Research trend of MRI application for lumbar disc degeneration with low back pain: a bibliometric analysis.

Background: Low Back Pain (LBP) is a pervasive and complex musculoskeletal condition affecting over 80% of the global population. Lumbar Disc Degeneration (LDD) significantly contributes to LBP, and MRI is crucial for its diagnosis and understanding. This study aimes to provide a comprehensive bibliometric analysis of MRI research on LDD with LBP, shedding light on research patterns, collaborations, and potential knowledge gaps. Methods: A comprehensive online search was conducted in the Scopus database to retrieve published literature on LDD with LBP. Bibliometric analysis was conducted to assess publication patterns, co-authorship networks, keyword co-occurrence, and co-citation analysis within the MRI applications for LDD research domain. Bibliometric analysis tools such as VOSviewer and the R package "bibliometrix" were utilized for quantitative assessments. Results: A total of 1,619 publications related to MRI and LDD were analyzed. The analysis indicated a consistent annual growth rate of 4.62% in publications related to MRI and lumbar disc degeneration, reflecting a steady increase in research output over the past two decades. The USA, China, and Japan emerged as leading contributors. "SPINE", "European Spine Journal", and "Spine Journal" were the most productive journals in this domain. Key research themes identified included lumbar spine, low back pain, and magnetic resonance imaging. Network visualization shows that low back pain and magnetic resonance imaging were the most widely used keywords. Conclusion: The comprehensive bibliometric analysis of MRI applications for Lumbar Disc Degeneration offers insights into prevailing research patterns, highlights key contributors and journals, and identifies significant research themes. This study provides a foundation for future research efforts and clinical practices in the field, ultimately contributing to the advancement of patient care for individuals suffering from LDD and associated Low Back Pain.

Structural and optical performance of NiV/Ti multilayer mirrors for Z-pinch plasma diagnostic at the wavelength region of 350-450†eV.

This Letter reports on investigations of novel, to the best of our knowledge, NiV(Ni93V7)/Ti multilayer mirrors for the operation in the wavelength region of 350-450 eV. Such mirrors are promising optical components for the Z-pinch plasma diagnostic. The NiV/Ti multilayers show superior structural and optical performance compared to conventional Ni/Ti multilayers. Replacing Ni with NiV in multilayers decreases interface widths and enhances the contrast of the refractive index between the absorber and spacer layers. The improvement of interface quality contributes to the enhancement in reflectance. Under the grazing incidence of 13°, a peak reflectivity of 25.1% at 429 eV is achieved for NiV/Ti multilayers, while 17.7% at 427 eV for Ni/Ti.

Feasibility of diffusion tensor imaging in cervical spondylotic myelopathy using MUSE sequence.

