Dissecting infectious bronchitis virus-induced host shutoff at the translation level.

Viruses have evolved a range of strategies to utilize or manipulate the host's cellular translational machinery for efficient infection, although the mechanisms by which infectious bronchitis virus (IBV) manipulates the host translation machinery remain unclear. In this study, we firstly demonstrate that IBV infection causes host shutoff, although viral protein synthesis is not affected. We then screened 23 viral proteins, and identified that more than one viral protein is responsible for IBV-induced host shutoff, the inhibitory effects of proteins Nsp15 were particularly pronounced. Ribosome profiling was used to draw the landscape of viral mRNA and cellular genes expression model, and the results showed that IBV mRNAs gradually dominated the cellular mRNA pool, the translation efficiency of the viral mRNAs was lower than the median efficiency (about 1) of cellular mRNAs. In the analysis of viral transcription and translation, higher densities of RNA sequencing (RNA-seq) and ribosome profiling (Ribo-seq) reads were observed for structural proteins and 5' untranslated regions, which conformed to the typical transcriptional characteristics of nested viruses. Translational halt events and the number of host genes increased significantly after viral infection. The translationally paused genes were enriched in translation, unfolded-protein-related response, and activation of immune response pathways. Immune- and inflammation-related mRNAs were inefficiently translated in infected cells, and IBV infection delayed the production of IFN-ß and IFN-λ. Our results describe the translational landscape of IBV-infected cells and demonstrate new strategies by which IBV induces host gene shutoff to promote its replication. IMPORTANCE: Infectious bronchitis virus (IBV) is a γ-coronavirus that causes huge economic losses to the poultry industry. Understanding how the virus manipulates cellular biological processes to facilitate its replication is critical for controlling viral infections. Here, we used Ribo-seq to determine how IBV infection remodels the host's biological processes and identified multiple viral proteins involved in host gene shutoff. Immune- and inflammation-related mRNAs were inefficiently translated, the translation halt of unfolded proteins and immune activation-related genes increased significantly, benefitting IBV replication. These data provide new insights into how IBV modulates its host's antiviral responses.

Terminal deoxynucleotidyl transferase expression in different subtypes of childhood B-cell acute lymphoblastic leukemia.

The lack of expression of terminal deoxynucleotidyl transferase (TdT) is frequently associated with KMT2A-rearranged subtype of pediatric acute lymphoblastic leukemia (ALL). However, this association has not been investigated extensively in the Asian population. A retrospective analysis of TdT expression in pediatric B-cell ALL (B-ALL) was performed in patients treated using the Taiwan Pediatric Oncology Group (TPOG) ALL 2002 and 2013 protocols. Among the 331 patients with B-ALL, 12 patients showed TdT negativity at initial diagnosis. Among these, eight patients showed KMT2A rearrangement (66.7%). Other patients showing negative TdT expression had ETV6::RUNX1, MEF2D-rearranged, and other B-ALL subtypes. However, in the context of KMT2A-rearranged B-ALL (n = 20), only eight patients showed TdT negativity. The 5-year event-free survival and overall survival of patients with and without TdT expression were 83.8% versus 46.8% (P <0.001) and 86.3% versus 55.4% (P = 0.004), respectively. Moreover, several aberrant markers, such as CD2, CD56, CD7, and CD117, were rarely expressed in the B-ALL samples, and if expressed, they were enriched in specific genetic subtypes. The results of this study indicate that immunophenotypic features are correlated with specific genetic subtypes of childhood B-ALL.

Accurate Finite Element Simulations of Dynamic Behaviour: Constitutive Models and Analysis with Deep Learning.

Owing to the challenge of capturing the dynamic behaviour of metal experimentally, high-precision numerical simulations have become essential for analysing dynamic characteristics. In this study, calculation accuracy was improved by analysing the impact of constitutive models using the finite element (FE) model, and the deep learning (DL) model was employed for result analysis. The results showed that FE simulations with these models effectively capture the elastic-plastic response, and the ZA model exhibits the highest accuracy, with a 26.0% accuracy improvement compared with other models at 502 m/s for Hugoniot elastic limit (HEL) stress. The different constitutive models offer diverse descriptions of stress during the elastic-plastic response because of temperature effects. Concurrently, the parameters related to the yield strength at quasi-static influence the propagation speed of elastic waves. Calculation show that the yield strength at quasi-static of 6061 Al adheres to y = ax + b for HEL stress. The R-squared (R2) and mean absolute error (MAE) values of the DL model for HEL stress predictions are 0.998 and 0.0062, respectively. This research provides a reference for selecting constitutive models for simulation under the same conditions.

