Viral DNA Research Articles

TRIM35 Negatively Regulates the cGAS-STING-Mediated Signaling Pathway by Attenuating K63-Linked Ubiquitination of STING.

The cGAS-STING-mediated antiviral response plays an important role in the defense against DNA virus infection. Tripartite motif protein 35 (TRIM35), an E3 ubiquitin ligase, was identified as a positive regulator of RLR-mediated antiviral signaling in our previous study, but the effect of TRIM35 on the cGAS-STING signaling pathway has not been elucidated. Herein, we showed that TRIM35 negatively regulates the cGAS-STING signaling pathway by directly targeting STING. TRIM35 overexpression significantly inhibited the cGAMP-triggered phosphorylation of TBK1 and IRF3, attenuating IFN-β expression and the downstream antiviral response. Mechanistically, TRIM35 colocalized and directly interacted with STING in the cytoplasm. TRM35 removed K63-linked ubiquitin from STING through the C36 and C44 sites in the RING domain, which impaired the interaction of STING with TBK1 or IKKε. In addition, we demonstrated that the RING domain is a key region for the antiviral effects of TIRM35. These results collectively indicate that TRIM35 negatively regulates type I interferon (IFN-I) production by targeting and deubiquitinating STING. TRIM35 may be a potential therapeutic target for controlling viral infection.

UBXN3B is crucial for B lymphopoiesis

The ubiquitin regulatory X (UBX) domain-containing proteins (UBXNs) are putative adaptors for ubiquitin ligases and valosin-containing protein; however, their invivo physiological functions remain poorly characterised. We recently showed that UBXN3B is essential for activating innate immunity to DNA viruses and controlling DNA/RNA virus infection. Herein, we investigate its role in adaptive immunity. We evaluated the antibody responses to multiple viruses and pathogenesis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza in tamoxifen-inducible global and constitutive B cell-specific Ubxn3b knockout mice; quantified various immune populations, B lineage progenitors/precursors, B cell receptor (BCR) signalling and apoptosis by flow cytometry, immunoblotting and immunofluorescence microscopy. We also performed bone marrow transfer, single-cell and bulk RNA sequencing. Both global and B cell-specific Ubxn3b knockout mice present a marked reduction in small precursor B-II (>60%), immature (>70%) and mature B (>95%) cell numbers. Transfer of wildtype bone marrow to irradiated global Ubxn3b knockouts restores normal B lymphopoiesis, while reverse transplantation does not. The mature B population shrinks rapidly with apoptosis and higher pro and activated caspase-3 protein levels were observed following induction of Ubxn3b knockout. Mechanistically, Ubxn3b deficiency leads to impaired pre-BCR signalling and cell cycle arrest. Ubxn3b knockout mice are highly vulnerable to respiratory viruses, with increased viral loads and prolonged immunopathology in the lung, and reduced production of virus-specific IgM/IgG. UBXN3B is essential for B lymphopoiesis by maintaining constitutive pre-BCR signalling and cell survival in a cell-intrinsic manner. United States National Institutes of Health grants, R01AI132526 and R21AI155820.

DNA damage response and inflammatory response: Two traffic lights for HPVs on the road to transformation.

Human papillomaviruses (HPVs) are non-enveloped double-stranded DNA viruses. When HPV infection persists, infected tissues can develop many HPV-related diseases such as cervical cancer and head and neck squamous cell carcinoma. To establish their persistent infection, HPVs have evolved mechanisms to manipulate the host cellular processes such as DNA damage response (DDR), which includes homologous recombination, nonhomologous end joining, and microhomology-mediated end joining. Additionally, HPVs utilize host inflammatory processes to facilitate their life cycles. Here, we bridge the concepts of DDR and inflammatory response, and discuss how HPV proteins orchestrate a sophisticated manipulation of DDR and inflammation to promote their viral replication, ultimately fostering the progression of infected cells towards oncogenic transformation to malignancy.

