibly from the same ancestral diploid species, not necessarily from different species, or for chromosome rearrangements after polyploidy. Overall, we discuss our novel and complementary findings in a polyploid crop with unique and complex chromosomal features.

Chromosome-specific painting reveals the Y genome origin and chromosome rearrangements of the St genome in Triticeae.

Karyotypes provide key cytogenetic information on phylogenetic relationships and evolutionary origins in related plant species. The St genome of Pseudoroegneria contributes to eight alloploid genera, representing over half of the species that are highly valuable for wheat (Triticum aestivum) breeding and for understanding Triticeae species evolution. However, St chromosome characterization is challenging due to limited cytogenetic markers and DNA information. We developed a complete set of St genome-specific chromosome painting probes for identification of the individual chromosomes 1St to 7St based on the genome sequences of Pse. libanotica and wheat. We revealed the conservation of St chromosomes in St-containing species by chromosome painting, including Pseudoroegneria, Roegneria, Elymus, and Campeiostachys. Notably, the Y genome showed hybridization signals, albeit weaker than those of the St genome. The awnless species harboring the Y genome exhibited more intense hybridization signals compare to the awned species in Roegneria and Campeiostachys, yet weaker than the hybridization signals of the St genome in autotetraploid Pse. strigosa. Although awnless species were morphologically more similar to each other, phenotypic divergence progressively increased from awnless to awned species. Our results indicate that the Y genome originated from the St genome and shed light on the possible origin of the Roegneria and Campeiostachys species, enhancing our understanding of St-genome-containing species evolution.

Multiregion exome sequencing indicates a monoclonal origin of esophageal spindle-cell squamous cell carcinoma.

Esophageal spindle-cell squamous cell carcinoma (ESS) is a rare biphasic neoplasm composed of a carcinomatous component (CaC) and a sarcomatous component (SaC). However, the genomic origin and gene signature of ESS remain unclear. Using whole-exome sequencing of laser-capture microdissection (LCM) tumor samples, we determined that CaC and SaC showed high mutational commonality, with the same top high-frequency mutant genes, mutation signatures, and tumor mutation burden; paired samples shared a median of 25.5% mutation sites. Focal gains were found on chromosomes 3q29, 5p15.33, and 11q13.3. Altered genes were mainly enriched in the RTK-RAS signaling pathway. Phylogenetic trees showed a monoclonal origin of ESS. The most frequently mutated oncogene in the trunk was TP53, followed by NFE2L2, KMT2D, and MUC16. Prognostic associations were found for CDC27, LRP2, APC, and SNAPC4. Our data highlight the monoclonal origin of ESS with TP53 as a potent driver oncogene, suggesting new targeted therapies and immunotherapies as treatment options. © 2024 The Pathological Society of Great Britain and Ireland.

The genome history of Bacillus thuringiensis subsp. tenebrionis: From discovery to present day product

AbstractBacillus thuringiensis subsp. tenebrionis (Btt) was discovered in 1982 and has been used as a biological control agent against the Colorado potato beetle (Leptinotarsa decemlineata) since the early 1990s. In this study, we sequenced, assembled and analysed the genome of strain DSM 2803, which is the patent deposit of the original Btt isolate BI 256‐82. A high‐quality reference genome is provided that allows investigating the genomic changes throughout the commercial history of Btt. Strain DSM 2803 (BI 256‐82) is particularly interesting since the subspecies description of ‘tenebrionis’ is based on it and because it represents the genomic origin of the commercial Btt strain currently used in plant protection. Comparative genome analysis of six Btt strains derived from BI 256‐82 revealed that the genomes were generally highly similar. However, some plasticity in the profile of extrachromosomal elements was detected, including recombination‐dependent plasmid differences and the presence/absence of a circular plasmid‐like prophage.

Multi-omics profiling the genomic origin and adeno-squamous transdifferentiation mechanism in adenosquamous carcinoma.