Short-term steaming during processing impacts the quality of Citri Reticulatae 'Chachi' peel.

Citrus peel is a commonly used food-medicine material in the production of fast-moving consumer goods (FMCGs). For instance, Ganpu tea is manufactured by combining the peel of Citri Reticulatae 'Chachi' (PCRC) with Pu-erh tea. The alleviated irritation of PCRC through years of aging makes Citri reticulatae Pericarpium a traditional Chinese medicine. Herein, we introduced short-term steaming into the processing of PCRC to favor the quick removal of its irritation while retaining its food-medicine properties. Sensory evaluation and volatile component analysis showed that 60-s steaming reduced irritation of freshly prepared PCRC. Biological evaluations indicated no effects of steaming on the neuroprotective activity of PCRC. The process increased the contents of several bioactive ingredients, including hesperidin, nobiletin, tangeretin, and synephrine. In addition, physical indications of accelerating PCRC aging were observed. Taken together, our findings suggest that short-term steaming may offer a promising new possibility for enhancing the quality of citrus peel.

Efficacy evaluation of a bivalent subunit vaccine against classical swine fever virus and porcine circovirus type 2.

Classical swine fever virus (CSFV) and porcine circovirus type 2 (PCV2) are two of the most devastating and economically significant pathogens affecting pig populations worldwide. Administration of a combination of vaccines against swine pathogens has been demonstrated to be as efficacious as the administration of single vaccines. In this study, we developed and tested a novel bivalent subunit vaccine against CSFV and PCV2. The safety and efficacy of this vaccine were demonstrated in mice and specific pathogen-free (SPF) piglets. In addition to investigating the serological responses after immunization, challenge studies with both viruses were also conducted. The results showed that this CSFV/PCV2 bivalent vaccine elicited a high level of neutralizing antibodies against both viruses and provided protection in challenge studies. In conclusion, the CSFV/PCV2 bivalent vaccine is safe and effective against CSFV or PCV2 challenge.

miR-345-3p Modulates M1/M2 Macrophage Polarization to Inhibit Inflammation in Bone Infection via Targeting MAP3K1 and NF-κB Pathway.

Infection after fracture fixation (IAFF), a complex infectious disease, causes inflammatory destruction of bone tissue and poses a significant clinical challenge. miR-345-3p is a biomarker for tibial infected nonunion; however, the comprehensive mechanistic role of miR-345-3p in IAFF is elusive. In this study, we investigated the role of miR-345-3p in IAFF pathogenesis through in vivo and in vitro experiments. In vivo, in a rat model of IAFF, miR-345-3p expression was downregulated, accompanied by increased M1 macrophage infiltration and secretion of proinflammatory factors. In vitro, LPS induced differentiation of primary rat bone marrow-derived macrophages into M1 macrophages, which was attenuated by miR-345-3p mimics. miR-345-3p promoted M1 to M2 macrophage transition-it reduced the expression of cluster of differentiation (CD) 86, inducible NO synthase, IL-1ß, and TNF-α but elevated those of CD163, arginase-1, IL-4, and IL-10. MAPK kinase kinase 1 (MAP3K1), a target mRNA of miR-345-3p, was overexpressed in the bone tissue of IAFF rats compared with that in those of the control rats. The M1 to M2 polarization inhibited MAP3K1 signaling pathways in vitro. Conversely, MAP3K1 overexpression promoted the transition from M2 to M1. miR-345-3p significantly inhibited NF-κB translocation from the cytosol to the nucleus in a MAP3K1-dependent manner. In conclusion, miR-345-3p promotes the polarization of M1 macrophages to the M2 phenotype by inhibiting the MAP3K1 and NF-κB pathways. These findings provide insight into the pathogenesis and immunotherapeutic strategies for IAFF and offer potential new targets for subsequent research.

Effects of silver nanoparticles and various forms of silver on nitrogen removal by the denitrifier Pseudomonas stutzeri and their toxicity mechanisms.