Evaluation of the diagnostic sensitivity and specificity of two pen-side tests for detecting African swine fever virus in experimentally infected pigs

African swine fever virus (ASFV) has spread through many countries and regions worldwide, causing significant losses. Timely detection of ASFV-infected pigs is crucial for disease control. In this study, we assessed the performance of two pen-side tests: a portable real-time PCR (qPCR) test for detecting viral genomic DNA and a lateral flow immunoassay (LFIA) for detecting viral antigens. To determine the time from infection to the earliest detection, 10 ASFV-seronegative pigs were inoculated intramuscularly with 104.0 hemadsorption dose 50 of a highly virulent ASFV strain. Whole blood and oral swab samples were alternately collected from each group of five pigs daily until all succumbed to the infection. Samples were promptly subjected to the two pen-side tests upon collection, and a subset was transported to a veterinary diagnostic laboratory for analysis using a reference qPCR assay. Viral genomic DNA was consistently detected by the reference qPCR assay in all blood samples from 2 days postinfection (dpi), preceding the onset of clinical signs, and in oral swabs from 4 dpi onwards. The portable qPCR test demonstrated comparable performance to the reference qPCR assay for both whole blood and oral swab samples. The LFIA exhibited 100% specificity when testing with whole blood samples but showed reduced sensitivity, particularly for blood samples collected early or late after infection. The antigen test did not perform well with oral swabs.

Dried Blood Matrix as a New Material for the Detection of DNA Viruses.

The gold standard for diagnosing viruses such as the Hepatitis B Virus has remained largely unchanged, relying on conventional methods involving extraction, purification, and polymerase chain reaction (PCR). This approach is hindered by limited availability, as it is time-consuming and requires highly trained personnel. Moreover, it suffers from low recovery rates of the nucleic acid molecules for samples with low copy numbers. To address the challenges of complex instrumentation and low recovery rate of DNA, a drying process coupled with thermal treatment of whole blood is employed, resulting in the creation of a dried blood matrix characterized by a porous structure with a high surface-to-volume ratio where it also inactivates the amplification inhibitors present in whole blood. Drawing on insights from Brunauer-Emmett-Teller (BET)- Barrett-Joyner-Halenda (BJH) analysis, scanning electron microscopy (SEM), and fluorescence recovery after photobleaching (FRAP), detection assay is devised for HBV, as a demonstration, from whole blood with high recovery of DNA and simplified instrumentation achieving a limit of detection (LOD) of 10 IU mL-1. This assay can be completed in <1.5 h using a simple heater, can be applied to other DNA viruses, and is expected to be suitable for point-of-care, especially in low-resource settings.

Association of preoperative antiviral treatment with incidences of post-hepatectomy liver failure in hepatitis B virus-related hepatocellular carcinoma.

Post-hepatectomy liver failure (PHLF) is a common consequence of radical partial hepatectomy in hepatocellular carcinoma (HCC). To investigate the relationship between preoperative antiviral therapy and PHLF, as well as assess the potential efficacy of hepatitis B virus (HBV) DNA level in predicting PHLF. A retrospective study was performed involving 1301 HCC patients with HBV who underwent radical hepatectomy. Receiver operating characteristic (ROC) analysis was used to assess the capacity of HBV DNA to predict PHLF and establish the optimal cutoff value for subsequent analyses. Logistic regression analyses were performed to assess the independent risk factors of PHLF. The increase in the area under the ROC curve, categorical net reclassification improvement (NRI), and integrated discrimination improvement (IDI) were used to quantify the efficacy of HBV DNA level for predicting PHLF. The P < 0.05 was considered statistically significant. Logistic regression analyses showed that preoperative antiviral therapy was independently associated with a reduced risk of PHLF (P < 0.05). HBV DNA level with an optimal cutoff value of 269 IU/mL (P < 0.001) was an independent risk factor of PHLF. All the reference models by adding the variable of HBV DNA level had an improvement in area under the curve, categorical NRI, and IDI, particularly for the fibrosis-4 model, with values of 0.729 (95%CI: 0.705-0.754), 1.382 (95%CI: 1.341-1.423), and 0.112 (95%CI: 0.110-0.114), respectively. All the above findings were statistically significant. In summary, preoperative antiviral treatment can reduce the incidence of PHLF, whereas an increased preoperative HBV DNA level has a correlative relationship with an increased susceptibility to PHLF.

First report of fesavirus 4 detection from cats in Japan.

