Neutralizing monoclonal antibodies against the Gc fusion loop region of Crimean-Congo hemorrhagic fever virus.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a highly pathogenic tick-borne virus, prevalent in more than 30 countries worldwide. Human infection by this virus leads to severe illness, with an average case fatality of 40%. There is currently no approved vaccine or drug to treat the disease. Neutralizing antibodies are a promising approach to treat virus infectious diseases. This study generated 37 mouse-derived specific monoclonal antibodies against CCHFV Gc subunit. Neutralization assays using pseudotyped virus and authentic CCHFV identified Gc8, Gc13, and Gc35 as neutralizing antibodies. Among them, Gc13 had the highest neutralizing activity and binding affinity with CCHFV Gc. Consistently, Gc13, but not Gc8 or Gc35, showed in vivo protective efficacy (62.5% survival rate) against CCHFV infection in a lethal mouse infection model. Further characterization studies suggested that Gc8 and Gc13 may recognize a similar, linear epitope in domain II of CCHFV Gc, while Gc35 may recognize a different epitope in Gc. Cryo-electron microscopy of Gc-Fab complexes indicated that both Gc8 and Gc13 bind to the conserved fusion loop region and Gc13 had stronger interactions with sGc-trimers. This was supported by the ability of Gc13 to block CCHFV GP-mediated membrane fusion. Overall, this study provides new therapeutic strategies to treat CCHF and new insights into the interaction between antibodies with CCHFV Gc proteins.

Elucidating the photodissociation fingerprint and quantifying the determination of organic hydroperoxides in gas-phase autoxidation.

Hydroperoxides are formed in the atmospheric oxidation of volatile organic compounds, in the combustion autoxidation of fuel, in the cold environment of the interstellar medium, and also in some catalytic reactions. They play crucial roles in the formation and aging of secondary organic aerosols and in fuel autoignition. However, the concentration of organic hydroperoxides is seldom measured, and typical estimates have large uncertainties. In this work, we developed a mild and environmental-friendly method for the synthesis of alkyl hydroperoxides (ROOH) with various structures, and we systematically measured the absolute photoionization cross-sections (PICSs) of the ROOHs using synchrotron vacuum ultraviolet-photoionization mass spectrometry (SVUV-PIMS). A chemical titration method was combined with an SVUV-PIMS measurement to obtain the PICS of 4-hydroperoxy-2-pentanone, a typical molecule for combustion and atmospheric autoxidation ketohydroperoxides (KHPs). We found that organic hydroperoxide cations are largely dissociated by loss of OOH. This fingerprint was used for the identification and accurate quantification of the organic peroxides, and it can therefore be used to improve models for autoxidation chemistry. The synthesis method and photoionization dataset for organic hydroperoxides are useful for studying the chemistry of hydroperoxides and the reaction kinetics of the hydroperoxy radicals and for developing and evaluating kinetic models for the atmospheric autoxidation and combustion autoxidation of the organic compounds.

AcMNPV P74 is cleaved at R325 and R334 by proteinases of both OB and BBMV to expose a potential fusion peptide for oral infection.

Baculoviruses enter insect midgut epithelial cells via a set of occlusion-derived virion (ODV) envelope proteins called per os infectivity factors (PIFs). P74 of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV), which was the first identified PIF, is cleaved by an endogenous proteinase embedded within the occlusion body during per os infection, but the target site(s) and function of the cleavage have not yet been ascertained. Here, based on bioinformatics analyses, we report that cleavage was predicted at an arginine and lysine-rich region in the middle of P74. A series of recombinant viruses with site-directed mutants in this region of P74 were generated. R325 or R334 was identified as primary cleavage site. In addition, we showed that P74 is also cleaved by brush border membrane vesicles (BBMV) of the host insect at R325 or R334, instead of R195, R196, and R199, as previously reported. Simultaneous mutations in R195, R196, and R199 lead to instability of P74 during ODV release. Bioassays showed that mutations at both R325 and R334 significantly affected oral infectivity. Taken together, our data show that both R325 and R334 of AcMNPV P74 are the primary cleavage site for both occlusion body endogenous proteinase and BBMV proteinase during ODV release and are critical for oral infection. IMPORTANCE: Cleavage of viral envelope proteins is usually an important trigger for viral entry into host cells. Baculoviruses are insect-specific viruses that infect host insects via the oral route. P74, a per os infectivity factor of baculoviruses, is cleaved during viral entry. However, the function and precise cleavage sites of P74 remain unknown. In this study, we found that R325 or R334 between the N- and C-conserved domains of P74 was the primary cleavage site by proteinase either from the occlusion body or host midgut. The biological significance of cleavage seems to be the release of the potential fusion peptide at the N-terminus of the cleaved C-terminal P74. Our results shed light on the cleavage model of P74 and imply its role in membrane fusion in baculovirus per os infection.

