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Sheng Wu Yi Xue Gong Cheng Xue Za Zhi =... Jun 2024The rapid development of high-throughput chromatin conformation capture (Hi-C) technology provides rich genomic interaction data between chromosomal loci for chromatin...
The rapid development of high-throughput chromatin conformation capture (Hi-C) technology provides rich genomic interaction data between chromosomal loci for chromatin structure analysis. However, existing methods for identifying topologically associated domains (TADs) based on Hi-C data suffer from low accuracy and sensitivity to parameters. In this context, a TAD identification method based on spatial density clustering was designed and implemented in this paper. The method preprocessed the raw Hi-C data to obtain normalized Hi-C contact matrix data. Then, it computed the distance matrix between loci, generated a reachability graph based on the core distance and reachability distance of loci, and extracted clustering clusters. Finally, it extracted TAD boundaries based on clustering results. This method could identify TAD structures with higher coherence, and TAD boundaries were enriched with more ChIP-seq factors. Experimental results demonstrate that our method has advantages such as higher accuracy and practical significance in TAD identification.
Topics: Chromatin; Cluster Analysis; Algorithms; Humans; Chromatin Immunoprecipitation Sequencing
PubMed: 38932542
DOI: 10.7507/1001-5515.202311059 -
Journal of Medical Virology Jun 2024Integration of the human papillomavirus (HPV) genome into the cellular genome is a key event that leads to constitutive expression of viral oncoprotein E6/E7 and drives...
Integration of the human papillomavirus (HPV) genome into the cellular genome is a key event that leads to constitutive expression of viral oncoprotein E6/E7 and drives the progression of cervical cancer. However, HPV integration patterns differ on a case-by-case basis among related malignancies. Next-generation sequencing technologies still face challenges for interrogating HPV integration sites. In this study, utilizing Nanopore long-read sequencing, we identified 452 and 108 potential integration sites from the cervical cancer cell lines (CaSki and HeLa) and five tissue samples, respectively. Based on long Nanopore chimeric reads, we were able to analyze the methylation status of the HPV long control region (LCR), which controls oncogene E6/E7 expression, and to identify transcriptionally-active integrants among the numerous integrants. As a proof of concept, we identified an active HPV integrant in between RUNX2 and CLIC5 on chromosome 6 in the CaSki cell line, which was supported by ATAC-seq, H3K27Ac ChIP-seq, and RNA-seq analysis. Knockout of the active HPV integrant, by the CRISPR/Cas9 system, dramatically crippled cell proliferation and induced cell senescence. In conclusion, identifying transcriptionally-active HPV integrants with Nanopore sequencing can provide viable targets for gene therapy against HPV-associated cancers.
Topics: Humans; Uterine Cervical Neoplasms; Female; Nanopore Sequencing; Virus Integration; Genetic Therapy; Papillomavirus Infections; Cell Line, Tumor; HeLa Cells; Oncogene Proteins, Viral; High-Throughput Nucleotide Sequencing; Papillomaviridae; Human Papillomavirus Viruses
PubMed: 38932482
DOI: 10.1002/jmv.29769 -
Viruses Jun 2024Human rotaviruses exhibit limited tropism and replicate poorly in most cell lines. Attachment protein VP4 is a key rotavirus tropism determinant. Previous studies in...
Human rotaviruses exhibit limited tropism and replicate poorly in most cell lines. Attachment protein VP4 is a key rotavirus tropism determinant. Previous studies in which human rotaviruses were adapted to cultured cells identified mutations in VP4. However, most such studies were conducted using only a single human rotavirus genotype. In the current study, we serially passaged 50 human rotavirus clinical specimens representing five of the genotypes most frequently associated with severe human disease, each in triplicate, three to five times in primary monkey kidney cells then ten times in the MA104 monkey kidney cell line. From 13 of the 50 specimens, we obtained 25 rotavirus antigen-positive lineages representing all five genotypes, which tended to replicate more efficiently in MA104 cells at late versus early passage. We used Illumina next-generation sequencing and analysis to identify variants that arose during passage. In VP4, variants encoded 28 mutations that were conserved for all P[8] rotaviruses and 12 mutations that were conserved for all five genotypes. These findings suggest there may be a conserved mechanism of human rotavirus adaptation to MA104 cells. In the future, such a conserved adaptation mechanism could be exploited to study human rotavirus biology or efficiently manufacture vaccines.