Fesaviruses, picorna-like RNA viruses, were discovered in 2014 in feces from cats in an animal shelter in the United States but have not since been reported elsewhere. In this study, we collected cat fecal samples from 20 adult cats from an animal shelter in Tokyo, Japan, and examined them for viral pathogens. Next generation sequencing (NGS) was performed to detect both RNA and DNA virus sequences. Sequences of a total of 7 RNA viruses including some common feline pathogenic viruses were detected across 8 samples, while no DNA virus sequences were identified in any sample. Of the RNA virus sequences detected in the samples, two sequences, 4,746 and 4,439 bp, demonstrated 90.3% and 85.0% similarity, respectively, to the fesavirus 4 sequence in the database. To confirm the NGS results, quantitative RT-PCR (qRT-PCR) assays were developed using specific primers and probes designed based on the contig sequences. Based on the qRT-PCR assays, we detected relatively high copy-numbers of fesavirus 4 RNA in the two fecal samples from which the fesavirus 4 sequences were originally obtained, and low copy numbers in other samples. These results demonstrate the presence of fesavirus 4 in cats in Japan for the first time.

Development of Fournier's gangrene after chemotherapy for the recurrence of testicular cancer despite the absence of anorectal lesions: A case report.

Fournier's gangrene usually occurs when a specific bacterium intrudes into soft tissue, causing a wound or tumor. We encountered a patient with Fournier's gangrene due to severe myelosuppression after chemotherapy, despite the absence of an initial lesion on the anus and rectum. A 54-year-old man with a left testicular cancer recurrence had undergone chemotherapy. He had asymptomatic hepatitis and high hepatitis B virus DNA levels, which were normalized by administering tenofovir alafenamide fumarate. Twelve days after the start of chemotherapy, he complained of severe pain around the anus. The following day, he went into septic shock. Visual inspection showed dark purple skin discoloration on the left side of the anus. Laboratory data revealed severe neutropenia. Computed tomography showed a high density of soft tissue on the left side of the anus and gas bubbles in the left femoral ring. We diagnosed the patient with Fournier's gangrene due to a severe immunosuppressive state resulting from chemotherapy. We emergently removed necrotic tissue to the fullest extent possible. However, because the patient was in severe sepsis status, careful management in the intensive care unit was required for 32 days. After the first emergency operation, we performed several additional excisions. Finally, 391 days after the initial surgery, the patient was discharged from our hospital. The tumor has not recurred, and he is under outpatient observation in the urology department. Fournier's gangrene should be considered in patients who are in a severe myelosuppressive state due to chemotherapy, have normal hepatitis B virus DNA levels but high hepatitis B surface antigen after tenofovir administration, complain of severe pain in the perianal area, and have a dark purple skin discoloration around the anus, despite having no initial anorectal lesions.

Imaging and quantification of human and viral circular RNAs.

We present a robust approach for cellular detection, imaging, localization, and quantification of human and viral encoded circular RNAs (circRNA) using amplified fluorescence in situ hybridization (ampFISH). In this procedure, a pair of hairpin probes bind next to each other at contiguous stretches of sequence and then undergo a conformational reorganization which initiates a target-dependent hybridization chain reaction (HCR) resulting in deposition of an amplified fluorescent signal at the site. By harnessing the capabilities of both ampFISH and single-molecule FISH (smFISH), we selectively identified and imaged circular RNAs and their linear counterparts derived from the human genome, SARS-CoV-2 (an RNA virus), and human cytomegalovirus (HCMV, a DNA virus). Computational image processing facilitated accurate quantification of circular RNA molecules in individual cells. The specificity of ampFISH for circular RNA detection was confirmed through an in situ RNase R treatment that selectively degrades linear RNAs without impacting circular RNAs. The effectiveness of circular RNA detection was further validated by using ampFISH probes with mismatches and probe pairs that do not bind to the continuous sequence in their target RNAs but instead bind at segregated sites. An additional specificity test involved probes against the negative strands of the circular RNA sequence, absent in the cell. Importantly, our technique allows simultaneous detection of circular RNAs and their linear counterparts within the same cell with single molecule sensitivity, enabling explorations of circular RNA biogenesis, subcellular localization, and functions.

CSF1R inhibition depletes brain macrophages and reduces brain virus burden in SIV-infected macaques.