Aleukemic Chronic Myeloid Leukemia Without Neutrophilia and Thrombocytosis: A Report From the BCR::ABL1 Pathology Group.

Chronic myeloid leukemia (CML) is characterized by leukocytosis with left-shifted neutrophilia, basophilia, eosinophilia, and variable thrombocytosis. However, extremely rare cases of patients with CML without significant leukocytosis and thrombocytosis (aleukemic phase [ALP] CML, or CML-ALP) have been reported. Due to its rarity and limited awareness, there remains a significant knowledge gap concerning the pathologic diagnosis, disease progression, and optimal patient management and outcomes. In this multi-institutional study, we investigated 31 patients with CML-ALP. Over half (54.8%) of patients had a history of or concurrent hematopoietic or nonhematopoietic malignancies. At time of diagnosis of CML-ALP, approximately 26.7% of patients exhibited neutrophilia, 56.7% had basophilia, and 13.3% showed eosinophilia. The median number of metaphases positive for t(9;22)(q34;q11.2) was 15, with a median of 38.5% of interphase nuclei positive for BCR::ABL1 by fluorescence in situ hybridization. The median BCR::ABL1 level was 26.14%. Remarkably, 14 (45.2%) patients were initially misdiagnosed or not diagnosed before karyotype or fluorescence in situ hybridization information for BCR::ABL1 became available. Twenty-five patients received tyrosine kinase inhibitors (TKIs). One patient developed blast crisis while on TKI treatment 8 months after initial diagnosis. With a median follow-up time of 46.1 months, 20 of 22 patients who received TKI therapy and had detailed follow-up information achieved complete cytogenetic remission or deeper, 15 achieved major molecular remission or deeper, and 10 achieved molecularly undetectable leukemia. In conclusion, given the frequent occurrence of prior or concurrent malignancies, aleukemic presentation, and low level of t(9;22)(q34;q11.2)/BCR::ABL1, misdiagnosis or delayed diagnosis is common among these patients. While these patients generally respond well to TKIs, rare patients may develop blastic transformation. It is therefore important for pathologists and hematologists to be aware of this highly unusual presentation of CML to ensure timely diagnosis and appropriate management.

Structural transitions during the cooperative assembly of baculovirus single-stranded DNA-binding protein on ssDNA.

Single-stranded DNA-binding proteins (SSBs) interact with single-stranded DNA (ssDNA) to form filamentous structures with various degrees of cooperativity, as a result of intermolecular interactions between neighboring SSB subunits on ssDNA. However, it is still challenging to perform structural studies on SSB-ssDNA filaments at high resolution using the most studied SSB models, largely due to the intrinsic flexibility of these nucleoprotein complexes. In this study, HaLEF-3, an SSB protein from Helicoverpa armigera nucleopolyhedrovirus, was used for in vitro assembly of SSB-ssDNA filaments, which were structurally studied at atomic resolution using cryo-electron microscopy. Combined with the crystal structure of ssDNA-free HaLEF-3 octamers, our results revealed that the three-dimensional rearrangement of HaLEF-3 induced by an internal hinge-bending movement is essential for the formation of helical SSB-ssDNA complexes, while the contacting interface between adjacent HaLEF-3 subunits remains basically intact. We proposed a local cooperative SSB-ssDNA binding model, in which, triggered by exposure to oligonucleotides, HaLEF-3 molecules undergo ring-to-helix transition to initiate continuous SSB-SSB interactions along ssDNA. Unique structural features revealed by the assembly of HaLEF-3 on ssDNA suggest that HaLEF-3 may represent a new class of SSB.

AC81 Is a Putative Disulfide Isomerase Involved in Baculoviral Disulfide Bond Formation.