BACKGROUND CONTEXT: The most frequent type of spinal cord injury is cervical spondylotic myelopathy (CSM). Conventional structural magnetic resonance imaging (MRI) is the gold diagnosis standard for CSM. Diffusion tensor imaging (DTI) could reflect microstructural changes in the spinal cord by tracing water molecular diffusion in early stages of CSM. However, due to the complex local anatomical structure and small field of view of the spinal cord, the imaging effect of traditional DTI imaging on the spinal cord is limited. MUSE (MUltiplexed Sensitivity-Encoding) -DTI is a novel diffusion-weighted imaging (DWI) sequence that achieves higher signal intensity through multiple excitation acquisition. MUSE sequence may improve the quality of spinal cord DTI imaging. STUDY DESIGN: Prospective study. PURPOSE: This study aimed to investigate the clinical diagnosis value of a novel protocol of MUSE-DTI in patients with cervical spondylotic myelopathy (CSM). PATIENT SAMPLE: From August 2021 to March 2022, a total of 60 subjects (22-71 years) were enrolled, including 51 CSM patients (22 males, 29 females) and 9 healthy subjects (4 males and 5 females). Each subject underwent a MUSE-DTI examination and a clinical Japanese Orthopedic Association (JOA) scale. OUTCOME MEASURES: We measured values of FA (Fractional Anisotropy), MD (Mean Diffusivity), AD (Axial Diffusivity), and RD (Radial Diffusivity), and collected the clinical JOA scores of each subject before the MR examination. METHODS: A 3.0T MR scanner (Signa Architect, GE Healthcare) performed the MUSE-DTI sequence on each subject. The cervical canal stenosis of subjects was classified from grade 0 to grade Ⅲ according to the method of an MRI grading system. FA, MD, AD, and RD maps were generated by postprocessing MUSE-DTI data on the GE workstation. Regions of interest (ROIs) were manually drawn at the C2 vertebral body level and C2/3-C6/7 intervertebral disc levels by covering the whole spinal cord. The clinical severity of myelopathy of subjects was assessed by the clinical Japanese Orthopedic Association scale (JOA). RESULTS: MUSE-DTI can acquire a high-resolution diffusion image compared to traditional DTI. The FAMCL values showed a decreasing trend from grade 0 to grade Ⅲ, while the MDMCL, ADMCL, and RDMCL values showed an overall increasing trend. Significant differences in MDMCL, ADMCL, and RDMCL values were found between adjacent groups among grades Ⅰ-Ⅲ (p<.05). The ADC2 values in CSM patients (grade I-Ⅲ) were significantly lower than in healthy individuals (grade 0) (p=.019). The clinical JOA score has a significant correlation with FAMCL (p=.035), MDMCL (p<.001), ADMCL (p<.001), and RDMCL (p<.001) values. CONCLUSIONS: MUSE-DTI displayed a better image quality compared to traditional DTI. MUSE-DTI parameters displayed a grade-dependent trend. All the MUSE-DTI parameters at MCL were correlated with the clinical JOA scores. The ADC2 values can reflect the secondary damage of distal spinal cord. Therefore, MUSE-DTI could be a reliable biomarker for clinical auxiliary diagnosis of spinal cord injury severity in cervical spondylotic myelopathy.

A GH19 lysozyme and peptidase from Myoviridae cyanophages lacking the typical holin-endolysin system exhibit lytic activity.

Most of the dsDNA cyanophages employ holin-endolysin lysis systems to damage the host cells. This study aimed to elucidate the lytic activity of ORF91 and ORF117 in the cyanophage MaMV-DH01, which lacked a conventional cholinesterase system. These two proteins contained Lyz-like superfamily domains and were annotated as a member of GH family 19 (named DHGH19) and peptidase (named DHpeptidase), respectively. Overexpression of DHGH19 in E. coli over a 5 h course demonstrated potent bactericidal activity, evident from significant growth inhibition, membrane damage, and leakage of intracellular enzymes of E. coli cells. However, the lytic activity of DHpeptidase was relatively weaker, exhibiting a bacteriostatic effect. It was important to highlight that the specific mutation of enzyme-catalyzed residues in DHGH19 (E122 and E131) showed that these were the essential amino acids for DHGH19 to exert its bactericidal activity. Furthermore, the lytic function of DHGH19 and DHpeptidase on cyanobacteria cells was confirmed by their overexpression in the cyanobacterium Synechocystis sp. PCC6803. Overall, this study provides novel insights into the lytic mechanism of Myoviridae cyanophage, offering potential alternatives for the development of GH19 and peptidase as new antibacterial agents in the future.

Intermittent theta burst stimulation for poststroke non-spatial attention deficit: a protocol of prospective, double-blinded, single-centre, randomised controlled trial in China.

INTRODUCTION: Attention deficit is the most common cognitive impairment after stroke, which can significantly hinder the recovery of both other cognitive domains and motor functions. Increasing evidence suggests that the left dorsolateral prefrontal cortex (DLPFC) is related to non-spatial attention functions, which indicates that it may be a promising target of repetitive transcranial magnetic stimulation (rTMS) for treating poststroke non-spatial attention deficit. Theta burst stimulation (TBS) is a modified pattern of rTMS that delivers shorter stimulation times and exhibits superior therapeutic efficacy. This study aims to provide evidence regarding the efficacy of intermittent TBS (iTBS) over the left DLPFC to improve poststroke non-spatial attention deficits and elucidate the potential neurophysiological mechanisms. METHODS AND ANALYSIS: In this single-centre, prospective, randomised, sham-controlled clinical trial, patients with non-spatial attention deficits (n=38) received 10 sessions of real iTBS (n=19) or sham iTBS (n=19) over the left DLPFC and a 30-min conventional attention training. Neuropsychological evaluations, electrophysiological examination and neuroimaging scan will be conducted at baseline, postintervention (second week) and 2-week follow-up (fourth week). The primary outcomes are the change in the Montreal Cognitive Assessment scores and the Digital Span Test scores from baseline to the end of the intervention (second week). The secondary outcomes comprise changes in magnetic resonance spectroscopy neuroimaging from baseline to the end of the intervention (second week) as well as attention test batteries (including tests of selective attention, sustained attention, divided attention and shifting attention) and ERP P300 from baseline to endpoint (fourth week). ETHICS AND DISSEMINATION: This study has been approved by the Institutional Ethical Committee of Tongji Hospital (ID: TJ-IRB20230879). All participants will sign the informed consent. Findings will be published in peer-reviewed journals and conference presentations. TRIAL REGISTRATION NUMBER: ChiCTR2300068669.