e20032 Background: Adenosquamous carcinoma (ASC) represents a rare subtype (0.4-4%) of non-small-cell lung cancer (NSCLC), with its multi-omic landscape yet to be fully elucidated. This study aims to explore the genomic origins and molecular mechanisms of adeno-squamous transdifferentiation in lung ASC through comprehensive multi-omics. Methods: We analyzed 69 samples from 31 lung ASC patients, including 16 FFPE samples with microdissected paired adenocarcinoma (AC) and squamous cell carcinoma (SCC) components, and additionally, incorporated pure Lung Adenocarcinoma (LUAD, n = 51) and pure Squamous Cell Carcinoma (LUSC, n = 28) for comparative analysis. Techniques included whole-exome sequencing, whole-genome methylation profiling, transcriptomics, single-cell RNA sequencing, and spatial transcriptomics. Results: AC and SCC exhibited similar genomic mutation landscapes, indicating a common genomic origin for AD and SCC. The SCC component in ASC displayed higher homologous recombination repair deficiency (HRD) and a notable 16p13.3 deletion in 31% of ASC transformations. Differential methylation analysis highlighted TP63's lower methylation in SCC, correlating negatively with RNA expression (P < 0.001), indicative of methylation-driven regulation. DMR binding motif analysis revealed hypo-methylation in p63, p53, and AP-1 motifs, and hyper-methylation in MKX2-1, FOXA1/2 motifs (P < 0.05). The transition from AC to SCC in ASC was marked by upregulated MYC signaling, proliferation, and DNA repair, with EMT changes not methylation-driven. Metabolic pathways were upregulated, and EMT/KRAS pathways downregulated in SCC compared to LUSC. Immune infiltration in LUAD-AC-ASC transformation showed an initial increase and subsequent decrease, transitioning from immune-infiltrated and immune-excluded to immune-desert phenotypes. IL36G of the IL1 family was highly expressed in the SCC component and strongly correlated with the squamous marker P63 (Pearson r = 0.72, P < 0.001). Single-cell RNA-seq data demonstrated a significant increase in IL36G in squamous cells of lung ASC. In vivo models showed adeno-squamous differentiation in adenocarcinoma cells expressing IL36G. Conclusions: This study provides a comprehensive multi-omic insight into the complex genomic and epigenomic landscape of ASC, highlighting a shared genomic origin and distinct molecular mechanisms in the adeno-squamous transdifferentiation process. The findings, particularly the role of TP63 methylation and IL36G expression in the SCC component, offer novel insights into the molecular heterogeneity of ASC. These results not only deepen our understanding of ASC but also pave the way for future investigations into targeted therapies that could potentially exploit these molecular distinctions for more effective treatment strategies in ASC.

Multi-region exome sequencing to determine the origin of esophageal spindle-cell squamous cell carcinoma.

4072 Background: Esophageal spindle-cell squamous cell carcinoma (ESS) is a rare biphasic neoplasm composed of carcinomatous and sarcomatous components. However, the genomic origin and gene signature of ESS remain unclear. Methods: A total of 24 patients diagnosed with ESS were enrolled from Shanghai Chest Hospital and underwent either radical resection (23/24) or endoscopic submucosal dissection (1/24). Laser-capture microdissection was employed to isolate cancerous and sarcomatous components from tumor and lymph node metastatic tissues. The cancerous components were subsequently microdissected based on morphology. Whole exome sequencing (WES) was conducted on both tumor tissues and paired white cell samples to analyze genomic profiles. Results: The tumor tissue samples of carcinomatous and sarcomatous components showed high mutational commonality. With the same top high-frequency mutant genes, mutation signatures, and tumor mutation burden, paired samples shared a median of 25.5% mutation sites. Focal gains were found on chromosomes 3q29, 5p15.33, and 11q13.3. Altered genes were mainly enriched in the RTK-RAS signaling pathway. Phylogenetic trees showed a monoclonal origin of ESS. The most frequently mutated oncogene in the trunk was TP53, followed by NFE2L2, KMT2D, and MUC16. It showed a significant association of CDC27, LRP2, APC, and SNAPC4 mutations with poor prognosis in ESS patients. Our data highlight the monoclonal origin of ESS with TP53 as a potent driver oncogene, suggesting new targeted therapies and immunotherapies as treatment options. Conclusions: In conclusion, our results regarding to the mutation characteristics of microdissected samples substantiate the monoclonal origin of both the carcinomatous and sarcomatous components of ESS.

Nucleosomal DNA has topological memory

One elusive aspect of the chromosome architecture is how it constrains the DNA topology. Nucleosomes stabilise negative DNA supercoils by restraining a DNA linking number difference (∆Lk) of about −1.26. However, whether this capacity is uniform across the genome is unknown. Here, we calculate the ∆Lk restrained by over 4000 nucleosomes in yeast cells. To achieve this, we insert each nucleosome in a circular minichromosome and perform Topo-seq, a high-throughput procedure to inspect the topology of circular DNA libraries in one gel electrophoresis. We show that nucleosomes inherently restrain distinct ∆Lk values depending on their genomic origin. Nucleosome DNA topologies differ at gene bodies (∆Lk = −1.29), intergenic regions (∆Lk = −1.23), rDNA genes (∆Lk = −1.24) and telomeric regions (∆Lk = −1.07). Nucleosomes near the transcription start and termination sites also exhibit singular DNA topologies. Our findings demonstrate that nucleosome DNA topology is imprinted by its native chromatin context and persists when the nucleosome is relocated.