Silver nanoparticles (AgNPs) are widely used in daily life and industry because of their excellent antibacterial properties. AgNPs can exist in wastewater in various forms, such as Ag+, Ag2SO4, Ag2CO3, Ag2S, Ag2O, and AgCl. To assess the potential environmental risk of AgNPs and various forms of Ag, their toxic effects were investigated using the common denitrifier species Pseudomonas stutzeri (P. stutzeri). The inhibitory effect of AgNPs and various forms of Ag on P. stutzeri growth and its denitrification performance occurred in a concentration-dependent manner. The denitrification efficiency of P. stutzeri decreased from 95%∼97% to 89∼95%, 74∼95%, and 56∼85% under low, medium, and high exposure doses, respectively, of AgNPs and various forms of Ag. The changes in cell membrane morphology and increases in lactate dehydrogenase (LDH) release indicated that AgNPs and various forms of Ag damaged the cell membrane of P. stutzeri. Oxidative stress caused by excessive accumulation of reactive oxygen species (ROS) increased superoxide dismutase (SOD) and catalase (CAT) activities and decreased glutathione (GSH) levels. Overall, this study will help elucidate the impact of AgNPs and their transformation products on nitrogen removal efficiency in wastewater biological treatment systems.

Discovery of Dual Function Agents That Exhibit Anticancer Activity via Catastrophic Nicotinamide Adenine Dinucleotide Depletion.

Nicotinamide adenine dinucleotide (NAD) is essentially involved in many biological processes of cancer cells, yet chemical intervention of NAD biosynthesis failed to obtain an optimal therapeutic benefit. We herein developed a new strategy to induce catastrophic NAD depletion by concurrently impairing NAD synthesis and promoting NAD consumption. We designed a series of new compounds that conjugate an inhibitor of nicotinamide phosphoribosyltransferase (NAMPT), a rate-limiting enzyme in the NAD salvage pathway, with a DNA-alkylating agent. Among them, compound 11b exhibited potent anticancer efficacy in cancer cell lines and mouse tumor models with intrinsic resistance to the parent compound FK866 or chlorambucil. Compound 11b caused catastrophic NAD depletion via a synergistic effect between the NAD salvage pathway blockade and DNA damage-triggered NAD consumption. Our findings suggest a new intervention strategy for causing catastrophic NAD depletion in cancer cells and provide basis for the development of new inhibitors targeting NAD metabolism.

Discovery of New Fusobacterium nucleatum Inhibitors to Attenuate Migratory Capability of Colon Cancer Cells by the Drug Repositioning Strategy.

Recent studies revealed that intestinal microbiota played important roles in colorectal cancer (CRC) carcinogenesis. Particularly, Fusobacterium nucleatum was confirmed to promote the proliferation and metastasis of CRC. Therefore, targeting F. nucleatum may be a potential preventive and therapeutic approach for CRC. Herein, 2,272 off-patent drugs were screened inhibitory activity against F. nucleatum. Among the hits, nitisinone was identified as a promising anti-F. nucleatum lead compound. Further optimization of nitisinone led to the discovery of more potent derivatives. Particularly, compounds 19q and 22c showed potent anti-F. nucleatum activity (MIC50 = 1 and 2 µg/mL, respectively) with low cytotoxicity. Among them, compound 19q effectively attenuated the migratory ability of MC-38 cells induced by F. nucleatum. Preliminary mechanism studies suggested that nitisinone and its derivatives might act by downregulating nitroreductase and tryptophanase. Thus, the development of small molecule F. nucleatum inhibitors represents an effective strategy to treat CRC.

Mifepristone inhibits hepatoma growth by enhancing the GR-HSP60-survivin interaction to facilitate survivin degradation.

Silencing of heat shock protein 60 (HSP60) suppresses the growth of hepatocellular carcinoma (HCC). Mifepristone inhibits HSP60 mRNA expression in Chlamydophila-infected epithelial cells. The aim of this study was to determine whether mifepristone could inhibit the growth of HCC cells by affecting the functions of HSP60. The effect of mifepristone on cell viability was examined by flow cytometry and a cell proliferation assay. Protein-protein interactions were examined using the immunoprecipitation assay. The anti-tumor effect of mifepristone was evaluated using a xenograft model. Our results indicated that mifepristone induces cell cycle arrest at the G1 phase and early-stage apoptosis in HCC cells. Instead of reducing the total amount of HSP60, mifepristone induced the release of mitochondrial HSP60 into the cytosol by causing a loss of ΔΨm, thereby enhancing glucocorticoid receptor (GR)-HSP60-survivin complex formation as well as survivin degradation. Animal models have confirmed the growth inhibitory effects of mifepristone on HCC, including changes in the abundance of HSP60 in mitochondria and cytosol, decreased survivin and Ki-67-positive cells, as well as increased cell apoptosis. In conclusion, the inhibition of HCC growth by mifepristone may be achieved by altering the subcellular distribution of HSP60 to enhance the formation of cytosolic GR-HSP60-survivin complexes in the cells, leading to the degradation of survivin.

