Goose Parvovirus

Overview and Taxonomy of Goose Parvovirus

Goose parvovirus (GPV) is the etiologic agent of Derzsy’s disease, an acute, highly contagious septicemia of goslings and Muscovy ducklings that has historically caused catastrophic losses to the global waterfowl industry. The virus was first recognized in the early 1960s, and for decades it was considered a relatively well‑circumscribed pathogen affecting only geese and Muscovy ducks. However, the emergence since 2015 of a novel genetic variant, termed novel goose parvovirus (NGPV), which causes short‑beak and dwarfism syndrome (SBDS) in Pekin ducks, Cherry Valley ducks, and mule ducks, has fundamentally reshaped our understanding of the virus’s host range, genetic plasticity, and economic impact. The disease is now recognized by the World Organisation for Animal Health (WOAH) as a significant transboundary threat to waterfowl production, and surveillance efforts are coordinated through networks such as FAO’s Emergency Prevention System for Animal Health. No evidence suggests zoonotic potential; GPV remains strictly confined to avian hosts.

Taxonomic Position

Goose parvovirus is classified within the family Parvoviridae, subfamily Parvovirinae, genus Dependoparvovirus [2, 23]. This genus historically comprised adeno‑associated viruses (AAVs) that require a helper virus for productive replication, but GPV and other avian dependoparvoviruses are considered autonomously replicating in permissive cells, despite their taxonomic placement. The type species is Dependoparvovirus anser, which includes GPV along with the closely related Muscovy duck parvovirus (MDPV) and the recombinant forms that have recently emerged. The virion is non‑enveloped, icosahedral, approximately 20–28 nm in diameter, and contains a linear, single‑stranded DNA genome of about 5.0–5.1 kb [3, 6]. The capsid is composed of three overlapping viral proteins, VP1, VP2, and VP3, generated from alternatively spliced transcripts and sharing a common C‑terminal region. Cryo‑electron microscopy at 2.4 Å resolution has revealed that the GPV capsid displays the canonical parvovirus surface features: depressions at the twofold axes, protrusions at the threefold axes, and channels at the fivefold axes [2]. These structural motifs are crucial for receptor binding, antigenicity, and host tropism.

Genetic Lineages and Nomenclature

Extensive genomic analyses have resolved the genetic diversity of waterfowl parvoviruses into three major clades:

1. Classical goose parvovirus (C‑GPV) – the original strains that cause Derzsy’s disease in geese and Muscovy ducks. These isolates share >95% nucleotide identity among themselves and are typically highly virulent in goslings, with mortality approaching 100% in the first month of life [4, 8, 37]. Examples include the reference strain B (European) and the Chinese vaccine strain SYG61v.

2. Muscovy duck parvovirus (MDPV) – a distinct lineage that primarily infects Muscovy ducklings and causes similar clinical signs. Although historically considered a separate serotype, MDPV and GPV share >80% nucleotide identity and undergo frequent recombination, blurring the boundaries between them [13, 39].

3. Novel goose parvovirus (NGPV) – also referred to as duck‑origin goose parvovirus, novel GPV, or variant GPV. NGPV is the causative agent of SBDS in Pekin, Cherry Valley, and mule ducks, and was first reported in China in 2015, subsequently spreading to Europe (Poland, Egypt, Turkey) and Southeast Asia [1, 18, 27, 30]. NGPV shares only 90–97% identity with C‑GPV, yet phylogenetic analyses consistently place it within the GPV clade, not as a separate species [14, 24, 35]. Key molecular hallmarks of NGPV include 12–16 conserved amino acid substitutions in the VP1 capsid protein and 12 substitutions in the non‑structural Rep1 protein, compared to C‑GPV [24, 29]. Additionally, NGPV strains often carry deletions in the inverted terminal repeats (ITRs) that may modulate replication and pathogenicity [9, 35].