Molecular Insights into Female Hybrid Sterility in Interspecific Crosses between Drosophila melanogaster and Drosophila simulans.

Species of the genus Drosophila have served as favorite models in speciation studies; however, genetic factors of interspecific reproductive incompatibility are under-investigated. Here, we performed an analysis of hybrid female sterility by crossing Drosophila melanogaster females and Drosophila simulans males. Using transcriptomic data analysis and molecular, cellular, and genetic approaches, we analyzed differential gene expression, transposable element (TE) activity, piRNA biogenesis, and functional defects of oogenesis in hybrids. Premature germline stem cell loss was the most prominent defect of oogenesis in hybrid ovaries. Because of the differential expression of genes encoding piRNA pathway components, rhino and deadlock, the functional RDCmel complex in hybrid ovaries was not assembled. However, the activity of the RDCsim complex was maintained in hybrids independent of the genomic origin of piRNA clusters. Despite the identification of a cohort of overexpressed TEs in hybrid ovaries, we found no evidence that their activity can be considered the main cause of hybrid sterility. We revealed a complicated pattern of Vasa protein expression in the hybrid germline, including partial AT-chX piRNA targeting of the vasasim allele and a significant zygotic delay in vasamel expression. We arrived at the conclusion that the hybrid sterility phenotype was caused by intricate multi-locus differences between the species.

Premeiotic 24-nt phasiRNAs are present in the Zea genus and unique in biogenesis mechanism and molecular function

Reproductive phasiRNAs (phased, small interfering RNAs) are broadly present in angiosperms and play crucial roles in sustaining male fertility. While the premeiotic 21-nt (nucleotides) phasiRNAs and meiotic 24-nt phasiRNA pathways have been extensively studied in maize (Zea mays) and rice (Oryza sativa), a third putative category of reproductive phasiRNAs-named premeiotic 24-nt phasiRNAs-have recently been reported in barley (Hordeum vulgare) and wheat (Triticum aestivum). To determine whether premeiotic 24-nt phasiRNAs are also present in maize and related species and begin to characterize their biogenesis and function, we performed a comparative transcriptome and degradome analysis of premeiotic and meiotic anthers from five maize inbred lines and three teosinte species/subspecies. Our data indicate that a substantial subset of the 24-nt phasiRNA loci in maize and teosinte are already highly expressed at the premeiotic phase. The premeiotic 24-nt phasiRNAs are similar to meiotic 24-nt phasiRNAs in genomic origin and dependence on DCL5 (Dicer-like 5) for biogenesis, however, premeiotic 24-nt phasiRNAs are unique in that they are likely i) not triggered by microRNAs, ii) not loaded by AGO18 proteins, and iii) not capable of mediating PHAS precursor cleavage. In addition, we also observed a group of premeiotic 24-nt phasiRNAs in rice using previously published data. Together, our results indicate that the premeiotic 24-nt phasiRNAs constitute a unique class of reproductive phasiRNAs and are present more broadly in the grass family (Poaceae) than previously known.

Genome constitution and evolution of Elymus atratus (Poaceae: Triticeae) inferred from cytogenetic and phylogenetic analysis.

Elymus atratus (Nevski) Hand.-Mazz. is perennial hexaploid wheatgrass. It was assigned to the genus Elymus L. sensu stricto based on morphological characters. Its genome constitution has not been disentangled yet. To identify the genome constitution and origin of E. atratus. In this study, genomic in situ hybridization and fluorescence in situ hybridization, and phylogenetic analysis based on the Acc1, DMC1 and matK sequences were performed. Genomic in situ hybridization and fluorescence in situ hybridization results reveal that E. atratus 2n = 6x = 42 is composed of 14 St genome chromosomes, 14 H genome chromosomes, and 14 Y genome chromosomes including two H-Y type translocation chromosomes, suggesting that the genome formula of E. atratus is StStYYHH. The phylogenetic analysis based on Acc1 and DMC1 sequences not only shows that the Y genome originated in a separate diploid, but also suggests that Pseudoroegneria (St), Hordeum (H), and a diploid species with Y genome were the potential donors of E. atratus. Data from chloroplast DNA showed that the maternal donor of E. atratus contains the St genome. Elymus atratus is an allohexaploid species with StYH genome, which may have originated through the hybridization between an allotetraploid Roegneria (StY) species as the maternal donor and a diploid Hordeum (H) species as the paternal donor.

