Search Results (211)

Diarrhea and impaired intestinal development are major constraints in newly hatched domestic pigeons raised for meat production, leading to reduced growth performance and increased reliance on antimicrobial interventions. Cecropin AD (CAD) is a broad-spectrum antimicrobial peptide with potential activity against bacterial pathogens and may represent an alternative strategy for improving intestinal health in poultry. In this study, we evaluated the safety and biological effects of a recombinant Newcastle disease virus (NDV) expressing CAD (rTS-CAD₃) in one-day-old domestic pigeons. Pigeons were inoculated with rTS-CAD₃, parental TS09-C, or PBS and monitored for growth performance, diarrhea incidence, viral RNA persistence, inflammatory responses, intestinal bacterial abundance, gut microbiota composition, intestinal morphology, and epithelial barrier function. rTS-CAD₃ caused no mortality, detected only transient viral RNA, and did not induce excessive inflammatory responses. Compared with TS09-C and PBS controls, rTS-CAD₃ significantly increased body weight gain and average daily gain while reducing diarrhea incidence. Importantly, rTS-CAD₃ decreased intestinal Escherichia coli abundance, reshaped the gut microbiota by reducing Proteobacteria and Escherichia–Shigella while increasing Firmicutes and Lactobacillus, improved villus height and the villus-to-crypt ratio, and enhanced occludin expression. These findings demonstrate that NDV-vectored CAD delivery can reduce intestinal bacterial burden, increase occludin expression, and alleviate diarrhea in domestic pigeons. The rTS-CAD₃ represents a promising antimicrobial peptide-based strategy for reducing spontaneous diarrhea and dependence on conventional antibiotics in poultry production. Full article

Newcastle disease virus (NDV) is an important avian pathogen that can circulate in migratory birds, including Larus ridibundus. This study examined the transcriptional responses and anti-NDV activities associated with recombinant IFN-α and Mx proteins from L. ridibundus. The IFN-α and Mx open reading frames were cloned (GenBank: OP263971 and OP263970), expressed using a pET32a(+)/E. coli system, and assessed using chicken embryos, DF-1 cells, and primary gull lymphocytes. qRT-PCR analysis showed that NDV infection increased IFN-α and Mx transcript abundance in gull lymphocytes, with peak levels at 48 h post-infection (p < 0.01). In chicken embryos, recombinant IFN-α was associated with higher embryo survival and lower NDV NP transcript abundance than recombinant Mx under the tested conditions. In DF-1 cells and primary gull lymphocytes, 0.25 mg/mL recombinant IFN-α was associated with reduced NDV NP transcript abundance, and morphological inspection suggested less severe cytopathic effects in DF-1 cells. IFN-α treatment was also accompanied by lower transcription of TLR7, MyD88, IRF7, and Mx compared with the NDV group at 48 h. These data support a role for gull IFN-α in limiting NDV-associated transcriptional responses in vitro, while further protein-level and infectious-virus assays are required to define the underlying mechanism. Full article

Background: The simultaneous circulation of highly pathogenic avian influenza (HPAI) H5N8 (clade 2.3.4.4b) and virulent Newcastle disease virus (NDV) genotype VII creates a cumulative risk for the poultry industry, rendering monovalent vaccination insufficient. This study aimed to develop and evaluate a combined inactivated vaccine against AIV H5N8 and NDV genotype VII based on local endemic strains. Methods: The vaccine was formulated as a water-in-oil emulsion (30:70) using Montanide™ ISA 78 VG. Evaluation included physicochemical characterization, safety assessment in chickens, immunogenicity evaluation via the hemagglutination inhibition (HI) test, and protective efficacy following challenge with virulent isolates. Results: The formulation demonstrated high stability and complete safety without adverse reactions. By 28 days post-immunization, the vaccine induced a robust, balanced humoral response with antibody titers reaching 9.10 ± 0.23 log₂ against NDV and 9.00 ± 0.26 log₂ against AIV, indicating no antigenic interference. Upon challenge, the vaccinated group exhibited 100% clinical protection and survival, compared to 100% mortality in controls. Furthermore, vaccination significantly reduced viral shedding, limiting horizontal spread. Conclusions: The developed combined inactivated vaccine demonstrates high safety, stability, and protective efficacy. It represents a promising candidate for comprehensive specific prophylaxis in endemic regions, effectively limiting horizontal pathogen transmission and mitigating economic losses in poultry production. Full article