The correct formation of native disulfide bonds is critical for the proper structure and function of many proteins. Cellular disulfide bond formation pathways commonly consist of two parts: sulfhydryl oxidase-mediated oxidation and disulfide isomerase-mediated isomerization. Some large DNA viruses, such as baculoviruses, encode sulfhydryl oxidases, but viral disulfide isomerases have not yet been identified, although G4L in poxvirus has been suggested to serve such a function. Here, we report that the baculovirus core gene ac81 encodes a putative disulfide isomerase. ac81 is conserved in baculoviruses, nudiviruses, and hytrosaviruses. We found that AC81 homologs contain a typical thioredoxin fold conserved in disulfide isomerases. To determine the role of AC81, a series of Autographa californica nucleopolyhedrovirus (AcMNPV) bacmids containing ac81 knockout or point mutations was generated, and the results showed that AC81 is essential for budded virus production, multinucleocapsid occlusion-derived virus (ODV) formation, and ODV embedding in occlusion bodies. Nonreducing Western blot analysis indicated that disulfide bond formation in per os infectivity factor 5 (PIF5), a substrate of the baculoviral sulfhydryl oxidase P33, was abnormal when ac81 was knocked out or mutated. Pulldown assays showed that AC81 interacted with PIF5 and P33 in infected cells. In addition, two critical regions that harbor key amino acids for function were identified in AC81. Taken together, our results suggest that AC81 is a key component involved in the baculovirus disulfide bond formation pathway and likely functions as a disulfide isomerase. IMPORTANCE Many large DNA viruses, such as poxvirus, asfarvirus, and baculovirus, encode their own sulfhydryl oxidase to facilitate the disulfide bond formation of viral proteins. Here, we show that AC81 functions as a putative disulfide isomerase and is involved in multiple functions of the baculovirus life cycle. Interestingly, AC81 and P33 (sulfhydryl oxidase) are conserved in baculoviruses, nudiviruses, and hytrosaviruses, which are all insect-specific large DNA viruses replicating in the nucleus, suggesting that viral disulfide bond formation is an ancient mechanism shared by these viruses.

Insights into Two-Metal-Ion Catalytic Mechanism of Cap-Snatching Endonuclease of Ebinur Lake Virus in Bunyavirales.

The cap-snatching endonuclease (EN) of segmented negative-strand RNA viruses (sNSVs) produces short capped primers for viral transcription by cleaving the host mRNAs. EN requires divalent metals as cofactors for nucleic acid substrates cleavage; however, the detailed mechanism of metal ion-dependent catalysis of ENs remains obscure. In this work, we reported the EN crystal structure of the Ebinur Lake virus (EBIV), an emerging mosquito-borne orthobunyavirus, and investigated its enzymatic properties and metal ion-based catalytic mechanism. In vitro biochemical data showed that EBIV EN is a specific RNA nuclease and prefers to cleave unstructured uridine-rich ssRNA. Structural comparison indicated that the overall structural architecture of EBIV EN is similar to that of other sNSV ENs, while the detailed active site configuration including the binding state of metal ions and the conformation of the LA/LB loop pair is different. Based on sequence conservation analysis, nine active site mutants were constructed, and seven crystal structures of them were determined. Mutations of active site residues associated with the two metal ions (Mn1 and Mn2) coordination abolished EN activity. Crystallographic analyses further revealed that none of these mutants bound two metal ions simultaneously in the active site. Importantly, we found that the perturbation of Mn1-coordination (metal site 1), resulted in the enhancement or elimination of Mn2-coordination (metal site 2). Taken together, our data provide structural evidence to support the two-metal-ion catalytic mechanism of EBIV EN and the correlation of metal binding at the two binding sites, which may be commonly shared by bunyaviruses or other sNSVs. IMPORTANCE The viral endonucleases (ENs) encoded by bunyaviruses and orthomyxoviruses play an essential role in initiating transcription by "snatching" capped primers from the host mRNAs. These ENs are metal-ion-dependent nucleases; however, the details of their catalytic mechanism remain elusive. Here, we reported high-resolution crystal structures of the wild-type and mutant ENs of a novel bunyavirus, the Ebinur Lake virus (EBIV), and revealed the structure and function relationship of EN. The EBIV EN exhibited differences in the details of active site structure compared to its homologues. Our data provided structural evidence to support a two-metal-ion catalytic mechanism of EBIV EN, and found the correlation of metal binding at both binding sites, which might reflect the dynamic structural properties that correlate to EN catalytic function. Taken together, our results revealed the structural characteristics of EBIV EN and made important implications for understanding the catalytic mechanism of cap-snatching ENs.

Structural and Biochemical Basis for Development of Diketo Acid Inhibitors Targeting the Cap-Snatching Endonuclease of the Ebinur Lake Virus (Order: Bunyavirales).