Variations in the genomic profiles and clinical behavior of meningioma by racial and ethnic group.

The influence of socioeconomic factors on racial disparities among patients with sporadic meningiomas is well established, yet other potential causative factors warrant further exploration. The authors of this study aimed to determine whether there is significant variation in the genomic profile of meningiomas among patients of different races and ethnicities and its correlation with clinical outcomes. The demographic, genomic, and clinical data of patients aged 18 years and older who had undergone surgery for sporadic meningioma between September 2008 and November 2021 were analyzed. Statistical analyses were performed to detect differences across all racial/ethnic groups, as were direct comparisons between Black and non-Black groups plus Hispanic and non-Hispanic groups. This study included 460 patients with intracranial meningioma. Hispanic patients were significantly younger at surgery (53.9 vs 60.2 years, p = 0.0006) and more likely to show symptoms. Black patients had a higher incidence of anterior skull base tumors (OR 3.2, 95% CI 1.7-6.3, p = 0.0008) and somatic hedgehog mutations (OR 5.3, 95% CI 1.6-16.6, p = 0.003). Hispanics were less likely to exhibit the aggressive genomic characteristic of chromosome 1p deletion (OR 0.28, 95% CI 0.07-1.2, p = 0.06) and displayed higher rates of TRAF7 somatic driver mutations (OR 2.96 95% CI 1.1-7.8, p = 0.036). Black patients had higher rates of recurrence (OR 2.6, 95% CI 1.3-5.2, p = 0.009) and shorter progression-free survival (PFS; HR 2.9, 95% CI 1.6-5.4, p = 0.002) despite extents of resection (EORs) similar to those of non-Black patients (p = 0.745). No significant differences in overall survival were observed among groups. Despite similar EORs, Black patients had worse clinical outcomes following meningioma resection, characterized by a higher prevalence of somatic hedgehog mutations, increased recurrence rates, and shorter PFS. Meanwhile, Hispanic patients had less aggressive meningiomas, a predisposition for TRAF7 mutations, and no difference in PFS. These findings could inform the care and treatment strategies for meningiomas, and they establish the foundation for future studies focusing on the genomic origins of these observed differences.

How to report and make sense of a new HIV-1 circulating recombinant form?

Co-circulation of multiple HIV-1 subtypes in the same high-risk groups leads to the on-going generation of various inter-subtype recombinants, including unique (URFs) and circulating (CRFs) recombinant forms, which brings a new challenge for the prevention and eradication of HIV/AIDS. Identification and prompt reporting of new CRFs will provide not only new insights into the understanding of genetic diversity and evolution of HIV-1, but also an early warning of potential prevalence of these variants. Currently, 140 HIV-1 CRFs have been described; however, their prevalence and clinical importance are less concerned. Apart from the mosaic genomic maps, less other valuable information, including the clinical and demographic data, genomic sequence characteristics, origin and evolutionary dynamics, as well as representative genomic fragments for determining the variants, are available for most of these CRFs. Accompanied with the growing increase of HIV-1 full-length genomic sequences, more and more CRFs will be identified in the near future due to the high recombination potential of HIV-1. Here, we discuss the prevalence and clinical importance of various HIV-1 CRFs and propose how to report and make sense of a new HIV-1 CRF.

Unravelling the genomic origins of lumpy skin disease virus in recent outbreaks.

Lumpy skin disease virus (LSDV) belongs to the genus Capripoxvirus and family Poxviridae. LSDV was endemic in most of Africa, the Middle East and Turkey, but since 2015, several outbreaks have been reported in other countries. In this study, we used whole genome sequencing approach to investigate the origin of the outbreak and understand the genomic landscape of the virus. Our study showed that the LSDV strain of 2022 outbreak exhibited many genetic variations compared to the Reference Neethling strain sequence and the previous field strains. A total of 1819 variations were found in 22 genome sequences, which includes 399 extragenic mutations, 153 insertion frameshift mutations, 234 deletion frameshift mutations, 271 Single nucleotide polymorphisms (SNPs) and 762 silent SNPs. Thirty-eight genes have more than 2 variations per gene, and these genes belong to viral-core proteins, viral binding proteins, replication, and RNA polymerase proteins. We highlight the importance of several SNPs in various genes, which may play an essential role in the pathogenesis of LSDV. Phylogenetic analysis performed on all whole genome sequences of LSDV showed two types of variants in India. One group of the variant with fewer mutations was found to lie closer to the LSDV 2019 strain from Ranchi while the other group clustered with previous Russian outbreaks from 2015. Our study highlights the importance of genomic characterization of viral outbreaks to not only monitor the frequency of mutations but also address its role in pathogenesis of LSDV as the outbreak continues.