Avian influenza A virus (AIV) and Newcastle disease virus (NDV) are among the most damaging pathogens of poultry, and AIV also carries zoonotic potential that places it within the One Health agenda. The two infections frequently produce overlapping clinical signs, so a method that detects both in a single reaction has clear practical value for surveillance. We developed a duplex one-step real-time reverse-transcription PCR (RT-qPCR) that detects AIV in the FAM channel and NDV in the ROX channel. Primers and hydrolysis probes were designed against conserved regions of the AIV nucleoprotein and matrix genes and the NDV matrix, phosphoprotein and nucleoprotein genes, after alignment of 500 sequences per virus retrieved from GenBank from different geographic regions. Annealing temperature, primer and probe concentrations were optimised, and recombinant plasmids carrying the target fragments served both as positive controls and as quantitative standards. The assay was specific for AIV and NDV, with no cross-reactivity with infectious bronchitis virus and no signal crossover between channels; its analytical sensitivity reached approximately 10–20 RNA copies per reaction. Qualitative results on reference isolates were fully concordant with two commercial veterinary kits (Cohen’s κ = 1.00), and the assay passed inter-laboratory commission testing. This study establishes the design and core analytical performance of a laboratory-developed duplex assay; expanded in silico inclusivity on contemporary sequences, a broader specificity panel, inhibitor-containing matrices, an internal amplification control, and prospective clinical validation are required before routine surveillance deployment. Full article

Background/Objectives: Waning immunity after primary COVID-19 vaccination supports evaluation of additional doses. HXP-GPOVac is an egg-based, inactivated Newcastle disease virus (NDV)-vectored vaccine expressing a prefusion-stabilized SARS-CoV-2 HexaPro spike antigen. We evaluated the safety, tolerability, and immunogenicity of a single additional 10 µg dose of HXP-GPOVac administered to adults previously primed with two doses of either HXP-GPOVac or BNT162b2. Methods: Study GPO NDV-HXP-S 203 was an open-label phase II extension enrolling adults (18–75 years) who previously completed a two-dose primary series in Study 202 with either HXP-GPOVac or BNT162b2 (Pfizer–BioNTech; Comirnaty). All participants received a single additional 10 µg intramuscular dose of HXP-GPOVac ≥ 6 months after their second primary dose. Solicited local/systemic adverse events (AEs) were recorded for 7 days, unsolicited AEs through Day 28, and serious AEs (SAEs) and adverse events of special interest (AESIs) throughout follow-up. Neutralizing antibody titers (pseudovirus 50% neutralization titer, NT₅₀) and anti-spike IgG (BAU/mL) were assessed pre-dose (Day 1) and post-vaccination through 12 months; a predefined subset underwent IFN-γ and IL-5 ELISpot. SARS-CoV-2 infection during follow-up was assessed using anti-nucleocapsid (anti-N) IgG. Symptomatic COVID-19 was identified through symptom-reported, symptom-triggered RT-PCR testing; sequencing was performed when feasible. Results: All 219 participants received HXP-GPOVac (167 primed with HXP-GPOVac and 52 with BNT162b2). Any solicited local reaction occurred in 22.2% (37/167) of HXP-GPOVac-primed and 26.9% (14/52) of BNT162b2-primed participants; any solicited systemic reaction occurred in 10.8% (18/167) and 13.5% (7/52), respectively. No vaccine-related unsolicited AEs or AESIs were reported. Three deaths occurred during the 12-month follow-up; one (a sudden cardiac death in an HXP-GPOVac-primed participant) was assessed by the safety medical team as possibly related to vaccination, and two were assessed as not related. Neutralizing antibody GMTs increased from 46.33 at baseline to 1569.04 at Day 15 in HXP-GPOVac-primed participants and from 77.25 to 841.34 in BNT162b2-primed participants; corresponding SCRs were 78.8% and 76.9%. Anti-spike IgG GMCs increased from 48.79 to 1480.14 BAU/mL and from 194.48 to 1547.88 BAU/mL, respectively. Responses declined over time but remained above baseline through 12 months. In the cellular immunity subset, post-vaccination IFN-γ responses increased, with comparatively modest IL-5 responses and no pattern suggestive of Th2 predominance. Conclusions: A single additional dose of HXP-GPOVac administered ≥6 months after primary vaccination with HXP-GPOVac or BNT162b2 was generally well tolerated and elicited robust recall humoral responses, with supportive findings of cellular immunity. Trial registration: Thai Clinical Trials Registry, TCTR20230213001. Full article