The Bunyavirales contain many important human pathogens that lack an antiviral therapy. The cap-snatching endonuclease (EN) of segmented negative-strand RNA viruses is an attractive target for broad-spectrum antivirals due to its essential role in initiating viral transcription. L-742,001, a previously reported diketo acid inhibitor against influenza virus EN, demonstrated potent EN inhibition and antiviral activity on various bunyaviruses. However, the precise inhibitory mechanism of the compound is still poorly understood. We recently characterized a highly active EN from Ebinur Lake virus (EBIV), a newly identified member of the Orthobunyavirus genus, and obtained its high-resolution structures, paving the way for structure-guided inhibitor development. Here, nine L-742,001 derivatives were designed and synthesized de novo, and their structure-activity relationship with EBIV EN was studied. In vitro biochemical data showed that the compounds inhibited the EBIV EN activity with different levels and could be divided into three categories. Five representative compounds were selected for further cell-based antiviral assay, and the results largely agreed with those of the EN assays. Furthermore, the precise binding modes of L-742,001 and its derivatives in EN were revealed by determining the high-resolution crystal structures of EN-inhibitor complexes, which suggested that the p-chlorobenzene is essential for the inhibitory activity and the flexible phenyl has the greatest exploration potential. This study provides an important basis for the structure-based design and optimization of inhibitors targeting EN of segmented negative-strand RNA viruses. IMPORTANCE The Bunyavirales contain many important human pathogens such as Crimean-Congo hemorrhagic fever virus and Lassa virus that pose serious threats to public health; however, currently there are no specific antiviral drugs against these viruses. The diketo acid inhibitor L-742,001 is a potential drug as it inactivates the cap-snatching endonuclease (EN) encoded by bunyaviruses. Here, we designed and synthesized nine L-742,001 derivatives and assessed the structure-activity relationship using EN of the newly identified Ebinur Lake virus (EBIV) as a research model. Our results revealed that the p-chlorobenzene of this broad-spectrum EN inhibitor is crucial for the inhibitory activity and the flexible phenyl "arm" has the best potential for further optimization. As cap-snatching ENs are present not only in bunyaviruses but also in influenza viruses, our data provide important guidelines for the development of novel and more potent diketo acid-based antiviral drugs against those viruses.

Functional Peroral Infectivity Complex of White Spot Syndrome Virus of Shrimp.

White spot syndrome virus (WSSV) is a major cause of disease in shrimp cultures worldwide. The infection process of this large circular double-stranded DNA virus has been well studied, but its entry mechanism remains controversial. The major virion envelope protein VP28 has been implicated in oral and systemic viral infection in shrimp. However, genetic analysis of viral DNA has shown the presence of a few genes related to proteins of per os infectivity factor (PIF) complex in baculoviruses. This complex is essential for the entry of baculoviruses, large terrestrial circular DNA viruses, into the midgut epithelial cells of insect larvae. In this study, we aimed to determine whether a PIF complex exists in WSSV, the components of this complex, whether it functions as an oral infectivity complex in shrimp, and the biochemical properties that contribute to its function in a marine environment. The results revealed a WSSV PIF complex (~720 kDa) comprising at least eight proteins, four of which were not identified as PIF homologs: WSV134, VP124 (WSV216), WSSV021, and WSV136. WSV134 is suggested to be a PIF4 homolog due to predicted structural similarity and amino acid sequence identity. The WSSV PIF complex is resistant to alkali, proteolysis, and high salt, properties that are important for maintaining infectivity in aquatic environments. Oral infection can be neutralized by PIF-specific antibodies but not by VP28-specific antibodies. These results indicate that the WSSV PIF complex is critical for WSSV entry into shrimp; the complex's evolutionary significance is also discussed. IMPORTANCE White spot disease, caused by the white spot syndrome virus (WSSV), is a major scourge in cultured shrimp production facilities worldwide. This disease is only effectively controlled by sanitation. Intervention strategies are urgently needed but are limited by a lack of appropriate targets. Our identification of a per os infectivity factor (PIF) complex, which is pivotal for the entry of WSSV into shrimp, could provide new targets for antibody- or dsRNA-based intervention strategies. In addition, the presence of a PIF complex with at least eight components in WSSV, which is ancestrally related to the PIF complex of invertebrate baculoviruses, suggests that this complex is structurally and functionally conserved in disparate virus taxa.

Structural Characterization of Per Os Infectivity Factor 5 (PIF5) Reveals the Essential Role of Intramolecular Interactions in Baculoviral Oral Infectivity.