Topics: Rotavirus; Humans; Capsid Proteins; Genotype; Mutation; Animals; Rotavirus Infections; Serial Passage; Cell Line; Virus Replication; High-Throughput Nucleotide Sequencing; Viral Tropism
PubMed: 38932271
DOI: 10.3390/v16060978 -
Viruses Jun 2024Viral integration within the host genome plays a pivotal role in carcinogenesis. Various disruptive mechanisms are involved, leading to genomic instability, mutations,...
Viral integration within the host genome plays a pivotal role in carcinogenesis. Various disruptive mechanisms are involved, leading to genomic instability, mutations, and DNA damage. With next-generation sequencing (NGS), we can now precisely identify viral and host genomic breakpoints and chimeric sequences, which are useful for integration site analysis. In this study, we evaluated a commercial hybrid capture NGS panel specifically designed for detecting three key viruses: HPV, HBV, and HIV-1. We also tested workflows for Viral Hybrid Capture (VHC) and Viral Integration Site (VIS) analysis, leveraging customized viral databases in CLC Microbial Genomics. By analyzing sequenced data from virally infected cancer cell lines (including SiHa, HeLa, CaSki, C-33A, DoTc2, 2A3, SCC154 for HPV; 3B2, SNU-182 for HBV; and ACH-2 for HIV-1), we precisely pinpointed viral integration sites. The workflow also highlighted disrupted and neighboring human genes that may play a crucial role in tumor development. Our results included informative virus-host read mappings, genomic breakpoints, and integration circular plots. These visual representations enhance our understanding of the integration process. In conclusion, our seamless end-to-end workflow bridges the gap in understanding viral contributions to cancer development, paving the way for improved diagnostics and treatment strategies.
Topics: Humans; Virus Integration; Hepatitis B virus; HIV-1; High-Throughput Nucleotide Sequencing; Workflow; Carcinogenesis; Genomics; Cell Line, Tumor; Papillomaviridae
PubMed: 38932267
DOI: 10.3390/v16060975 -
Viruses Jun 2024Hepatitis A virus (HAV), a member of the genus ( HepV), remains a significant viral pathogen, frequently causing enterically transmitted hepatitis worldwide. In this...
Hepatitis A virus (HAV), a member of the genus ( HepV), remains a significant viral pathogen, frequently causing enterically transmitted hepatitis worldwide. In this study, we conducted an epidemiological survey of HepVs carried by small terrestrial mammals in the wild in Yunnan Province, China. Utilizing HepV-specific broad-spectrum RT-PCR, next-generation sequencing (NGS), and QNome nanopore sequencing (QNS) techniques, we identified and characterized two novel HepVs provisionally named EpMa-HAV and EpLe-HAV, discovered in the long-tailed mountain shrew () and long-tailed brown-toothed shrew (), respectively. Our sequence and phylogenetic analyses of EpMa-HAV and EpLe-HAV indicated that they belong to the species (HepV-I) clade II, also known as the Chinese shrew HepV clade. Notably, the codon usage bias pattern of novel shrew HepVs is consistent with that of previously identified Chinese shrew HepV. Furthermore, our structural analysis demonstrated that shrew HepVs differ from other mammalian HepVs in RNA secondary structure and exhibit variances in key protein sites. Overall, the discovery of two novel HepVs in shrews expands the host range of HepV and underscores the existence of genetically diverse animal homologs of human HAV within the genus HepV.
Topics: Animals; Shrews; China; Phylogeny; Genome, Viral; RNA, Viral; Genomics; High-Throughput Nucleotide Sequencing; Picornaviridae Infections
PubMed: 38932262
DOI: 10.3390/v16060969 -
Viruses Jun 2024The diversity of and species in tomatoes was assessed via high-throughput sequencing of 154 symptomatic foliar samples collected from 2002 to 2017 across seven...