Perivascular macrophages (PVMs) and, to a lesser degree, microglia are targets and reservoirs of HIV and simian immunodeficiency virus (SIV) in the brain. Previously, we demonstrated that colony-stimulating factor 1 receptor (CSF1R) in PVMs was upregulated and activated in chronically SIV-infected rhesus macaques with encephalitis, correlating with SIV infection of PVMs. Herein, we investigated the role of CSF1R in the brain during acute SIV infection using BLZ945, a brain-penetrant CSF1R kinase inhibitor. Apart from three uninfected historic controls, nine Indian rhesus macaques were infected acutely with SIVmac251 and divided into three groups (n = 3 each): an untreated control and two groups treated for 20-30 days with low- (10 mg/kg/day) or high- (30 mg/kg/day) dose BLZ945. With the high-dose BLZ945 treatment, there was a significant reduction in cells expressing CD163 and CD206 across all four brain areas examined, compared with the low-dose treatment and control groups. In 9 of 11 tested regions, tissue viral DNA (vDNA) loads were reduced by 95%-99% following at least one of the two doses, and even to undetectable levels in some instances. Decreased numbers of CD163+ and CD206+ cells correlated significantly with lower levels of vDNA in all four corresponding brain areas. In contrast, BLZ945 treatment did not significantly affect the number of microglia. Our results indicate that doses as low as 10 mg/kg/day of BLZ945 are sufficient to reduce the tissue vDNA loads in the brain with no apparent adverse effect. This study provides evidence that infected PVMs are highly sensitive to CSF1R inhibition, opening new possibilities to achieve viral clearance.

Improving grapevine virus diagnostics: Comparative analysis of three dsRNA enrichment methods for high-throughput sequencing

The extraction of double stranded (ds) RNA is a common enrichment method for the study, characterization, and detection of RNA viruses. In addition to RNA viruses, viroids, and some DNA viruses, can also be detected from dsRNA enriched extracts which makes it an attractive method for detecting a wide range of viruses when coupled with HTS. Several dsRNA enrichment strategies have been developed. The oldest utilizes the selective binding properties of dsRNA to cellulose. More recent methods are based on the application of anti-dsRNA antibodies and viral proteins with a specific affinity for dsRNA. All three methods have been used together with HTS for plant virus detection and study. To our knowledge, this is the first comparative study of three alternative dsRNA enrichment methods for virus and viroid detection through HTS using virus-infected, and healthy grapevine test plants. Extracts were performed in triplicate using methods based on, the anti-dsRNA antibody mAb rJ2 (Millipore Sigma Canada Ltd, Oakville, ON, Canada), the B2 dsRNA binding protein, and ReliaPrep™ Resin (Promega Corporation, Madison, WI, USA). The results show that the workflows for all three methods are effectively comparable, apart from purification steps related to antibody and binding protein construct. Both the cellulose resin and dsRNA binding protein construct methods provide highly enriched dsRNA extracts suitable for HTS with the B2 method providing a 36× and the ReliaPrep™ Resin a 163× increase in dsRNA enrichment compared to the mAb rJ2 antibody. The overall consistency and cost effectiveness of the ReliaPrep™ cellulose resin-based method and the potentially simpler adaptation to robotics made it the method of choice for future transfer to a semi-automated workflow.

The Generation and Characterization of Monoclonal Antibodies against the MPXV A29L Protein

Mpox (formerly known as monkeypox) is a zoonotic disease caused by monkeypox virus (MPXV), a DNA virus belonging to the Orthopoxvirus genus, in the Poxviridae family. The disease constitutes a moderate risk to public health at the global level. The MPXV A29L protein plays a crucial role in coordinating virion assembly and facilitating important virus-host interactions. This study focused on the expression, purification, and recombinant protein synthesis of the A29L protein of MPXV using prokaryotic systems. Using hybridoma technology, we successfully generated the monoclonal antibodies (mAbs) 1E12 and 4B2, which specifically recognize the A29L protein. These mAbs were found to be suitable for use in indirect immunofluorescence assays (IFA), Western blotting, and immunoprecipitation (IP). Our investigation also revealed that mAbs 1E12 and 4B2 could detect the A27L protein, a homologous protein found in the vaccinia virus Western Reserve (VACV WR) strain, using IFA, Western blotting, and immunoprecipitation (IP). Using mAbs 1E12 and 4B2 as primary immunological probes, A27L protein expression was detected as early as 6 h postinfection with VACV WR, with increasing protein levels being observed throughout the infection. This study enhances our understanding of the protein structure and function of MPXV and contributes to the development of specific MPXV detection methods.