Eukaryotic translation elongation factor 1 alpha (eEF1A) inhibits Siniperca chuatsi rhabdovirus (SCRV) infection through two distinct mechanisms.

IMPORTANCE: Although a virus can regulate many cellular responses to facilitate its replication by interacting with host proteins, the host can also restrict virus infection through these interactions. In the present study, we showed that the host eukaryotic translation elongation factor 1 alpha (eEF1A), an essential protein in the translation machinery, interacted with two proteins of a fish rhabdovirus, Siniperca chuatsi rhabdovirus (SCRV), and inhibited virus infection via two different mechanisms: (i) inhibiting the formation of crucial viral protein complexes required for virus transcription and replication and (ii) promoting the ubiquitin-proteasome degradation of viral protein. We also revealed the functional regions of eEF1A that are involved in the two processes. Such a host protein inhibiting a rhabdovirus infection in two ways is rarely reported. These findings provided new information for the interactions between host and fish rhabdovirus.

Advances on genomes studies of large DNA viruses in aquaculture: A minireview.

Genomic studies of viral diseases in aquaculture have received more and more attention with the growth of the aquaculture industry, especially the emerging and re-emerging viruses whose genome could contain recombination, mutation, insertion, and so on, and may lead to more severe diseases and more widespread infections in aquaculture animals. The present review is focused on aquaculture viruses, which is belonged to two clades, Varidnaviria and Duplodnaviria, and one class Naldaviricetes, and respectively three families: Iridoviridae (ranaviruses), Alloherpesviridae (fish herpesviruses), and Nimaviridae (whispoviruses). The viruses possessed DNA genomes nearly or larger than 100 kbp with gene numbers more than 100 and were considered large DNA viruses. Genome analysis and experimental investigation have identified several genes involved in genome replication, transcription, and virus-host interactions. In addition, some genes involved in virus genetic variation or specificity were also discussed. A summary of these advances would provide reference to future discovery and research on emerging or re-emerging aquaculture viruses.

Fusing a TurboID tag with the Andrias davidianus ranavirus 2L reduced virus adsorption efficiency.

Andrias davidianus ranavirus (ADRV) is a member of the genus ranavirus (family Iridoviridae). ADRV 2L is an envelope protein that could be essential in viral infection. In the present study, the function of ADRV 2L was investigated by fusion with the biotin ligase TurboID tag. A recombinant ADRV with a V5-TurboID tag fused in the N-terminal of 2L (ADRVT-2L) and a recombinant ADRV expressing V5-TurboID (ADRVT) were constructed, respectively. Infection of the recombinant viruses and wild-type ADRV (ADRVWT) in the Chinese giant salamander thymus cell line (GSTC) showed that ADRVT-2L had reduced cytopathic effect and lower virus titers than the other two viruses, indicating the fusion of a big tag affected ADRV infection. Analysis of the temporal expression profile showed that the expression of V5-TurboID-2L was delayed than wild-type 2L. However, electron microscopy found that the virion morphogenesis was not affected in ADRVT-2L-infected cells. Furthermore, the virus binding assay revealed that the adsorption efficiency of ADRVT-2L was considerably decreased compared to the other two viruses. Therefore, these data showed that linking the TurboID tag to ADRV 2L affected virus adsorption to the cell membrane, which suggested an important role of 2L in virus entry into cells.

Genome Characteristics of Two Ranavirus Isolates from Mandarin Fish and Largemouth Bass.