Newcastle disease (ND), a highly contagious poultry disease caused by NDV, primarily triggers gastrointestinal lesions. Microecological preparations, novel biological additives for restoring intestinal microbiota diversity, improve nutrient absorption, reinforce intestinal barrier function, and modulate host immune responses. This study investigated the effects of a compound microecological preparation on intestinal pathogenicity in chickens infected with genotype VII Newcastle disease virus (NDV). SPF chickens were allocated to four dietary groups with or without a compound microecological preparation, followed by NDV challenge in two groups. Survival, intestinal morphology, and transcriptomic responses were assessed. The results showed that chickens fed with the compound microecological preparation exhibited improved intestinal development. Following NDV infection, these chickens displayed milder cecal lesions without obvious hemorrhage and a higher survival rate. Furthermore, differential gene transcription analysis revealed that supplementation with the compound microecological preparation regulated the expression of genes associated with metabolic processes, biological regulation, immune response, and growth and development pathways, consistent with the clinical findings. In conclusion, we demonstrated that the compound microecological preparation promotes intestinal development in chickens, delays disease progression following NDV infection, and alleviates pathological damage. Transcriptomic analysis further indicated that the preparation enhances intestinal mucosal immunity by stimulating IgA production and strengthening the immune response against genotype VII NDV. These findings provide a scientific basis for the application of compound microecological preparations in regulating the intestinal immune system and in the prevention of Newcastle disease. Full article

The search for novel antiviral agents against Newcastle disease virus (NDV) remains a priority in industrial poultry farming due to the virus’s high contagiousness and associated economic losses, prompting evaluation of polyene macrolides as potential therapeutic candidates. We employed a comprehensive approach combining computational modeling (molecular docking and dynamics simulation) and laboratory experiments to investigate the antiviral potential of natamycin, nystatin, and filipin complex against three NDV strains. Molecular docking analysis indicated binding sites for macrolides within the hydrophobic regions of surface glycoproteins HN and F, with binding energies ranging from −6.5 to −10.5 kcal/mol, while 50 ns molecular dynamics simulation confirmed complex stability. Laboratory testing using fluorescence-based neuraminidase assays demonstrated dose-dependent inhibitory activity with IC₅₀ values of 0.0043 ± 0.0015 mg/mL for filipin complex, 0.0117 ± 0.0029 mg/mL for nystatin, and 0.0220 ± 0.0138 mg/mL for natamycin, with similar ranking observed for fusion inhibition (EC₅₀ values of 0.00053 ± 0.00039, 0.00545 ± 0.00560, and 0.01196 ± 0.00965 mg/mL, respectively). While filipin complex exhibited the highest antiviral activity, its significant cytotoxicity limits therapeutic application, whereas natamycin demonstrated a favorable safety profile consistent with its GRAS status. These findings indicate that natamycin exhibits a favorable safety-to-efficacy profile in vitro, warranting further in vivo investigation to clarify its mechanism of action and establish practical application protocols for NDV control in poultry. Full article

To address the lack of a commercially available vaccine for goose circovirus (GoCV), we developed and evaluated a prokaryotically expressed subunit vaccine targeting the viral capsid (Cap) protein. A truncated Cap protein (GoCV-ΔCap) was expressed in Escherichia coli (E. coli) and formulated with aluminum hydroxide as a subunit vaccine (GoCVsubvac). Goslings were primed intramuscularly (i.m.) with high (75 µg) or low (15 µg) doses GoCVsubvac, followed by a boost 14 days later. At 14 days post-boost, goslings were challenged with GoCV and were administered a bivalent inactivated vaccine against Newcastle disease virus (NDV) and H9-subtype Avian influenza virus (AIV). Using our established gosling pathogenicity model, vaccine efficacy was evaluated via body weight, lesions, viral load, antibody titers, cytokine responses, and interference with NDV/AIV immunity. Results demonstrated that the GoCV-ΔCap vaccine, especially the high-dose formulation, provided effective immunoprotection. It elicited robust humoral and cellular immune responses, reduced lymphoid pathology, and decreased the viral detection rate in lymphoid tissues from 100% (5/5) in infected controls to 40% (2/5). Importantly, it alleviated GoCV-induced immunosuppression and preserved the immunogenicity of co-administered vaccines. This novel subunit vaccine is a promising candidate for controlling GoCV disease (GoCVD). Full article