Uncovering the first complete chloroplast genomics, comparative analysis, and phylogenetic relationships of the medicinal plants Rhamnus cathartica and Frangula alnus (Rhamnaceae).

Rhamnus cathartica and Frangula alnus are economically valuable medicinal plants from the Rhamnaceae family. However, their chloroplast genome structure, phylogenetic position, relationships, and evolution remain poorly understood. Herein, the complete chloroplast genome resources of R. cathartica and F. alnus have been added. The first comparative analysis of the Rhamnus and Frangula species based on complete chloroplast genomes was provided. The chloroplast genomes of R. cathartica and F. alnus exhibited a quadripartite structure, with total lengths of 161,149 bp and 161,255 bp, respectively. The lack of the infA and psbL genes does not negatively impact the normal functioning of Rhamnus and Frangula species. The rpl20 and rpl33 genes are undergoing rapid evolution. Rhamnus and Frangula species prefer amino acids with A/U-terminal codons. There were between 100 and 126 simple sequence repeats and between 38 and 100 long repeats. Several highly divergent intergenic regions (trnK-UUU-trnQ-UUG, atpH-atpI, trnY-GUA-trnE-UUC, trnG-GCC-trnfM-CAU, trnT-UGU-trnF-GAA, rpl20-rps12, and rpl22-rps19) and highly divergent genes (ycf3, ndhA, rpl32, and ycf1) were identified, which could serve as potential phylogenetic markers due to their variability. We reconstructed the phylogenetic relationships among Rhamnus species and F. alnus using complete chloroplast genomes. There is no significant correlation between the medicinal value of the species analyzed and their phylogenetic relationships. These results provide valuable insights for understanding the phylogenetic relationship and evolution of Rhamnus and Frangula species. These findings could serve as a foundation for future studies on the Rhamnaceae. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01331-7.

Controlling antifungal activity with light: Optical regulation of fungal ergosterol biosynthetic pathway with photo-responsive CYP51 inhibitors.

Invasive fungal infections (IFIs) have been associated with high mortality, highlighting the urgent need for developing novel antifungal strategies. Herein the first light-responsive antifungal agents were designed by optical control of fungal ergosterol biosynthesis pathway with photocaged triazole lanosterol 14α-demethylase (CYP51) inhibitors. The photocaged triazoles completely shielded the CYP51 inhibition. The content of ergosterol in fungi before photoactivation and after photoactivation was 4.4% and 83.7%, respectively. Importantly, the shielded antifungal activity (MIC80 ≥ 64 µg/mL) could be efficiently recovered (MIC80 = 0.5-8 µg/mL) by light irradiation. The new chemical tools enable optical control of fungal growth arrest, morphological conversion and biofilm formation. The ability for high-precision antifungal treatment was validated by in vivo models. The light-activated compound A1 was comparable to fluconazole in prolonging survival in Galleria mellonella larvae with a median survival of 14 days and reducing fungal burden in the mouse skin infection model. Overall, this study paves the way for precise regulation of antifungal therapy with improved efficacy and safety.

Discovery of potent natural product higenamine derivatives as novel Anti-Fusobacterium nucleatum agents.

Fusobacterium nucleatum (F. nucleatum) is closely associated with the occurrence and development of colorectal cancer (CRC). Discovery of specific antibacterial agents against F. nucleatum was urgent for the prevention and treatment of CRC. We screened a natural product library and successfully identified higenamine as an antibacterial hit against F. nucleatum. Further hit optimizations led to the discovery of new higenamine derivatives with improved anti-F. nucleatum activity. Among them, compound 7c showed potent antibacterial activity against F. nucleatum (MIC50 = 0.005 µM) with good selectivity toward intestinal bacteria and normal cells. It significantly inhibited the migration of CRC cells induced by F. nucleatum. Mechanism study revealed that compound 7c impaired the integrity of biofilm and cell wall, which represents a good starting point for the development of novel anti-F. nucleatum agents.

Discovery of BRD4-HDAC Dual Inhibitors with Improved Fungal Selectivity and Potent Synergistic Antifungal Activity against Fluconazole-Resistant Candida albicans.