Ranaviruses are promiscuous pathogens that threaten lower vertebrates globally. In the present study, two ranaviruses (SCRaV and MSRaV) were isolated from two fishes of the order Perciformes: mandarin fish (Siniperca chuatsi) and largemouth bass (Micropterus salmoides). The two ranaviruses both induced cytopathic effects in cultured cells from fish and amphibians and have the typical morphologic characteristics of ranaviruses. Complete genomes of the two ranaviruses were then sequenced and analyzed. Genomes of SCRaV and MSRaV have a length of 99, 405, and 99, 171 bp, respectively, and both contain 105 predicted open reading frames (ORFs). Eleven of the predicted proteins have differences between SCRaV and MSRaV, in which only one (79L) possessed a relatively large difference. A comparison of the sequenced six ranaviruses from the two fish species worldwide revealed that sequence identities of the six proteins (11R, 19R, 34L, 68L, 77L, and 103R) were related to the place where the virus was isolated. However, there were obvious differences in protein sequence identities between the two viruses and iridoviruses from other hosts, with more than half lower than 55%. Especially, 12 proteins of the two isolates had no homologs in viruses from other hosts. Phylogenetic analysis revealed that ranaviruses from the two fishes clustered in one clade. Further genome alignment showed five groups of genome arrangements of ranaviruses based on the locally collinear blocks, in which the ranaviruses, including SCRaV and MSRaV, constitute the fifth group. These results provide new information on the ranaviruses infecting fishes of Perciformes and also are useful for further research of functional genomics of the type of ranaviruses.

Quantitative evaluation of the spinal cord compression in patients with cervical spondylotic myelopathy using synthetic MRI.

Objectives: This work aimed to investigate the feasibility and diagnostic value of synthetic MRI, including T1, T2 and PD values in determining the severity of cervical spondylotic myelopathy (CSM). Methods: All subjects (51 CSM patients and 9 healthy controls) underwent synthetic MRI scan on a 3.0T GE MR scanner. The cervical canal stenosis degree of subjects was graded 0-III based on the method of a MRI grading system. Regions of interest (ROIs) were manually drawn at the maximal compression level (MCL) by covering the whole spinal cord to generate T1MCL, T2MCL, and PDMCL values in grade I-III groups. Besides, anteroposterior (AP) and transverse (Trans) diameters of the spinal cord at MCL were measured in grade II and grade III groups, and relative values were calculated as follows: rAP = APMCL/APnormal, rTrans = TransMCL/Transnormal. rMIN = rAP/rTrans. Results: T1MCL value showed a decreasing trend with severity of grades (from grade 0 to grade II, p < 0.05), while it increased dramatically at grade III. T2MCL value showed no significant difference among grade groups (from grade 0 to grade II), while it increased dramatically at grade III compared to grade II (p < 0.05). PDMCL value showed no statistical difference among all grade groups. rMIN of grade III was significantly lower than that of grade II (p < 0.05). T2MCL value was negatively correlated with rMIN, whereas positively correlated with rTrans. Conclusion: Synthetic MRI can provide not only multiple contrast images but also quantitative mapping, which is showed promisingly to be a reliable and efficient method in the quantitative diagnosis of CSM.

Biological and Genomic Characteristics of MaMV-DH01, a Novel Freshwater Myoviridae Cyanophage Strain.