Baculoviruses initiate oral infection in the highly alkaline midgut of insects via a group of envelope proteins called per os infectivity factors (PIFs). To date, no high-resolution structural information has been reported for any PIF. Here, we present the crystal structure of the PIF5 ectodomain (PIF5e) from Autographa californica multiple nucleopolyhedrovirus (AcMNPV) at a 2.2-Å resolution. It revealed an open cavity between the N-terminal E1 domain and the C-terminal E2 domain and a cysteine-rich region with three pairs of disulfide bonds in the E2 domain. Multiple conserved intramolecular interactions within PIF5 are essential for maintaining its tertiary structure. Two conserved arginines (Arg8 and Arg74) play critical roles in E1-E2 interactions, and mutagenesis analysis supported their crucial role in oral infection. Importantly, the reduction in the oral infectivity of the Arg8, Arg74, or cysteine mutant viruses was related to the proteolytic cleavage of PIF5 by the endogenous protease embedded in occlusion bodies during alkaline treatment. This suggested that the structural stability of PIF5 under physiological conditions in the insect midgut is critical for baculoviral oral infectivity. IMPORTANCEPer os infection mediated by PIFs is the highly complex mechanism by which baculoviruses initiate infection in insects. Previous studies revealed that multiple PIF proteins form a large PIF complex on the envelope of virions, while PIF5 functions independently of the PIF complex. Here, we report the crystal structure of AcMNPV PIF5e, which, to our knowledge, is the first atomic structure reported for a PIF protein. The structure revealed the precise locations of three previously proposed disulfide bonds and other conserved intramolecular interactions, which are important for the structural stability of PIF5 and are also essential for oral infectivity. These findings advance our understanding of the molecular mechanism of baculovirus oral infection under alkaline conditions.

Sphingomyelin-Sequestered Cholesterol Domain Recruits Formin-Binding Protein 17 for Constricting Clathrin-Coated Pits in Influenza Virus Entry.

Influenza A virus (IAV) is a global health threat. The cellular endocytic machineries harnessed by IAV remain elusive. Here, by tracking single IAV particles and quantifying the internalized IAV, we found that sphingomyelin (SM)-sequestered cholesterol, but not accessible cholesterol, is essential for the clathrin-mediated endocytosis (CME) of IAV. The clathrin-independent endocytosis of IAV is cholesterol independent, whereas the CME of transferrin depends on SM-sequestered cholesterol and accessible cholesterol. Furthermore, three-color single-virus tracking and electron microscopy showed that the SM-cholesterol complex nanodomain is recruited to the IAV-containing clathrin-coated structure (CCS) and facilitates neck constriction of the IAV-containing CCS. Meanwhile, formin-binding protein 17 (FBP17), a membrane-bending protein that activates actin nucleation, is recruited to the IAV-CCS complex in a manner dependent on the SM-cholesterol complex. We propose that the SM-cholesterol nanodomain at the neck of the CCS recruits FBP17 to induce neck constriction by activating actin assembly. These results unequivocally show the physiological importance of the SM-cholesterol complex in IAV entry. IMPORTANCE IAV infects cells by harnessing cellular endocytic machineries. A better understanding of the cellular machineries used for its entry might lead to the development of antiviral strategies and would also provide important insights into physiological endocytic processes. This work demonstrated that a special pool of cholesterol in the plasma membrane, SM-sequestered cholesterol, recruits FBP17 for the constriction of clathrin-coated pits in IAV entry. Meanwhile, the clathrin-independent cell entry of IAV is cholesterol independent. The internalization of transferrin, the gold-standard cargo endocytosed solely via CME, is much less dependent on the SM-cholesterol complex. These results provide new insights into IAV infection and the pathway/cargo-specific involvement of the cholesterol pool(s).

Developments in the classification and nomenclature of arthropod-infecting large DNA viruses that contain pif genes.

Viruses of four families of arthropod-specific, large dsDNA viruses (the nuclear arthropod large DNA viruses, or NALDVs) possess homologs of genes encoding conserved components involved in the baculovirus primary infection mechanism. The presence of such homologs encoding per os infectivity factors (pif genes), along with their absence from other viruses and the occurrence of other shared characteristics, suggests a common origin for the viruses of these families. Therefore, the class Naldaviricetes was recently established, accommodating these four families. In addition, within this class, the ICTV approved the creation of the order Lefavirales for three of these families, whose members carry homologs of the baculovirus genes that code for components of the viral RNA polymerase, which is responsible for late gene expression. We further established a system for the binomial naming of all virus species in the order Lefavirales, in accordance with a decision by the ICTV in 2019 to move towards a standardized nomenclature for all virus species. The binomial species names for members of the order Lefavirales consist of the name of the genus to which the species belongs (e.g., Alphabaculovirus), followed by a single epithet that refers to the host species from which the virus was originally isolated. The common names of viruses and the abbreviations thereof will not change, as the format of virus names lies outside the remit of the ICTV.