Whole-genome resequencing provides insights into the diversity and adaptation to desert environment in Xinjiang Mongolian cattle

BackgroundXinjiang Mongolian cattle is an indigenous breed that inhabits the Taklimakan Desert and is characterized by its small body size. However, the genomic diversity, origin, and genetic basis underlying the adaptation to the desert environment have been poorly studied.ResultsWe analyzed patterns of Xinjiang Mongolian cattle genetic variation by sequencing 20 genomes together with seven previously sequenced genomes and comparing them to the 134 genomes of nine representative breeds worldwide. Among the breeds of Bos taurus, we found the highest nucleotide diversity (0.0024) associated with the lower inbreeding coefficient (2.0110-6), the lowest linkage disequilibrium (r2 = 0.3889 at distance of 10 kb), and the highest effective population size (181 at 20 generations ago) in Xinjiang Mongolian cattle. The genomic diversity pattern could be explained by a limited introgression of Bos indicus genes. More importantly, similarly to desert-adapted camel and same-habitat sheep, we also identified signatures of selection including genes, GO terms, and/or KEGG pathways controlling water reabsorption and osmoregulation, metabolic regulation and energy balance, as well as small body size in Xinjiang Mongolian cattle.ConclusionsOur results imply that Xinjiang Mongolian cattle might have acquired distinct genomic diversity by virtue of the introgression of Bos indicus, which helps understand the demographic history. The identification of selection signatures can provide novel insights into the genomic basis underlying the adaptation of Xinjiang Mongolian cattle to the desert environment.

Genomic origin, fragmentomics, and transcriptional properties of long cell-free DNA molecules in human plasma.

Recent studies have revealed an unexplored population of long cell-free DNA (cfDNA) molecules in human plasma using long-read sequencing technologies. However, the biological properties of long cfDNA molecules (>500 bp) remain largely unknown. To this end, we have investigated the origins of long cfDNA molecules from different genomic elements. Analysis of plasma cfDNA using long-read sequencing reveals an uneven distribution of long molecules from across the genome. Long cfDNA molecules show overrepresentation in euchromatic regions of the genome, in sharp contrast to short DNA molecules. We observe a stronger relationship between the abundance of long molecules and mRNA gene expression levels, compared with short molecules (Pearson's r = 0.71 vs. -0.14). Moreover, long and short molecules show distinct fragmentation patterns surrounding CpG sites. Leveraging the cleavage preferences surrounding CpG sites, the combined cleavage ratios of long and short molecules can differentiate patients with hepatocellular carcinoma (HCC) from non-HCC subjects (AUC = 0.87). We also investigated knockout mice in which selected nuclease genes had been inactivated in comparison with wild-type mice. The proportion of long molecules originating from transcription start sites are lower in Dffb-deficient mice but higher in Dnase1l3-deficient mice compared with that of wild-type mice. This work thus provides new insights into the biological properties and potential clinical applications of long cfDNA molecules.

Analysis of ddRAD-seq data provides new insights into the genomic structure and patterns of diversity in Italian donkey populations.

With more than 150 recognized breeds, donkeys assume relevant economic importance, especially in developing countries. Even if the estimated number of heads worldwide is 53M, this species received less attention than other livestock species. Italy has traditionally been considered one of the cradles of European donkey breeding, and despite a considerable loss of biodiversity, today still counts nine autochthonous populations. A total of 220 animals belonging to 9 different populations were genotyped using the double-digest restriction site associated DNA (ddRAD) sequencing to investigate the pattern of diversity using a multi-technique approach. A total of 418,602,730 reads were generated and successfully demultiplexed to obtain a medium-density SNP genotypes panel with about 27K markers. The diversity indices showed moderate levels of variability. The genetic distances and relationships, largely agree with the breeding history of the donkey populations under investigation. The results highlighted the separation of populations based on their genetic origin or geographical proximity between breeding areas, showed low to moderate levels of admixture, and indicated a clear genetic difference in some cases. For some breeds, the results also validate the success of proper management conservation plans. Identified ROH islands, mapped within genomic regions related to immune response and local adaptation, are consistent with the characteristics of the species known for its rusticity and adaptability. This study is the first exhaustive genome-wide analysis of the diversity of Italian donkey populations. The results emphasized the high informativeness of genome-wide markers retrieved through the ddRAD approach. The findings take on great significance in designing and implementing conservation strategies. Standardized genotype arrays for donkey species would make it possible to combine worldwide datasets to provide further insights on the evolution of the genomic structure and origin of this important genetic resource.