The genomic traits of cyanophages and their potential for metabolic reprogramming of the host cell remain unknown due to the limited number of studies on cyanophage isolates. In the present study, a lytic Microcystis cyanophage, MaMV-DH01, was isolated and identified. MaMV-DH01 has an icosahedral head approximately 100 nm in diameter and a tail 260 nm in length. Its burst size is large, with approximately 145 phage particles/infected cell; it has a latent period of 2 days, and it shows high stability under pH and temperature stresses. Multiple infection (multiplicity of infection [MOI] 0.0001 to 100) results showed that when the MOI was 0.0001, MaMV-DH01 needed a longer time to lyse host cells. Cyanophage MaMV-DH01 has a double-stranded DNA genome of 182,372 bp, with a GC content of 45.35% and 210 predicted open reading frames (ORFs). These ORFs are related to DNA metabolism, structural proteins, lysis, host-derived metabolic genes, and DNA packaging. Phylogenetic trees based on the whole genome and two conserved genes (TerL and capsid) indicate that MaMV-DH01 is clustered with Ma-LMM01 and MaMV-DC, which are independent of other cyanophages. Collinearity analysis showed that the complete genome of MaMV-DH01 was longer than those of Ma-LMM01 and MaMV-DC, with lengths of 20,263 bp and 13,139 bp, respectively. We verified the authenticity of these excess DNA fragments and found that they are involved to various degrees in the MaMV-DH01 transcription process. Map overlays of environmental virus macrogenomic reads onto the MaMV-DH01 genome revealed that viral sequences similar to that of MaMV-DH01 are widespread in the environment. IMPORTANCE A novel freshwater Myoviridae cyanophage strain, MaMV-DH01, was isolated; this strain infects Microcystis aeruginosa FACHB-524, and the biological and genomic characteristics of MaMV-DH01 provide new insights for understanding the mechanism by which cyanophages infect cyanobacterial blooms.

Siniperca chuatsi Rhabdovirus (SCRV)-Induced Key Pathways and Major Antiviral Genes in Fish Cells.

Fish rhabdoviruses, including Siniperca chuatsi rhabdovirus (SCRV), are epidemic pathogens that harm fish aquaculture. To clarify the interactions between SCRV and its host and explore antiviral targets, the present study performed transcriptome analysis in a cultured S. chuatsi skin cell line (SCSC) after SCRV infection at 3, 12, 24, and 36 h post-infection (hpi). Comparison with control obtained 38, 353, 896, and 1452 differentially expressed genes (DEGs) in the detected time points, respectively. Further analysis of the Go terms and KEGG pathways revealed the key pathways "Cytokine-cytokine receptor interaction" and "interferon related pathways" in SCSC cells responding to SCRV infection. The significantly up-regulated genes in the pathways were also verified by qPCR. Furthermore, gene cloning and overexpression revealed that five interferon-stimulated genes (ISGs) IFI4407, IFI35, Viperin, IFIT1, and IFIT5 had the ability to inhibit SCRV replication in FHM (Fathead minnow) cells, especially an inhibition efficiency more than 50% was observed in IFI35 overexpressed cells. In summary, current study revealed the main innate immune pathways in S. chuatsi cells induced by SCRV infection and the major ISGs of S. chuatsi in controlling SCRV replication.

Environmental Factors and Their Threshold Affecting the Survival of Five Aquatic Animal Viruses in Different Animal Cells.

Aquatic animal viruses infect and transmit in aquatic environments, causing serious harm to the aquaculture industry and a variety of wild aquatic animals. How are they affected by environmental factors and do they represent potential threat to mammalian heath or not? Here, the effects of environmental factors (ultraviolet radiation (UV), temperature, pH, and drying) and their threshold on five epidemic aquatic animal viruses infecting amphibians and bony fish, including Rana grylio virus (RGV), Andrias davidianus ranavirus (ADRV), Grass carp reovirus (GCRV), Paralichthys olivaceus rhabdovirus (PORV), and Scophthalmus maximus rhabdovirus (SMRV), were measured and compared in a fish cell line. The examination of virus titers after different treatment in fish cells showed that the two iridoviruses, RGV and ADRV, had a higher tolerance to all of the environmental factors, such as they only had a decay rate of 22-36% when incubated at 37 °C for 7 days. However, the rhabdovirus SMRV was sensitive to all of the factors, with a decay rate of more than 80% in most of the treatments; even a complete inactivation (100%) can be observed after drying treatment. To address the potential threat to mammals, infectivity and limitation factors of the five viruses in Baby hamster kidney fibroblast cells (BHK-21) were tested, which showed that three of the five viruses can replicate at a low temperature, but a high temperature strongly inhibited their infection and none of them could replicate at 37 °C. This study clarified the sensitivity or tolerance of several different types of aquatic animal viruses to the main environmental factors in the aquatic environment and proved that the viruses cannot replicate in mammalian cells at normal physiological temperature.

Antiviral Activity of Crude Polysaccharide Derived from Seaweed against IHNV and IPNV In Vitro.