The term mutated GPV (MGPV) has also been used in some studies to distinguish highly pathogenic strains that retain the classical tropism for goslings but possess distinct genetic markers [4]. In practice, MGPV overlaps with C‑GPV, and the nomenclature remains fluid. Importantly, the World Organisation for Animal Health (WOAH) does not currently differentiate between these lineages for notification purposes, but the Food and Agriculture Organization (FAO) has flagged NGPV as an emerging disease of ducks requiring active surveillance.

Recombinant and Chimeric Strains

The co‑circulation of C‑GPV, MDPV, and NGPV in overlapping geographic regions has driven extensive recombination. Natural recombinants have been identified with one, two, or even three crossover regions. For example, Muscovy duck‑origin goose parvovirus (MDGPV) strains result from recombination between MDPV (as the major parent) and GPV (minor parent), with breakpoints in the P9 promoter region and the VP3 gene [39]. More recently, a three‑region recombinant (2022FZ) was reported, containing exchanges in the P9‑NS region, the NS2 gene, and the VP3 gene [13]. Another isolate, HLJ2023, was shown to be a potential recombinant between a NGPV strain and a MDPV‑like strain, and its virulence for goslings was exceptionally high (100% mortality within 36 h) [3]. These findings underscore that recombination is a major driver of GPV evolution, generating novel variants that can shift host tropism and pathogenicity. The use of live attenuated vaccines (e.g., SYG61v) has been implicated in some recombination events, raising concerns about vaccine‑derived strains contributing to genetic diversity [33, 39].

Host Range and Cross‑Species Transmission

Historically, C‑GPV was considered restricted to geese and Muscovy ducks, with Pekin ducks and other Anas species showing only subclinical infection or limited replication in immune organs [36]. The emergence of NGPV shattered this paradigm. NGPV is now known to cause overt SBDS in Pekin, Cherry Valley, and mule ducks, while remaining less pathogenic in geese [4, 20]. Conversely, C‑GPV (MGPV) is highly pathogenic to goslings but induces only mild signs in ducks [4]. This reciprocal host tropism implies that the genetic differences between C‑GPV and NGPV, particularly in the VP1 and Rep proteins, determine cellular receptor usage and species‑specific pathogenesis. The VP3 protein, which contains the major antigenic epitopes and is involved in receptor binding, shows distinct amino acid residues in NGPV that align with European SBDS strains from the 1970s, suggesting that NGPV may have originated from a European lineage that later spread to Asia [24, 30].

Host range is not absolute: experimental infection of Cherry Valley ducklings with C‑GPV can produce mild lesions in immune organs, and NGPV has been isolated from geese with feathering disorders [31, 36]. Moreover, co‑infections with duck circovirus (DuCV) or goose circovirus (GoCV) dramatically exacerbate the severity of SBDS and Derzsy’s disease, indicating that immunosuppressive co‑pathogens can broaden the clinical spectrum [16, 22, 34]. The detection of GPV in 100% of tested commercial goose flocks in Poland, often in mixed infections, highlights the ubiquity of the virus even in vaccinated populations [1].

Genomic Organization and Functional Elements

The GPV genome is typical of dependoparvoviruses: two large open reading frames (ORFs) encoding the non‑structural proteins (Rep1, Rep2) and the structural proteins (VP1, VP2, VP3), flanked by complex inverted terminal repeats (ITRs) of approximately 400–500 nt that form T‑shaped hairpins essential for replication [6, 9]. The ITRs contain cis‑acting elements such as E‑box motifs (CACATG) that influence packaging and transcriptional regulation. Deletion of one E‑box in the left ITR of an MDGPV infectious clone reduced viral replication and attenuated virulence in Muscovy duck embryos [41]. The 5′ end of the genome also harbors the P9 promoter driving expression of the Rep proteins, while the P41 promoter controls capsid gene expression.

The non‑structural protein NS1 (Rep1) is a multifunctional helicase and endonuclease required for rolling‑circle replication. NS1 induces apoptosis in goose embryo fibroblasts via the AIF‑mitochondrial pathway [25]. It also contributes to viral pathogenicity: chimeric viruses exchanging the Rep1 gene between NGPV and C‑GPV showed that the NGPV Rep1 is associated with the ability to cause SBDS in ducks [20]. The NS2 protein, though less studied, appears to be involved in host‑dependent replication and is a recombination hotspot [13].