Over the past several decades, invasive fungal infections, especially candidiasis, have caused dramatic morbidity and mortality due to ineffective antifungal drugs and severe drug resistance. Herein, new BRD4-histone deacetylase (HDAC) inhibitors were designed to restore the susceptibility of Candida albicans (C. albicans) to fluconazole (FLC). Interestingly, several compounds showed excellent selectivity against fungal HDACs. In particular, compound B2 showed excellent synergistic effect with FLC against resistant C. albicans (FICI = 0.063) with high selectivity against fungal HDACs (SI = 1653) and low cytotoxicity. Compound B2 effectively synergized with FLC and prevented biofilm formation and morphological transition in resistant C. albicans, potentiating the antifungal activity of FLC in vivo and significantly reducing kidney fungal loads. Thus, this drug combination is promising in the treatment of resistant C. albicans infections.

Target- and prodrug-based design for fungal diseases and cancer-associated fungal infections.

Invasive fungal infections (IFIs) are emerging as a serious threat to public health and are associated with high incidence and mortality. IFIs also represent a frequent complication in patients with cancer who are undergoing chemotherapy. However, effective and safe antifungal agents remain limited, and the development of severe drug resistance further undermines the efficacy of antifungal therapy. Therefore, there is an urgent need for novel antifungal agents to treat life-threatening fungal diseases, especially those with new mode of action, favorable pharmacokinetic profiles, and anti-resistance activity. In this review, we summarize new antifungal targets and target-based inhibitor design, with a focus on their antifungal activity, selectivity, and mechanism. We also illustrate the prodrug design strategy used to improve the physicochemical and pharmacokinetic profiles of antifungal agents. Dual-targeting antifungal agents offer a new strategy for the treatment of resistant infections and cancer-associated fungal infections.

Discovery of a new chemical scaffold for the treatment of superbug Candida auris infections.

Candida auris has emerged as a serious threat of public health and caused global epidemic due to multi-drug resistance, remarkable transmissibility and high mortality. To tackle the challenging super fungus, novel benzoanilide antifungal agents were discovered by an integrated strategy of phenotypic screen, hit optimization, antifungal assays and mechanism exploration. The most promising compound A1 showed potent in vitro and in vivo efficacy against Candida auris infection. Mechanism investigation revealed that compound A1 blocked the biosynthesis of virulence factors and fungal cell walls through the inhibition of glycosylphosphatidylinositol (GPI) and GPI-anchored proteins. Thus, compound A1 represents a promising lead compound to combat drug-resistant candidiasis.

Identification and Comparison of the Sialic Acid-Binding Domain Characteristics of Avian Coronavirus Infectious Bronchitis Virus Spike Protein.

Infectious bronchitis virus (IBV) infections are initiated by the transmembrane spike (S) glycoprotein, which binds to host factors and fuses the viral and cell membranes. The N-terminal domain of the S1 subunit of IBV S protein binds to sialic acids, but the precise location of the sialic acid binding domain (SABD) and the role of the SABD in IBV-infected chickens remain unclear. Here, we identify the S1 N-terminal amino acid (aa) residues 19 to 227 (209 aa total) of IBV strains SD (GI-19) and GD (GI-7), and the corresponding region of M41 (GI-1), as the minimal SABD using truncated protein histochemistry and neuraminidase assays. Both α-2,3- and α-2,6-linked sialic acids on the surfaces of CEK cells can be used as attachment receptors by IBV, leading to increased infection efficiency. However, 9-O acetylation of the sialic acid glycerol side chain inhibits IBV S1 and SABD protein binding. We further constructed recombinant strains in which the S1 gene or the SABD in the GD and SD genomes were replaced with the corresponding region from M41 by reverse genetics. Infecting chickens with these viruses revealed that the virulence and nephrotropism of rSDM41-S1, rSDM41-206, rGDM41-S1, and rGDM41-206 strains were decreased to various degrees compared to their parental strains. A positive sera cross-neutralization test showed that the serotypes were changed for the recombinant viruses. Our results provide insight into IBV infection of host cells that may aid vaccine design. IMPORTANCE To date, only α-2,3-linked sialic acid has been identified as a potential host binding receptor for IBV. Here, we show the minimum region constituting the sialic acid binding domain (SABD) and the binding characteristics of the S1 subunit of spike (S) protein of IBV strains SD (GI-19), GD (GI-7), and M41 (GI-1) to various sialic acids. The 9-O acetylation modification partially inhibits IBV from binding to sialic acid, while the virus can also bind to sialic acid molecules linked to host cells through an α-2,6 linkage, serving as another receptor determinant. Substitution of the putative SABD from strain M41 into strains SD and GD resulted in reduced virulence, nephrotropism, and a serotype switch. These findings suggest that sialic acid binding has diversified during the evolution of γ-coronaviruses, impacting the biological properties of IBV strains. Our results offer insight into the mechanisms by which IBV invades host cells.