Both infectious hematopoietic necrosis virus (IHNV) and infectious pancreatic necrosis virus (IPNV) are the causative agents of acute and highly contagious diseases of juvenile salmonids, resulting in severe economic losses to these cold-water fish globally. There is an urgent need to explore antiviral agents against IHNV and IPNV due to the lack of commercially available vaccines and antiviral drugs. More importantly, the co-infection of IHNV and IPNV is prevalent in nature, which not only aggravates extensive damage to the salmonids but also poses challenges to its prevention and control. The antiviral effects of a crude polysaccharide derived from seaweed (CSP) on IHNV and IPNV were evaluated in this study separately. Furthermore, the underlying antiviral mechanisms of CSP to IHNV and IPNV were analyzed, respectively. The results showed that CSP possessed excellent safety and good ability to inhibit IHNV, IPNV, and their co-infection. CSP preferred to act at the early stage of viral infection. The antiviral mechanism of CSP on IHNV is possibly involved in preventing viral attachment and release, while in IPNV, it is involved in suppressing viral attachment, entry, and release. Taken together, the results of this study shed new light on developing novel agents against viral infection in salmonid fish.

Supplementation of Dietary Crude Lentinan Improves the Intestinal Microbiota and Immune Barrier in Rainbow Trout (Oncorhynchus mykiss) Infected by Infectious Hematopoietic Necrosis Virus.

The effects of crude lentinan (CLNT) on the intestinal microbiota and the immune barrier were evaluated in rainbow trout (Oncorhynchus mykiss) infected by infectious hematopoietic necrosis virus (IHNV). The results showed that supplementary CLNT declined the rainbow trout mortality caused by IHNV, which suggested that CLNT has preventive effects on IHNV infection. IHNV destroyed intestinal integrity, as well as caused the intestinal oxidative and damage in rainbow trout. Supplementary CLNT significantly strengthened the intestinal immune barrier by declining intestinal permeability, as well as enhancing intestinal antioxidant and anti-inflammatory abilities in IHNV-infected rainbow trout (P<0.05). In addition, CLNT modified the aberrant changes of intestinal microbiota induced by IHNV, mainly represented by promoting the growths of Carnobacterium and Deefgea and inhibiting Mycobacterium and Nannocystis. Especially, supplementing with CLNT significantly promoted the growth of short-chain fatty acid-producing bacteria (P<0.05) and consequently increased the production of acetic acid, butanoic acid, and hexanoic acid in the intestine of IHNV-infected rainbow trout. Furthermore, it was speculated that CLNT could regulate the self-serving metabolic pathways of intestinal microbiota induced by IHNV, such as fatty acid metabolism and amino acid metabolism. Together, CLNT played the antiviral effects on IHNV infection through strengthening the intestinal immune barrier, as well as regulating intestinal microbiota and SCFA metabolism in rainbow trout. The present data revealed that CLNT exerted a promising prebiotic role in preventing the rainbow trout from IHNV infection.

Development and characterization of a skin cell line from Chinese perch (Siniperca chuatsi) and its application in aquatic animal viruses.

Chinese perch (Siniperca chuatsi), an important fish for the aquaculture industry of China, is often affected by viral diseases. A stable and sensitive cell line can play an important role in virus identification and isolation, functional gene identification, virus pathogenic mechanism and antiviral immunity study. In the present study, a new cell line (S. chuatsi skin cell, SCSC) derived from the skin of S. chuatsi was established. The SCSC mainly consisted of fibroblastic-like cells, which grew well in M199 medium supplemented with 10% foetal bovine serum at 25°C. Chromosome analysis revealed that the SCSC (44%) has a diploid chromosome number of 2n = 48. The SCSC can be transfected and expressed exogenous gene efficiently. It also showed high sensitivity to several aquatic animal viruses from different families including Rhabdoviridae, Iridoviridae and Reoviridae. In addition, RT-PCR showed that S. chuatsi rhabdovirus (SCRV) started genome replication as early as 3 h post infection in the cells, which also induced the up-regulation of a variety of immune-related genes including these related to interleukin family, pattern recognition receptors, JAK-STAT pathway and interferon regulatory factors. In summary, current study provided a new tool in research of fish viruses and its interaction with host.

ADRV 12L: A Ranaviral Putative Rad2 Family Protein Involved in DNA Recombination and Repair.