The structural proteins are derived from a common precursor by alternative splicing and proteolytic cleavage. VP1 contains a unique N‑terminal region harboring a phospholipase A2 (PLA2) domain and a nuclear localization signal (NLS) spanning residues 160YPVVKKPKLTEE171; the lysines at positions 164, 165, and 167 are critical for nuclear import of the VP1 protein [38, 42]. VP2 is the major capsid component, and VP3 is generated by trypsin‑like cleavage of VP2 and forms the core of the capsid. The VP3 gene is highly conserved and is the target of most diagnostic assays, including CRISPR‑Cas12a, RPA, and LAMP methods [5, 12, 19]. VP1 of NGPV has been shown to suppress type I interferon induction by targeting IRF7, thereby facilitating immune evasion [15].

Phylogeography and Global Spread

Phylogeographic analyses indicate that China is a major epicenter of GPV diversity and a source of export [18]. From China, NGPV appears to have spread to Europe via trade in live ducks or contaminated poultry products; the Polish NGPV isolates from 2019 share 98–99% identity with Chinese strains [1, 30]. Egyptian NGPV strains also cluster with Chinese isolates, suggesting a common origin [10, 17, 27]. In Turkey, both classical and novel variants circulate, with field strains clustering in the European group 2 lineage [26, 28]. The detection of NGPV in skin and feather follicles of duck carcasses at slaughterhouses indicates that the virus can persist in feather burrs and may be mechanically carried over long distances by transport vehicles [7, 11]. Contaminated transport vehicles have been proposed as a vector for GPV introduction into naïve flocks despite vaccination [11].

The WOAH and FAO have emphasized the need for standardized molecular surveillance to track the international movement of GPV variants. The development of differential diagnostic assays, such as TaqMan qPCR that distinguishes C‑GPV from NGPV based on host specificity, has facilitated this effort [32]. Nevertheless, the rapid evolution of GPV through recombination and point mutation necessitates continuous genomic monitoring.

Economic Significance

Goose parvovirus, in all its guises, remains a scourge of waterfowl production. Derzsy’s disease can cause mortality rates exceeding 90% in goslings, with survivors suffering growth retardation and dwarfism that render them unmarketable [8, 23]. SBDS, though associated with lower mortality (4–20%), leads to severe growth depression, beak deformities, and reduced feed conversion, causing substantial economic losses in the duck industry [4, 30, 40]. In Poland, GPV was detected in all surveyed flocks, and co‑infections with circovirus and polyomavirus compounded production losses [1]. The virus also induces angel wing syndrome in Muscovy ducks, further impairing mobility and carcass quality [21]. Given its global distribution and the lack of effective cross‑protective vaccines between lineages, GPV remains a high‑priority pathogen for veterinary authorities and international trade bodies.

Molecular Pathogenesis of Goose Parvovirus

The molecular pathogenesis of goose parvovirus (GPV) represents a multifaceted interplay between viral structural and non-structural proteins, host cellular machinery, and immune surveillance systems. As a member of the genus Dependoparvovirus within the family Parvoviridae, GPV exhibits a single-stranded DNA genome of approximately 5,100 nucleotides, flanked by complex inverted terminal repeats (ITRs) that are critical for replication and pathogenicity [2, 9]. The pathogenic cascade initiated by GPV infection encompasses viral entry and uncoating, modulation of host cell cycle and apoptotic pathways, subversion of innate immune responses, and the induction of systemic pathological changes that manifest as Derzsy’s disease in geese and short beak and dwarfism syndrome (SBDS) in ducks. The emergence of novel GPV (NGPV) and recombinant variants has further complicated our understanding of pathogenesis, as these strains exhibit altered host tropism, tissue distribution, and virulence profiles [3, 4, 43].