Endoscopic Balloon Dilatation-Based Membrane Resection for Membranous Duodenal Stenosis: A Feasibility and Safety Study (With Video).

OBJECTIVES: Surgery is generally considered as the first-line therapy for membranous duodenal stenosis (MDS) in children. However, abdominal surgery leaves permanent scars and may even cause intestinal adhesion. Therefore, an effective, safe, and minimally invasive method is urgently needed. This study aimed to evaluate the safety, efficacy, and feasibility of endoscopic balloon dilatation-based membrane resection (EBD-MR) to treat MDS in children. METHODS: We retrospectively reviewed patients with MDS treated with EBD-MR in Shanghai Children's Hospital from May 2016 to August 2021. Primary study outcome was clinical success, defined as weight gain and complete remission of vomiting, without the need for repeat endoscopic or surgical intervention during follow-up. Secondary outcomes included technical success, diameter changes of the membrane opening, and adverse events. RESULTS: Nineteen children (9 females, mean age 14.5 ± 11.2 months) received endoscopic treatment for MDS, and clinical success was achieved in 18 of 19 patients (94.7%). No bleeding, perforation, and jaundice occurred. Diameters of the membrane opening increased from 2.97 ± 2.87 mm to 9.78 ± 1.27 mm after the treatment, symptoms of vomiting did not reappear during 10-73 months of follow-up, and body mass index of the children increased from 14.9 ± 2.2 kg/m 2 (pre-operation) to 16.2 ± 3.7 kg/m 2 (6 months after operation). One patient required surgical revision because of existence of a second web; three patients received 2-3 sessions of endoscopic treatment to obtain the final remission. CONCLUSIONS: The EBD-MR technique is safe, effective, and feasible for MDS, which provided an excellent alternative to surgical management for the disease in pediatric patients.

Genome Sequencing and Organization of Three Geographically Different Isolates of Nucleopolyhedrovirus from the Gypsy Moth Reveal Significant Genomic Differences.

Background: The gypsy moth (Lymantria dispar L., Lepidoptera: Erebidae) is a worldwide pest of trees and forests. Lymantria dispar nucleopolyhedrovirus (LdMNPV) belongs to the Baculoviridae family and is an insect virus specific to gypsy moth larvae. In this study, we describe the complete genome sequences of three geographically diverse isolates, H2 (China), J2 (Japan), and T3 (Turkey), of Lymantria dispar multiple nucleopolyhedrovirus (LdMNPV). Methods: The genomes of isolates H2, J2, and T3 were subjected to shotgun pyrosequencing using Roche 454 FLX and assembled using Roche GS De Novo Assembler. Comparative analysis of all isolates was performed using bioinformatics methods. Results: The genomes of LdMNPV-H2, J2, and T3 were 164,746, 162,249, and 162,614 bp in size, had GC content of 57.25%, 57.30%, and 57.46%, and contained 162, 165, and 164 putative open reading frames (ORFs ≥ 150 nt), respectively. Comparison between the reference genome LdMNPV-5/6 (AF081810) and the genomes of LdMNPV-H2, J2, and T3 revealed differences in gene content. Compared with LdMNPV-5/6, ORF5, 6, 8, 10, 31, and 67 were absent in LdMNPV-H2, ORF5, 13, and 66 were absent in LdMNPV-J2, and ORF10, 13, 31, and 67 were absent in LdMNPV-T3. In addition, the gene encoding the mucin-like protein (ORF4) was split into two parts in isolates H2 and T3 and designated ORF4a and ORF4b. Phylogenetic analysis grouped isolates H2 and J2 in a different cluster than isolate T3, which is more closely related to the Turkish and Polish isolates. In addition, H2 was found to be closely related to a South Korean LdMNPV isolate. Conclusion: This study provided a more detailed overview of the relationships between different geographic LdMNPV isolates. The results showed remarkable differences between groups at the genome level.

Improving the ranging performance of chaos LiDAR.