Background/Objectives: HXP-GPOVac is a locally produced, inactivated Newcastle disease virus-based (NDV-HXP-S) COVID-19 vaccine manufactured in Thailand. This phase II trial compared its safety and immunogenicity with the mRNA vaccine BNT162b2 in adults aged 18–75 years. Methods: In this randomized, double-blind, active-controlled trial registered with the Thai Clinical Trials Registry (TCTR20220819003), 300 participants were assigned 3:1 to receive HXP-GPOVac or BNT162b2 on Days 1 and 29. Solicited adverse events (AEs) were recorded for 7 days after each dose, AEs were summarized through 28 days after each dose, and serious adverse events (SAEs), medically attended AEs (MAAEs), and adverse events of special interest (AESIs) were collected through Day 197. Humoral immunogenicity was assessed by pseudovirus 50% neutralization titers (NT₅₀) and anti-spike IgG concentrations at baseline, Day 29, Day 43, and Day 197. Seroconversion was defined as a ≥4-fold increase from baseline. A predefined subset underwent interferon-γ (IFN-γ) and interleukin-5 (IL-5) ELISpot assays to assess cell-mediated immune responses. The primary immunogenicity analysis assessed non-inferiority of HXP-GPOVac compared with BNT162b2 based on the NT₅₀ geometric mean titer ratio, with a prespecified non-inferiority margin of 0.5. Results: Solicited AEs were predominantly mild and occurred more frequently after the first dose in both groups; one or more solicited local or systemic AEs were reported by 23.7% (95% CI: 18.3–29.8) of HXP-GPOVac recipients and 44.7% (95% CI: 33.3–56.6) of BNT162b2 recipients after the first dose. AEs through 28 days after vaccination and SAEs were uncommon; MAAEs occurred in 17.0% of HXP-GPOVac recipients and 22.4% of BNT162b2 recipients, and none were considered related to vaccination. In the HXP-GPOVac group, NT₅₀ geometric mean titers increased from 5.6 at baseline to 65.5 at Day 29 and 505 at Day 43, declining to 63.6 at Day 197. Anti-spike IgG geometric mean concentrations rose from 7.5 BAU/mL at baseline to 102.7 BAU/mL at Day 29 and 514.6 BAU/mL at Day 43, decreasing to 61.0 BAU/mL at Day 197. BNT162b2 induced higher antibody levels at all time points. The NT₅₀ GMT ratio (HXP-GPOVac/BNT162b2) at Day 43 was 0.51 (95% CI: 0.39–0.67); the lower bound did not exceed the prespecified non-inferiority margin of 0.5, and non-inferiority was not established. Seroconversion rates at Day 43 were 97.6% for HXP-GPOVac and 97.1% for BNT162b2 (neutralizing antibody) and 98.6% and 97.1%, respectively (anti-spike IgG). ELISpot analyses demonstrated increased IFN-γ responses after the second dose without evidence of Th2-dominant skewing. Conclusions: HXP-GPOVac was well tolerated and induced substantial humoral and cellular immune responses, with high seroconversion rates and balanced T-cell polarization. Although absolute antibody levels were lower than those induced by BNT162b2 and the prespecified non-inferiority criterion was not met, these findings support continued evaluation of the inactivated NDV-HXP-S vaccine platform. Full article

Avian orthoavulavirus 1 (AOaV-1), commonly known as Newcastle disease virus (NDV), despite widespread vaccination, remains a significant threat to domestic chickens (Gallus gallus domesticus). Currently available live-attenuated NDV vaccines are derived from genotypes I and II lentogenic strains, whereas genetically divergent velogenic strains predominantly caused recent NDV outbreaks. This study examined the extent of genotypic divergence between NDV vaccine strains and field strains using phylogenetic and multivariate analyses of two major antigenic and virulence-associated genes: fusion (F) and haemagglutinin-neuraminidase (HN). A total of 121 full-length NDV-F and 81 NDV-HN gene sequences, representing reported NDV genotypes, were downloaded from GenBank and analysed using maximum-likelihood (ML) phylogenetic trees and principal coordinates analysis (PCoA). The phylogeny revealed genotype-specific clustering for both genes, consistent with current NDV classification. NDV vaccine strains belonging to genotypes I and II formed distinct clades, segregated from the majority of NDV field strains, including velogenic or virulent NDV genotypes. The principal coordinates analysis of both genes further confirmed the phylogenetic clustering of NDV genotypes, indicating increased genomic heterogeneity. These findings suggest genetic segregation of divergent velogenic or virulent genotypes from lentogenic NDV vaccines, requiring biological experiments for determining their efficacy against field strains. This study highlights the importance of molecular surveillance of NDV to monitor its genomic diversity, which is crucial for developing strategies to combat NDV outbreaks in domestic chickens. This study provides an updated, NDV-glycoprotein-gene-based comparative analysis across reported NDV genotypes using phylogenetic and multivariate approaches. Full article