Capsid Architecture and Host Cell Entry

The GPV capsid, resolved at 2.4 Å resolution through cryogenic electron microscopy (cryo-EM), displays the canonical parvoviral T=1 icosahedral symmetry characterized by surface depressions at the two-fold axes, prominent protrusions at the three-fold axes, and channel-like structures at the five-fold axes [2]. The capsid is assembled from 60 copies of the viral structural proteins VP1, VP2, and VP3, which share a common C-terminal core but differ in their N-terminal extensions. The VP1 unique region (VP1u) contains a phospholipase A2 (PLA2) domain essential for endosomal escape during viral entry, while the nuclear localization signals (NLS) within VP1 are critical for trafficking of incoming virions to the nucleus. Specifically, the basic region spanning residues 160YPVVKKPKLTEE171 in the VP1 N-terminus functions as a bona fide NLS, with lysine residues at positions 164, 165, and 167 being absolutely required for nuclear import [38, 42]. Mutagenesis studies have demonstrated that alanine substitutions at these positions (K164A, K165A, or K167A) completely abrogate VP1 nuclear localization and render the virus non-viable in goose embryo fibroblasts (GEFs), underscoring the essential role of this motif in the GPV life cycle [38].

Structural comparisons between GPV and other dependoparvoviruses, including human adeno-associated virus 2 (AAV2), AAV5, and quail AAV, have revealed unique conformations in surface-exposed variable regions (VRs) that likely dictate host specificity [2]. Notably, VR-III exhibits structural divergence between GPV and Muscovy duck parvovirus (MDPV), implicating this region in species-specific receptor engagement. Unlike many AAVs that utilize sialic acid, heparan sulfate proteoglycan, or the AAV receptor (AAVR) for attachment, GPV capsids do not bind these canonical receptors, suggesting the involvement of distinct, yet-to-be-identified cellular receptors [6]. The GPV capsid demonstrates thermal stability at physiological pH but undergoes conformational destabilization under acidic conditions, a property that may facilitate pH-dependent uncoating within endosomal compartments [2].

Viral Proteins and Their Roles in Pathogenicity

The GPV genome encodes two major non-structural proteins (NS1 and NS2) and three capsid proteins (VP1, VP2, and VP3), all derived from overlapping reading frames. NS1 is a multifunctional phosphoprotein possessing helicase, ATPase, and site-specific endonuclease activities essential for viral DNA replication. Beyond its replicative functions, NS1 is a primary determinant of pathogenicity. Comparative studies between classical GPV (cGPV) and NGPV have identified 12 conserved amino acid substitutions in the Rep1 (NS1) protein of NGPV strains relative to cGPV, which are hypothesized to contribute to the altered host tropism and attenuated virulence observed in duck hosts [24, 43]. The critical role of Rep1 in pathogenicity was definitively demonstrated using chimeric viruses: replacement of the NGPV Rep1 gene with its cGPV counterpart altered the virulence profile in duck infection models, confirming that non-structural protein determinants are key drivers of host-specific pathogenesis [20].

NS1 also functions as a potent inducer of apoptosis. In GEFs, GPV infection triggers apoptosis through an apoptosis-inducing factor (AIF)-mediated mitochondrial pathway that is both NS1-dependent and independent of classical caspase activation [25]. Mechanistically, NS1 expression leads to mitochondrial membrane depolarization, a progressive increase in reactive oxygen species (ROS) over 48 hours post-infection, and elevated cathepsin D activity. These events culminate in the translocation of AIF from the mitochondria to the nucleus, where it executes caspase-independent chromatin condensation and DNA fragmentation [25]. This AIF-mediated pathway represents a non-canonical apoptotic mechanism that may facilitate viral dissemination while evading the inflammatory responses typically associated with caspase-dependent cell death.