The diversity of and species in tomatoes was assessed via high-throughput sequencing of 154 symptomatic foliar samples collected from 2002 to 2017 across seven Brazilian biomes. The first pool (BP1) comprised 73 samples from the North (13), Northeast (36), and South (24) regions. Sixteen begomoviruses and one were detected in BP1. Four begomovirus-like contigs were identified as putative novel species (NS). NS#1 was reported in the semi-arid (Northeast) region and NS#2 and NS#4 in mild subtropical climates (South region), whereas NS#3 was detected in the warm and humid (North) region. The second pool (BP2) comprised 81 samples from Southeast (39) and Central-West (42) regions. Fourteen viruses and subviral agents were detected in BP2, including two topileviruses, a putative novel begomovirus (NS#5), and two alphasatellites occurring in continental highland areas. The five putative novel begomoviruses displayed strict endemic distributions. Conversely, tomato mottle leaf curl virus (a monopartite species) displayed the most widespread distribution occurring across the seven sampled biomes. The overall diversity and frequency of mixed infections were higher in susceptible (16 viruses + alphasatellites) in comparison to tolerant (carrying the -1 or -3 introgressions) samples, which displayed 9 viruses. This complex panorama reinforces the notion that the tomato-associated diversity is yet underestimated in Neotropical regions.
Topics: Solanum lycopersicum; Brazil; Plant Diseases; Metagenomics; Phylogeny; Geminiviridae; Animals; Genetic Variation; Genome, Viral; Begomovirus; High-Throughput Nucleotide Sequencing
PubMed: 38932191
DOI: 10.3390/v16060899 -
Viruses May 2024Viral diseases pose a significant threat to tomato crops ( L.), one of the world's most economically important vegetable crops. The limited genetic diversity of...
Viral diseases pose a significant threat to tomato crops ( L.), one of the world's most economically important vegetable crops. The limited genetic diversity of cultivated tomatoes contributes to their high susceptibility to viral infections. To address this challenge, tomato breeding programs must harness the genetic resources found in native populations and wild relatives. Breeding efforts may aim to develop broad-spectrum resistance against the virome. To identify the viruses naturally infecting 19 advanced lines, derived from native tomatoes, high-throughput sequencing (HTS) of small RNAs and confirmation with PCR and RT-PCR were used. Single and mixed infections with tomato mosaic virus (ToMV), tomato golden mosaic virus (ToGMoV), and pepper huasteco yellow vein virus (PHYVV) were detected. The complete consensus genomes of three variants of Mexican ToMV isolates were reconstructed, potentially forming a new ToMV clade with a distinct 3' UTR. The absence of reported mutations associated with resistance-breaking to ToMV suggests that the , , and genes could theoretically be used to confer resistance. However, the high mutation rates and a 63 nucleotide insertion in the 3' UTR, as well as amino acid mutations in the ORFs encoding 126 KDa, 183 KDa, and MP of Mexican ToMV isolates, suggest that it is necessary to evaluate the capacity of these variants to overcome , , and resistance genes. This evaluation, along with the characterization of advanced lines using molecular markers linked to these resistant genes, will be addressed in future studies as part of the breeding strategy. This study emphasizes the importance of using HTS for accurate identification and characterization of plant viruses that naturally infect tomato germplasm based on the consensus genome sequences. This study provides crucial insights to select appropriate disease management strategies and resistance genes and guide breeding efforts toward the development of virus-resistant tomato varieties.
Topics: Plant Diseases; Solanum lycopersicum; High-Throughput Nucleotide Sequencing; Plant Breeding; Plant Viruses; Genome, Viral; Phylogeny; Disease Resistance; RNA, Viral
PubMed: 38932180
DOI: 10.3390/v16060888 -
Viruses May 2024Recently, high-throughput sequencing of influenza A viruses has become a routine test. It should be noted that the extremely high diversity of the influenza A virus...
Recently, high-throughput sequencing of influenza A viruses has become a routine test. It should be noted that the extremely high diversity of the influenza A virus complicates the task of determining the sequences of all eight genome segments. For a fast and accurate analysis, it is necessary to select the most suitable reference for each segment. At the same time, there is no standardized method in the field of decoding sequencing results that allows the user to update the sequence databases to which the reads obtained by virus sequencing are compared. The IAVCP (influenza A virus consensus and phylogeny) was developed with the goal of automatically analyzing high-throughput sequencing data of influenza A viruses. Its goals include the extraction of a consensus genome directly from paired raw reads. In addition, the pipeline enables the identification of potential reassortment events in the evolutionary history of the virus of interest by analyzing the topological structure of phylogenetic trees that are automatically reconstructed.
Topics: High-Throughput Nucleotide Sequencing; Phylogeny; Genome, Viral; Influenza A virus; Humans; Genomics; Influenza, Human; Computational Biology
PubMed: 38932165
DOI: 10.3390/v16060873 -
Viruses May 2024Recombination events in human adenovirus (HAdV) have led to some new highly pathogenic or infectious types. It is vital to monitor recombinant HAdVs, especially in...