Vaccination remains the core strategy for the prevention and control of Newcastle disease (ND). The inherent thermosensitivity of traditional Newcastle disease virus (NDV) vaccines imposes major limitations on their transportation, storage, and field application. To address these challenges, a novel liquid, thermostable, live ND vaccine was developed in the present study. Firstly, Tris/HCl buffer at near-neutral pH was identified as the optimal basic buffer system. On this basis, further screening and formulation optimization of vaccine stabilizers were conducted, and NDV strains with excellent thermal stability were used to verify the stability-conferring properties of the developed stabilizer. The results showed that the formulation composed of 0.5% gelatin, 4% trehalose, 0.1% L-glutamic acid, and 0.5% thiourea was confirmed as the optimal stabilizer for ND liquid vaccines. This formulation maintained the stable storage of the tested NDV for 12 months at 4 °C and exhibited promising stability for 30 days at 25 °C, marking a significant advancement toward development thermostable NDV vaccines that are independent of a continuous cold chain. More importantly, the liquid vaccine stored at 4 °C for 12 months still induced high levels of NDV-specific antibodies in specific pathogen-free chicks and provided 100% protective efficacy against challenge with virulent NDV. In conclusion, the liquid vaccine stabilizer developed in this study not only significantly enhanced the thermostability of the vaccine but also effectively maintained its immunogenicity, thereby providing an important theoretical basis for the research and development of liquid ND vaccines. Full article

Background/Objectives: Combined vaccination against avian influenza (A/H5N3, A/H7N7) and Newcastle disease is of practical interest for reducing handling during immunization and for achieving timely protection in poultry. The aim of this study was to evaluate an inactivated combined (associated) vaccine containing antigenic variants of avian influenza viruses A/H5N3 and A/H7N7 and Newcastle disease virus (NDV). The vaccine is protected by Patent No. 87417. Methods: Viruses with initial reproductive titers of 10^7.5 EID₅₀/mL were inactivated with formaldehyde and formulated as mono-, bi-, or trivalent combinations. Antigens were adsorbed onto aluminum hydroxide gel (1.5%). Immunogenicity was assessed in chicks naïve to avian influenza and Newcastle disease using hemagglutination inhibition (HI) antibody kinetics. Vaccination was performed twice with a 21-day interval. Group administration via drinking water (5 mL/bird) was compared with parenteral administration (1.0 mL/bird). Protective efficacy was evaluated by challenge with virulent viruses at day 30. Sterility and safety/reactogenicity were assessed, and immunobiological performance was additionally evaluated under household farm conditions (337 chickens). Results: Following vaccination, protective immunity was observed starting from day 14. HI titers peaked by day 30 (7.6–7.8 log₂ for A/H5N3 and A/H7N7; 9.2 log₂ for NDV) and remained detectable through 180 days (4.3–4.7 log₂ for avian influenza antigens; 5.1 log₂ for NDV). Group administration via drinking water produced antibody kinetics comparable to parenteral vaccination, and vaccinated birds were resistant to challenge at day 30. The tested batches met sterility requirements and showed acceptable safety/reactogenicity in laboratory studies. Conclusions: The developed inactivated combined vaccine induced HI antibodies and protective immunity against avian influenza (A/H5N3, A/H7N7) and Newcastle disease. The formulation concept supports flexible antigen combinations and enables group administration via drinking water, which may reduce handling compared with separate vaccinations. Full article

Background/Objectives: We previously developed a low-cost vaccine based on Newcastle disease virus expressing a stabilized pre-fusion spike of SARS-CoV-2 (NDV-HXP-S), which has shown safety and immunogenicity in pre-clinical and clinical studies. Due to the emergence of immune-evasive variants and the need to protect vulnerable populations, we evaluated adjuvanted NDV-HXP-S vaccine formulations to enhance and broaden immune responses. Methods: We tested the antibody responses of mice immunized intramuscularly with an inactivated NDV-HXP-S vaccine adjuvanted with AddaVax, AddaS03, Alhydrogel adjuvant 2% (Alum), or Quil-A. Results: AddaVax, AddaS03, and Alum induced the strongest IgG responses to the ancestral spike protein, boosted cross-reactive antibodies against both S1 and S2 subunits, and elicited high cross-neutralizing titers. Conclusions: The present results highlight the critical role of adjuvant selection in shaping both the magnitude and breadth of the immune response induced by the NDV-HXP-S vaccine. AddaVax, AddaS03, and Alum stand out as promising candidates to enhance NDV-HXP-S vaccine immunogenicity, with potential applications in booster strategies against SARS-CoV-2, enabling dose sparing and reducing costs. Full article