Anticancer Effects of Antidepressants in Hepatocellular Carcinoma Cells.

BACKGROUND/AIM: An epidemiological investigation indicated that tricyclic antidepressants (TCAs) and selective serotonin reuptake inhibitors (SSRIs) were associated with a lower risk of hepatocellular carcinoma (HCC). Another previous study showed that seven antidepressants inhibited glucocorticoid receptor (GR)-mediated gene transcription, a pathway that is linked to various diseases, including cancer. It is known that the expression levels of GR in cancerous tissues are higher than those in noncancerous tissues in patients with HCC. Notably, among the seven antidepressants, amitriptyline (TCA), desipramine (TCA), and fluoxetine (SSRI) were found to induce apoptosis in HCC cells. Given this, we investigated whether four other GR-specific antidepressants, including mianserin (atypical antidepressant), tianeptine (atypical antidepressant), imipramine (TCA), and moclobemide (monoamine oxidase inhibitor, MAOI) affect the cell viability of HCC. MATERIALS AND METHODS: Cell proliferation and IC50 curves were determined by MTT assays. RESULTS: Imipramine and mianserin significantly inhibited HCC cell viability, whereas moclobemide and tianeptine did not. IC50 showed that the same dose of imipramine or mianserin led to significant inhibitory effects on HCC cells whereas there were only slight effects on normal human hepatocytes (HH). CONCLUSION: According to previous and present findings, TCAs, SSRIs and mianserin may have anti-tumor activity in HCC. However, the appropriate dose, frequency, and route of the administration still need to be determined in future preclinical and clinical studies.

Soft tissue tuberculosis detected by next-generation sequencing: A case report and review of literature.

BACKGROUND: Soft tissue tuberculosis is rare and insidious, with most patients presenting with a localized enlarged mass or swelling, which may be factors associated with delayed diagnosis and treatment. In recent years, next-generation sequencing has rapidly evolved and has been successfully applied to numerous areas of basic and clinical research. A literature search revealed that the use of next-generation sequencing in the diagnosis of soft tissue tuberculosis has been rarely reported. CASE SUMMARY: A 44-year-old man presented with recurrent swelling and ulcers on the left thigh. Magnetic resonance imaging suggested a soft tissue abscess. The lesion was surgically removed and tissue biopsy and culture were performed; however, no organism growth was detected. Finally, Mycobacterium tuberculosis was confirmed as the pathogen responsible for infection through next-generation sequencing analysis of the surgical specimen. The patient received a standardized anti-tuberculosis treatment and showed clinical improvement. We also performed a literature review on soft tissue tuberculosis using studies published in the past 10 years. CONCLUSION: This case highlights the importance of next-generation sequencing for the early diagnosis of soft tissue tuberculosis, which can provide guidance for clinical treatment and improve prognosis.

Mutagenesis of the di-leucine motif in the cytoplasmic tail of newcastle disease virus fusion protein modulates the viral fusion ability and pathogenesis.

BACKGROUND: Newcastle disease virus (NDV) is a highly infectious viral disease, which can affect chickens and many other kinds of birds. The main virulence factor of NDV, the fusion (F) protein, is located on the viral envelope and plays a major role in the virus' ability to penetrate cells and cause host cell fusion during infection. Multiple highly conserved tyrosine and di-leucine (LL) motifs in the cytoplasmic tail (CT) of the virus may contribute to F protein functionality in the viral life cycle. METHODS: To examine the contribution of the LL motif in the biosynthesis, transport, and function of the F protein, we constructed and rescued a NDV mutant strain, rSG10*-F/L537A, with an L537A mutation using a reverse genetic system. Subsequently, we compared the differences in the syncytium formation ability, pathogenicity, and replication levels of wild-type rSG10* and the mutated strain. RESULTS: Compared with rSG10*, rSG10*-F/L537A had attenuated syncytial formation and pathogenicity, caused by a viral budding defect. Further studies showed that the LL-motif mutation did not affect the replication, transcription, or translation of the virus genome but affected the expression of the F protein at the cell surface. CONCLUSIONS: We concluded that the LL motif in the NDV F CT affected the regulation of F protein expression at the cell surface, thus modulating the viral fusion ability and pathogenic phenotype.