Profiling of viral load, antibody and inflammatory response of people with monkeypox during hospitalization: a prospective longitudinal cohort study in China

The dynamics of viral shedding and the specific humoral response against monkeypox virus (MPXV) have not been well characterized in patients across their disease course during hospitalisation. The aim of this study was to determine the viral load and the levels of antibodies against MPXV using longitudinal paired-collected samples from hospitalized patients. Patients who were hospitalised with mpox were recruited at Beijing Ditan Hospital Capital Medical University in China between June 2 and September 23, 2023. Paired samples, including samples from skin lesions, the oropharynx, saliva, faeces, urine, plasma, and serum, were serially collected at days 1, 3, 7, and 14 after admission until discharge. Not all of the patients had samples obtained at all of the timepoints. All the samples were analysed via quantitative PCR. Virus isolation was performed by using clinical samples and Vero cells. The presence of IgM, IgA, IgG, and neutralising antibodies (NAbs) against MPXV was evaluated. The first collected plasma sample was taken when the patient was hospitalised, and the levels of cytokines and chemokines were measured in the sample. The demographic data, smallpox vaccination status, history of known exposure to MPVX, HIV status and other clinical data were collected using a standard case report form. A total of 510 specimens were serially collected from 39 recruited people with mpox. Among all the samples, the skin lesions had the highest viral DNA detection rates and viral loads, and the saliva samples had the second highest rates and viral loads. One day before discharge, 85% of the dry scrabs (median Ct 28.2, range 19.0-38.3) and 70% of the saliva samples (median Ct 32.4, range 24.5-38.1) were positive for viral DNA, Of which, 23.1% of dry scrabs were positive in viral culture. The rate of viral DNA detection in the oropharyngeal, saliva, and faecal samples decreased with time, while the rates in the plasma, serum, and urine samples increased quickly before 10 days post symptom onset (PSO). The median days of appearance of MPXV-IgM, MPXV-IgA, MPXV-IgG, and NAb were at 8(interquartile range [IQR] 7-9), 9 (7-10), 12 (9-15), and 12 (9-15) PSO, respectively. The IgM, IgA, IgG, and NAb titres increased with time. Between days 11 and 21 PSO, the NAb titres were lower in people living with HIV (PWH) than in people living without HIV (PWOH). Increased NAb titres were associated with decreased viral loads in the saliva (r=0.28, p=0.025), faeces (r=0.35, p=0.021), plasma (r=0.30, p=0.0044), and serum samples (r=0.37, p=0.001). Compared with PWOH, PWH had higher plasma levels of MIP-1α, MIP-1β, G-CSF, IL-4, and FGF-basic. The high positive viral culture rate of clinical samples of patients when they are discharged from the hospital indicates that effective public health management strategies are needed for people with mpox. The low NAb titres and high levels of cytokines in PWH shows that earlier treatment is needed to control inflammation in high-risk populations. National Natural Science Foundation of China, Chinese Academy of Medical Sciences, Fundamental Research Funds for the Central Universities for Peking Union Medical College, National Key R&D Program of China.

Exploring protein-mediated compaction of DNA by coarse-grained simulations and unsupervised learning

Protein-DNA interactions and protein-mediated DNA compaction play key roles in a range of biological processes. The length scales typically involved in DNA bending, bridging, looping, and compaction (≥ 1 kbp) are challenging to address experimentally or by all-atom molecular dynamics simulations, making coarse-grained simulations a natural approach. Here we present a simple and generic coarse-grained model for the DNA-protein and protein-protein interactions, and investigate the role of the latter in the protein-induced compaction of DNA. Our approach models the DNA as a discrete worm-like chain. The proteins are treated in the grand-canonical ensemble and the protein-DNA binding strength is taken from experimental measurements. Protein-DNA interactions are modeled as an isotropic binding potential with an imposed binding valency, without specific assumptions about the binding geometry. To systematically and quantitatively classify DNA-protein complexes, we present an unsupervised machine learning pipeline that receives a large set of structural order parameters as input, reduces the dimensionality via principal component analysis, and groups the results using a Gaussian mixture model. We apply our method to recent data on the compaction of viral genome-length DNA by HIV integrase and we find that protein-protein interactions are critical to the formation of looped intermediate structures seen experimentally. Our methodology is broadly applicable to DNA-binding proteins and to protein-induced DNA compaction and provides a systematic and semi-quantitative approach for analyzing their mesoscale complexes.