Chaos lidar has gained significant attention due to its high spatial resolution, natural anti-interference capability, and confidentiality. However, constrained by the power of the chaos laser, the sensitivity of the linear detector, and the hardware bandwidth, chaos lidar is greatly restricted in the application of long-distance target detection and imaging. To overcome these constraints, we propose a novel, to the best of our knowledge, chaos lidar based on Geiger mode avalanched photodetectors (GM-APDs) in a previous study called chaos single-photon (CSP) lidar. In this paper, we compare the CSP lidar with the linear mode chaos lidars by combining with lidar equation. Regarding the ranging principle, the CSP lidar is fully digital and breaks through the constraints of a detector's bandwidth and ADC's sampling rate. The simulation results indicate that the detection range of the CSP lidar is approximately 35 times and 8 times greater than that of a continuous-wave chaos lidar and pulsed chaos lidar, respectively. Although the detection accuracy of the CSP lidar is only at the centimeter level and is lower than the linear mode chaos lidars, its consumption of storage resources and power is greatly reduced due to 1-bit quantization in the GM-APD. Additionally, we investigate the impact of GM-APD parameters on the signal-to-noise ratio (SNR) of the CSP lidar system and demonstrate that the dead time difference between GM-APDs has a negligible effect. In conclusion, we present and demonstrate a new chaos lidar system with a large detection range, high SNR, low storage resources and power consumption, and on-chip capability.

Leukemic phase follicular lymphoma in a young adult.

BACKGROUND: Leukemic presentation of follicular lymphoma (FL) is uncommon, with most cases reported in older adults. DESIGN: This report describes an unusual case of a young adult diagnosed with leukemic phase of FL. We reviewed the existing literature on this rare presentation of the disease and its potential impact on patient outcomes. RESULTS: Leukemic phase of FL in young adults can be mistaken for other high-grade hematologic malignancies. Morphology assessment and ancillary testing, such as flow cytometry and FISH analysis, can assist in achieving an accurate diagnosis of the leukemic phase of FL. Notably, our young patient responded well to therapy, which is different from what is typically observed in older patients who have a poorer prognosis. Further cases are needed to investigate the prognostic impact of the leukemic phase of FL in younger patients.

From the archives of MD Anderson Cancer Center Castleman disease involving the thymus gland: Case report and literature review.

Castleman disease is a nodal based disease and very rarely involves the thymus gland. We report a 52-year-old man who was found incidentally to have a single thymic mass by computerized tomography scan. Thymectomy was performed, and the gross specimen showed a well-circumscribed, multi-loculated cystic mass. Histologic examination showed thymus involved by Castleman disease, hyaline-vascular variant. The lesion was characterized by lymphoid follicles with wide mantle zones, variably lymphocyte-depleted germinal centers with sclerotic radial blood vessels, and prominent interfollicular/stromal changes including numerous endothelial venules with sclerotic walls and hyaline sclerosis, scattered and frequent dysplastic follicular dendritic cells and foci of dystrophic calcification. Immunohistochemical analysis showed that the follicle mantle zones were composed of numerous B-cells positive for CD20, PAX5, and IgD. Antibodies specific for CD21 and CD23 highlighted prominent follicular dendritic cell networks within follicles. There was no evidence of human herpes virus 8. We searched the literature and could identify only 10 additional cases of thymic CD. Previously reported cases included 8 unicentric and 2 multicentric, classified pathologically as plasma cell variant (n = 4), hyaline vascular variant (n = 3), and mixed (n = 3). Thymectomy, as was done in the currently reported case, most often leads to the diagnosis of Castleman disease and was a mainstay of treatment in other reported cases.

Identification of Six Thiolases and Their Effects on Fatty Acid and Ergosterol Biosynthesis in Aspergillus oryzae.