The Andrias davidianus ranavirus (ADRV) is a member of the family Iridoviridae and belongs to the nucleocytoplasmic large DNA viruses. Based on genomic analysis, an ADRV-encoding protein, ADRV 12L, and its homologs from other iridoviruses were predicted as Rad2 family proteins based on the conserved amino acids, domains, and secondary structures. Expression analysis showed that the transcription of ADRV 12L started at 4 h post infection, and its expression was not inhibited by a DNA-replication inhibitor. Meanwhile, immunofluorescence localization showed that ADRV 12L mainly localized in viral factories and colocalized with the viral nascent DNA, which hinted at a possible role in DNA replication. Furthermore, a mutant ADRV lacking 12L (ADRV-Δ12L) was constructed. In both luciferase assays based on homologous recombination (HR) and double-strand break repair (DSBR) that followed, ADRV-Δ12L induced less luciferase activity than the wild-type ADRV, indicating that HR and DSBR were impaired in ADRV-Δ12L infected cells. In addition, infection with ADRV-Δ12L resulted in smaller plaque sizes and lower viral titers than that with wild-type ADRV, indicating an important role for 12L in efficient virus infection. Therefore, the results suggest that Rad2 homologs encoded by iridovirus have important roles in HR- and DSBR-process of the viral DNA and, thus, affect virus replication and the production of progeny virions.

Andrias davidianus Ranavirus (ADRV) Genome Replicate Efficiently by Engaging Cellular Mismatch Repair Protein MSH2.

As nucleocytoplasmic large DNA viruses, replication of ranaviruses (genus Ranavirus, family Iridoviridae) involves a series of viral and host proteins. We have described that the replication and transcription machinery of Andrias davidianus ranavirus (ADRV) which was isolated from the Chinese giant salamander contained host factors. Here, a new host factor, the MutS homolog 2 (MSH2), was proved as an important protein that participated in ADRV infection. Expression of MSH2 was stable during ADRV infection in cultured cells and it localized at the cytoplasmic viral factories and colocalized with virus nascent DNA, indicating its possible role in virus genome replication. Investigation of the viral proteins that interacted with MSH2 by co-immunoprecipitation showed that A. davidianus MSH2 can interact with ADRV-35L (possible components associated with virus transcription), ADRV-47L (virus DNA polymerase), and ADRV-98R. Further knockdown MSH2 expression by RNAi significantly reduced the late gene expression of ADRV. Additionally, MSH2 knockout by CRISPR/Cas9 significantly reduced viral titers, genome replication, and late gene transcription of ADRV. Thus, the current study proved that ADRV can engage cellular MSH2 for its efficient genome replication and late gene transcription, which provided new information for understanding the roles of host factors in ranavirus replication and transcription.

Functional Analysis of the Endopeptidase and Holin From Planktothrix agardhii Cyanophage PaV-LD.

A cyanophage PaV-LD, previously isolated from harmful filamentous cyanobacterium Planktothrix agardhii, was sequenced, and co-expression of its two ORFs in tandem, ORF123 and ORF124, inhibited growth on the model cyanobacterium Synechocystis sp. PCC6803 cells. However, the mechanism of action of ORF123 and ORF124 alone remains to be elucidated. In this study, we aimed to study the individual function of ORF123 or ORF124 from PaV-LD. Our data showed that the ORF123 encoded an endopeptidase, which harbored an M23 family peptidase domain and a transmembrane region. The expression of the endopeptidase in Escherichia coli alone revealed that the protein exhibited remarkable bacteriostatic activity, as evidenced by observation of growth inhibition, membrane damage, and leakage of the intracellular enzyme. Similarly, the holin, a membrane-associated protein encoded by the ORF124, showed weak bacteriostatic activity on E. coli. Moreover, deletion mutations indicated that the transmembrane domains of endopeptidase and holin were indispensable for their bacteriostatic activity. Meanwhile, the bacteriostatic functions of endopeptidase and holin on cyanobacteria cells were confirmed by expressing them in the cyanobacterium Synechocystis sp. PCC6803. Collectively, our study revealed the individual role of endopeptidase or holin and their synergistic bacteriolytic effect, which would contribute to a better understanding of the lytic mechanism of cyanophage PaV-LD.