High production of recombinant protein using geminivirus-based deconstructed vectors in Nicotiana benthamiana

We focused on the geminiviral vector systems to develop an efficient vector system for plant biotechnology. Begomoviruses and curtoviruses, which belong to the Geminiviridae family, contain an intergenic region (IR) and four genes involved in replication, including replication-associated protein (Rep, C1), transcriptional activator (TrAP, C2), and replication enhancer (REn, C3). Geminiviruses can amplify thousands of copies of viral DNA using plant DNA polymerase and viral replication-related enzymes and accumulate viral proteins at high concentrations. In this study, we optimized geminiviral DNA replicon vectors based on tomato yellow leaf curl virus (TYLCV), honeysuckle yellow vein virus (HYVV), and mild curly top virus (BMCTV) for the rapid, high-yield plant-based production of recombinant proteins. Confirmation of the optimal combination by co-delivery of each replication-related gene and each IR harboring the Pontellina plumata-derived turbo green fluorescence protein (tGFP) gene via agroinfiltration in Nicotiana benthamiana leaves resulted in efficient replicon amplification and robust protein production within 3 days. Co-expression with the p19 protein of the tomato bush stunt virus, a gene-silencing suppressor, further enhanced tGFP accumulation by stabilizing mRNA. With this system, tGFP protein was produced at 0.7–1.2 mg/g leaf fresh weight, corresponding to 6.9–12.1% in total soluble protein. These results demonstrate the advantages of rapid and high-level production of recombinant proteins using the geminiviral DNA replicon system for transient expression in plants.

Influenza virus infection activates TAK1 to suppress RIPK3-independent apoptosis and RIPK1-dependent necroptosis

Many DNA viruses develop various strategies to inhibit cell death to facilitate their replication. However, whether influenza A virus (IAV), a fast-replicating RNA virus, attenuates cell death remains unknown. Here, we report that IAV infection induces TAK1 phosphorylation in a murine alveolar epithelial cell line (LET1) and a murine fibroblastoma cell line (L929). The TAK1-specific inhibitor 5Z-7-Oxzeneonal (5Z) and TAK1 knockout significantly enhance IAV-induced apoptosis, as evidenced by increased PARP, caspase-8, and caspase-3 cleavage. TAK1 inhibition also increases necroptosis as evidenced by increased RIPK1S166, RIPK3T231/S232, and MLKLS345 phosphorylation. Mechanistically, TAK1 activates IKK, which phosphorylates RIPK1S25 and inhibits its activation. TAK1 also activates p38 and its downstream kinase MK2, which phosphorylates RIPK1S321 but does not affect RIPK1 activation. Further investigation revealed that the RIPK1 inhibitor Nec-1 and RIPK1 knockout abrogate IAV-induced apoptosis and necroptosis; re-expression of wild-type but not kinase-dead (KD)-RIPK1 restores IAV-induced cell death. ZBP1 knockout abrogates IAV-induced cell death, whereas RIPK3 knockout inhibits IAV-induced necroptosis but not apoptosis. 5Z treatment enhances IAV-induced cell death and slightly reduces the inflammatory response in the lungs of H1N1 virus-infected mice and prolongs the survival of IAV-infected mice. Our study provides evidence that IAV activates TAK1 to suppress RIPK1-dependent apoptosis and necroptosis, and that RIPK3 is required for IAV-induced necroptosis but not apoptosis in epithelial cells.

Analysis of the Unique Historical Isolate of African Swine Fever Virus Isolate Spencer from Outbreaks in 1951

African swine fever (ASF) is a deadly hemorrhagic disease of domestic and wild swine that was first described in the early 20th century after the introduction of European pigs to Kenya. The etiological agent, the African swine fever virus (ASFV), is a large DNA virus within the Asfarviridae family that is broadly categorized epidemiologically into genotypes based on the nucleotide sequence of B646L, the gene encoding the major capsid protein p72. ASF outbreaks in Africa have been linked historically to 25 genotypes by p72 nucleotide analysis and, recently, to 6 genotypes by amino acid comparison, whereas global outbreaks of ASF outside of Africa have only been linked to 2 genotypes: genotype I, which led to an outbreak in Europe during the 1960s that later spread to South America, and genotype II, responsible for the current pandemic that began in Georgia in 2007 and has since spread to Europe, Asia, and Hispaniola. Here, we present an analysis of the genome of ASFV Spencer, an isolate that was collected in 1951 near Johannesburg, South Africa. While nucleotide analysis of Spencer indicates the p72 coding sequence is unique, differentiating from the closest reference by five nucleotides, the predicted amino acid sequence indicates that it is 100% homologous to contemporary genotype 1. Full genome analysis reveals it is more similar to Mkuzi1979 and encodes genes that share similarity with either genotype 1 or genotype 2 outbreak strains.