Recombination events in human adenovirus (HAdV) have led to some new highly pathogenic or infectious types. It is vital to monitor recombinant HAdVs, especially in children with acute respiratory tract infections (ARIs). In the retrospective study, HAdV positive specimens were collected from pediatric patients with ARIs during 2015 to 2021, then typed by sequence analysis of the penton base, hexon and fiber gene sequence. For those with inconsistent typing results, a modified method with species-specific primer sets of a fiber gene sequence was developed to distinguish co-infections of different types from recombinant HAdV infections. Then, plaque assays combined with meta-genomic next-generation sequencing (mNGS) were used to reveal the HAdV genomic characteristics. There were 466 cases positive for HAdV DNA (2.89%, 466/16,097) and 350 (75.11%, 350/466) successfully typed with the most prevalent types HAdV-B3 (56.57%, 198/350) and HAdV-B7 (32.00%, 112/350), followed by HAdV-C1 (6.00%, 21/350). Among 35 cases (7.51%, 35/466) with inconsistent typing results, nine cases were confirmed as co-infections by different types of HAdVs, and 26 cases as recombinant HAdVs in six genetic patterns primarily clustered to species C (25 cases) in pattern 1-5, or species D (1 case) in pattern 6. The novel recombinant HAdV of species D was identified with multiple recombinant events among HAdV-D53, HAdV-D64, and HAdV-D8, and officially named as HAdV-D115. High-frequency recombination of HAdVs in six genetic recombination patterns were identified among children with ARIs in Beijing. Specifically, there is a novel Adenovirus D human/CHN/S8130/2023/115[P22H8F8] designed as HAdV D115.
Topics: Humans; Adenoviruses, Human; Respiratory Tract Infections; Recombination, Genetic; Adenovirus Infections, Human; Child, Preschool; Retrospective Studies; Male; Child; Infant; Female; Phylogeny; Beijing; Genotype; High-Throughput Nucleotide Sequencing; Coinfection; DNA, Viral; Genome, Viral; Adolescent; China
PubMed: 38932121
DOI: 10.3390/v16060828 -
Molecules (Basel, Switzerland) Jun 2024Due to the intricate complexity of the original microbiota, residual heat-resistant enzymes, and chemical components, identifying the essential factors that affect dairy...
Due to the intricate complexity of the original microbiota, residual heat-resistant enzymes, and chemical components, identifying the essential factors that affect dairy quality using traditional methods is challenging. In this study, raw milk, pasteurized milk, and ultra-heat-treated (UHT) milk samples were collectively analyzed using metagenomic next-generation sequencing (mNGS), high-throughput liquid chromatography-mass spectrometry (LC-MS), and gas chromatography-mass spectrometry (GC-MS). The results revealed that raw milk and its corresponding heated dairy products exhibited different trends in terms of microbiota shifts and metabolite changes during storage. Via the analysis of differences in microbiota and correlation analysis of the microorganisms present in differential metabolites in refrigerated pasteurized milk, the top three differential microorganisms with increased abundance, ( 0.01), unclassified class ( 0.05), and ( 0.01), were detected; these were highly correlated with certain metabolites in pasteurized milk ( > 0.8). This indicated that these genera were the main proliferating microorganisms and were the primary genera involved in the metabolism of pasteurized milk during refrigeration-based storage. Microorganisms with decreased abundance were classified into two categories based on correlation analysis with certain metabolites. It was speculated that the heat-resistant enzyme system of a group of microorganisms with high correlation ( > 0.8), such as and , was the main factor causing milk spoilage and that the group with lower correlation ( < 0.3) had a lower impact on the storage process of pasteurized dairy products. By comparing the metabolic pathway results based on metagenomic and metabolite annotation, it was proposed that protein degradation may be associated with microbial growth, whereas lipid degradation may be linked to raw milk's initial heat-resistant enzymes. By leveraging the synergy of metagenomics and metabolomics, the interacting factors determining the quality evolution of dairy products were systematically investigated, providing a novel perspective for controlling dairy processing and storage effectively.
Topics: Microbiota; Animals; Milk; Food Storage; Pasteurization; High-Throughput Nucleotide Sequencing; Dairy Products; Metagenomics; Gas Chromatography-Mass Spectrometry; Food Handling; Bacteria; Metabolome
PubMed: 38930811
DOI: 10.3390/molecules29122745