Thiolase plays important roles in lipid metabolism. It can be divided into degradative thiolases (thioase I) and biosynthetic thiolases (thiolases II), which are involved in fatty acid ß-oxidation and acetoacetyl-CoA biosynthesis, respectively. The Saccharomyces cerevisiae genome harbors only one gene each for thioase I and thiolase II, namely, Pot1 and Erg10, respectively. In this study, six thiolases (named AoErg10A to AoErg10F) were identified in Aspergillus oryzae genome using bioinformatics analysis. Quantitative reverse transcription-PCR (qRT-PCR) indicated that the expression of these six thiolases varied at different growth times and under different forms of abiotic stress. Subcellular localization analysis showed that AoErg10A was located in the cytoplasm, AoErg10B and AoErg10C were in the mitochondria, and AoErg10D, AoErg10E, and AoErg10F were in the peroxisome. Yeast heterologous complementation assays revealed that AoErg10A, AoErg10D, AoErg10E, AoErg10F, and cytoplasmic AoErg10B (AoErg10BΔMTS) recovered the phenotypes of S. cerevisiae erg10 weak and lethal mutants and that only AoErg10D, AoErg10E, and AoErg10F recovered the phenotype of the pot1 mutant that cannot use oleic acid as the carbon source. Overexpression of AoErg10s affected either the growth speed or the sporulation of the transgenic strains. In addition, the fatty acid and ergosterol content changed in all the AoErg10-overexpressing strains. This study revealed the function of six thiolases in A. oryzae and their effect on growth and fatty acid and ergosterol biosynthesis, which may lay the foundation for genetic engineering for lipid metabolism in A. oryzae or other fungi. IMPORTANCE Thiolases, including thioase I and thiolase II, play important roles in lipid metabolism. Aspergillus oryzae, one of the most industrially important filamentous fungi, has been widely used for manufacturing oriental fermented food such as sauce, miso, and sake for a long time. In addition, A. oryzae has a high capability in production of high lipid content and has been used for lipid production. Thus, it is very important to investigate the function of thiolases in A. oryzae. In this study, six thiolase (named AoErg10A to AoErg10F) were identified by bioinformatics analysis. Unlike other reported thiolases in fungi, three of the six thiolases showed dual functions of thioase I and thiolase II in S. cerevisiae, indicating that the lipid metabolism is more complex in A. oryzae. The reveal of function of these thiolases in A. oryzae can lay the foundation for genetic engineering for lipid metabolism in A. oryzae or other fungi.

Chaos single photon LIDAR and the ranging performance analysis based on Monte Carlo simulation.

With the advent of serial production lidars, single photon lidar faces an increasingly severe threat of crosstalk. In this paper, we first propose the concept of Chaos Single Photon (CSP) lidar and establish the theoretical model. In CSP lidar system, chaos laser replaces pulsed laser, and the physical random sequence generated by a Geiger mode avalanche photodiode (GM-APD) responding to chaos laser substitutes the traditional pseudo-random sequence. The mean density of '1' code of the CSP lidar system can exceed 10 million counts per second (Mcps) with a dead time immunity. The theoretical models of detection probability and false alarm rate are derived and demonstrated based on Poisson distribution. The bit error rate (BER) is introduced into the CSP lidar system for evaluating the range walk error intuitively. Additionally, the simulation results indicate that the CSP lidar system has a robust anti-crosstalk capability. Compared with the traditional pseudo-random single photon (PRSP) lidar system, the CSP lidar system not only overcomes range ambiguity but also has a signal to noise rate (SNR) of 60 times, reaching 10000 when the mean echo photoelectron number is 10 per nanosecond. Benefited from large-scale arrays and extremely high sensitivity properties of GM-APDs, we are looking forward to the applications of the CSP lidar in weak signal detection, remote mapping, autonomous driving, etc.

Inhibition of AoAur1 increases mycelial growth, hyphal fusion and improves physiological adaptation to high-temperature stress in Aspergillus oryzae.

Inositol phosphorylceramide (IPC) participates in hyphal growth and serves as a signaling molecule that enables fungi to adapt to diverse environments. Here, a gene, encodes IPC synthase, was identified from the Aspergillus oryzae 3.042 genome and designated AoAur1. The characteristics, phylogenetic evolution, and resistance to aureobasidin A of AoAur1 were analyzed. The expression pattern of AoAur1 was markedly downregulated under temperature stress. Additionally, an RNAi-AoAur1 strain in which the AoAur1 expression was inhibited had mycelial that grew more quickly, had a higher frequency of hyphal fusion, and was more resistant to high-temperature stress than the control. Gene expression profiles showed that the genes related to IPC biosynthesis were obviously downregulated, while AoCerS, which participates in dihydroceramide biosynthesis, increased in the RNAi-AoAur1 strain at the three temperature treatments. A metabolomic analysis revealed that the intracellular IPC content decreased, and the accumulation of dihydroceramide and galactosylceramide increased significantly in the RNAi-AoAur1 strain. Thus, the inhibition of AoAur1 reduced IPC level followed by an increase in the contents of dihydroceramide and galactosylceramide that promote mycelial growth and the formation of spores in the RNAi-AoAur1 strain. Interestingly, the inhibition of AoAur1 also induced the expression of hyphal fusion-related genes, which promote hyphal fusion, thus, contributing to the transduction of stress signal to enhance the ability of cells to adapt to temperature stress. Our results demonstrated that the downregulation of AoAur1 and a decrease in the accumulation of IPC is one of the mechanisms that enables A. oryzae to adapt low- and high-temperature stress.