Epidemiology and genotypic diversity of feline bocavirus identified from cats in Harbin, China

Feline bocavirus (FBoV) is a globally distributed linear, single-stranded DNA virus infect cats, currently classified into three distinct genotypes. Although FBoV can lead to systemic infections, its complete pathogenic potential remains unclear. In this study, 289 blood samples were collected from healthy cats in Harbin, revealing an overall FBoV prevalence of 12.1%. Notably, genotypes 1 and 3 of FBoV were found co-circulating among the cat population in Harbin. Additionally, recombination events were detected, particularly in the newly discovered NG/104 and DL/102 strains. Furthermore, negative selection sites were predominantly observed across the protein coding genes of FBoV. These findings suggest a co-circulation of genetically diverse FBoV strains among cats in Harbin, indicate that purifying selection is the primary driving force shaping the genomic evolution of FBoV, and also underscore the importance of comprehensive surveillance efforts to enhance our understanding of the epidemiology and evolutionary characteristics of FBoV.

The plasma viral communities associate with clinical profiles in a large-scale haematological patients cohort

BackgroundHaematological patients exhibit immune system abnormalities that make them susceptible to viral infections. Understanding the relationship between the virome in the blood plasma of haematological patients and their clinical characteristic is crucial for disease management. We aimed to explore the presence of viral pathogens and identify close associations between viral infections and various clinical features.ResultsA total of 21 DNA viruses and 6 RNA viruses from 12 virus families were identified from 1383 patients. Patients with haematological diseases exhibited significantly higher diversity, prevalence, and co-detection rates of viral pathogens. During fever episodes, pathogen detection was notably higher, with Epstein-Barr virus (EBV) and Mucorales infections being the most probable culprits for fever symptoms in non-haematological patients. The detection rate of torque teno virus (TTV) significantly increases in haematological patients after transplantation and during primary lung infections. Additionally, TTV-positive patients demonstrate significantly higher absolute neutrophil counts, while C-reactive protein and procalcitonin levels are notably lower. Furthermore, TTV, cytomegalovirus, and parvovirus B19 (B19V) were found to be more prevalent in non-neutropenic patients, while non-viral pathogenic infections, such as Gram-negative bacteria and Mucorales, were more common in neutropenic patients. Pegivirus C (HPgV-C) infection often occurred post-transplantation, regardless of neutropenia. Additionally, some viruses such as TTV, B19V, EBV, and HPgV-C showed preferences for age and seasonal infections.ConclusionsAnalysis of the plasma virome revealed the susceptibility of haematological patients to plasma viral infections at specific disease stages, along with the occurrence of mixed infections with non-viral pathogens. Close associations were observed between the plasma virome and various clinical characteristics, as well as clinical detection parameters. Understanding plasma virome aids in auxiliary clinical diagnosis and treatment, enabling early prevention to reduce infection rates in patients and improve their quality of life.EnXnzQJ5tBc4iYM41USpCeVideo

HIV-1 Intasomes Assembled with Excess Integrase C-Terminal Domain Protein Facilitate Structural Studies by Cryo-EM and Reveal the Role of the Integrase C-Terminal Tail in HIV-1 Integration.

Retroviral integration is mediated by intasome nucleoprotein complexes wherein a pair of viral DNA ends are bridged together by a multimer of integrase (IN). Atomic-resolution structures of HIV-1 intasomes provide detailed insights into the mechanism of integration and inhibition by clinical IN inhibitors. However, previously described HIV-1 intasomes are highly heterogeneous and have the tendency to form stacks, which is a limiting factor in determining high-resolution cryo-EM maps. We have assembled HIV-1 intasomes in the presence of excess IN C-terminal domain protein, which was readily incorporated into the intasomes. The purified intasomes were largely homogeneous and exhibited minimal stacking tendencies. The cryo-EM map resolution was further improved to 2.01 Å, which will greatly facilitate structural studies of IN inhibitor action and drug resistance mechanisms. The C-terminal 18 residues of HIV-1 IN, which are critical for virus replication and integration in vitro, have not been well resolved in previous intasome structures, and its function remains unclear. We show that the C-terminal tail participates in intasome assembly, resides within the intasome core, and forms a small alpha helix (residues 271-276). Mutations that disrupt alpha helix integrity impede IN activity in vitro and disrupt HIV-1 infection at the step of viral DNA integration.