Chicken respiratory diseases represent multifactorial conditions resulting from viral, bacterial, mycoplasmal pathogens, and environmental factors, causing significant economic losses within the poultry industry. A specific respiratory disease characterized by breathing difficulties and bronchial occlusion due to caseous exudates is termed chicken bronchial obstruction. However, the absence of rapid, precise, and highly sensitive diagnostic methods for differentiation of primary respiratory disease pathogens or opportunistic pathogens, including avian influenza virus (AIV), infectious bronchitis virus (IBV), Pseudomonas aeruginosa (P. aeruginosa), and Escherichia coli (E. coli), constitutes a substantial challenge. This study developed a quadruplex droplet digital polymerase chain reaction (ddPCR) method that targeted the HA gene of H9 subtype AIV, the M gene of IBV, the Pal gene of P. aeruginosa, and the UidA gene of E. coli. Following the optimization of annealing temperature, sensitivity, and repeatability, the minimum detectable concentrations were determined as 3.02 copies/μL for the HA gene of H9 subtype AIV, 3.08 copies/μL for the M gene of IBV, 3.19 copies/μL for the Pal gene of P. aeruginosa, 3.39 copies/μL for the UidA gene of E. coli. No cross-reactivity was observed with Newcastle disease virus (NDV), H5 subtype AIV, H7 subtype AIV, fowl adenovirus serotype 4 (FAdV-4), infectious laryngotracheitis virus (ILTV), Avibacterium paragallinarum, Streptococcus, Salmonella, Pasteurella multocida, and Staphylococcus aureus. The method demonstrated excellent repeatability, with a coefficient of variation (CV) below 9%. The 185 clinical samples collected in Hebei Province China are tested by both quadruplex ddPCR and quadruplex qPCR method and the results compared. The sensitivity of the quadruplex ddPCR method (57.30%; 106/185) slightly exceeded that of the quadruplex qPCR method (49.73%; 92/185). Pathogens or opportunistic pathogens positive rates obtained via the quadruplex ddPCR were 40.00% for H9 subtype AIV, 33.51% for IBV, 24.32% for P. aeruginosa, and 27.57% for E. coli. In comparison, the positive rates of H9 subtypes AIV, IBV, P. aeruginosa, and E. coli from the quadruplex qPCR were 36.22%, 30.81%, 21.62%, and 24.32%, respectively. The coincidence rates between the two methods were 96.22% for H9 AIV, 97.30% for IBV, 97.30% for P. aeruginosa, and 96.76% for E. coli. These results demonstrated that the quadruplex ddPCR method represented a highly sensitive, specific, and rapid technique for identifying H9 subtype AIV, IBV, P. aeruginosa, and E. coli. Full article

Species of the orders Charadriiformes and Anseriformes serve as the primary long-distance disseminators of various avulaviruses. The most economically significant among them is Newcastle disease virus (NDV), or Avian orthoavulavirus 1 (AOAV-1), which causes diseases of varying severity in both domestic and wild birds. Other avulaviruses have been studied to a much lesser extent, and for most of them, only single isolates are known, which does not allow a comprehensive assessment of their potential threat. To evaluate the biological diversity and potential risks posed by avian paramyxoviruses spread by wild waterfowl during autumn migration, fecal samples from mallards (Anas platyrhynchos) (n = 3604) were collected at water bodies in Moscow and the Moscow Region between 2008 and 2024. From these samples, AOAV-1 (n = 4) and Avian paraavulavirus 4 (APMV-4) (n = 9) were isolated and partially sequenced. Phylogenetic analysis revealed that all AOAV-1 isolates belong to genotype 1 of class II, while all APMV-4 isolates belong to the Eurasian subgenotype of genotype 1. Analysis of the F protein cleavage site motif indicated conformity with the consensus sequences characteristic of lentogenic and non-pathogenic avian paramyxoviruses in all isolates. Full article