The VP3 capsid protein, which constitutes the core of the viral shell, is a major immunogen and target for neutralizing antibodies. However, VP3 also contributes to pathogenesis through its role in receptor binding and host range determination. Structural modeling of the VP3 protein from Vietnamese NGPV isolates (DuPV-BAFU) revealed that clusters of receptor-interacting amino acid residues are conserved among GPV strains capable of infecting ducks, whereas these residues differ from those found in MDPV and non-duck-adapted GPV strains [47]. This suggests that subtle structural alterations in VP3 dictate the ability of GPV to cross species barriers and establish infection in novel hosts.

Modulation of Host Innate Immunity

A cornerstone of GPV molecular pathogenesis is its capacity to subvert the host type I interferon (IFN) response. NGPV infection of duck embryo fibroblasts (DEFs) stimulates the mRNA expression of cyclic GMP-AMP synthase (cGAS), yet paradoxically results in weak IFN-β induction [15]. This antagonism is mediated primarily by the VP1 protein, which targets interferon regulatory factor 7 (IRF7), a master transcription factor for IFN-α/β production. Co-immunoprecipitation experiments have demonstrated that both VP1 and VP2 physically interact with IRF7, although only full-length VP1 possesses the ability to suppress IFN-β expression [15]. Truncated VP1 forms, including the VP1 unique region alone (VP1U) and VP2, fail to inhibit IFN-β, indicating that the intact VP1 protein is required for immune evasion. This blockade occurs upstream of IRF7 activation, as NGPV infection impairs the cGAS-STING signaling pathway, leading to reduced phosphorylation and nuclear translocation of IRF7 [15]. Consequently, downstream interferon-stimulated genes (ISGs) are poorly induced, creating a permissive environment for viral replication.

Transcriptomic profiling of GPV-infected goslings has revealed a distinct immunological signature characterized by elevated levels of immunosuppressive cytokines, including transforming growth factor-beta (TGF-β) and interleukin-10 (IL-10), while pro-inflammatory cytokines such as IL-4, IFN-γ, and tumor necrosis factor-alpha (TNF-α) remain unaffected [8]. This shift toward an anti-inflammatory state likely facilitates persistent viral infection and contributes to the growth retardation and dwarfism observed in surviving birds. Furthermore, GPV infection activates the IL-17 signaling pathway and upregulates genes involved in ferroptosis, including PTGS2, transferrin (TF), and ASCL1, suggesting that programmed cell death pathways beyond classical apoptosis contribute to tissue pathology [8].

Induction of Apoptosis and Cell Cycle Dysregulation

GPV infection exerts profound effects on cellular proliferation and survival. In goose embryo fibroblasts, infection leads to the identification of 285 differentially expressed genes, with significant enrichment in pathways associated with negative regulation of cell proliferation and skeletal system development [8]. Key genes involved in growth plate regulation and bone morphogenesis, including matrix metalloproteinases (MMP2, MMP9, MMP13), connective tissue growth factor (CCN3), and prostaglandin D2 synthase (PTGDS), are dysregulated, providing a molecular explanation for the skeletal abnormalities characteristic of SBDS [8].

The NS1 protein not only induces apoptosis via the AIF-mitochondrial pathway but also influences cell cycle progression. Recombinant Muscovy duck-origin GPV (MDGPV) with a deletion of the E-box motif (CACATG) within the left ITR induces G0/G1 phase arrest in infected mesenchymal stem cells, preventing entry into S phase and thereby limiting viral replication [41]. E-box elements are cis-acting transcriptional regulatory sequences that bind host transcription factors, and their deletion attenuates virus replication capacity and virulence [41]. This finding highlights the importance of ITR-encoded regulatory elements in modulating the cellular environment to favor viral propagation.

The Role of Inverted Terminal Repeats in Pathogenicity

The ITR regions of the GPV genome have emerged as critical determinants of viral pathogenicity. Using a reverse genetics system for NGPV, researchers have demonstrated that swapping the ITR sequences between different viral strains results in rescued viruses with distinct virulence profiles in duck embryos [9]. The ITRs contain palindromic sequences that form hairpin structures essential for priming DNA replication, but they also harbor binding sites for host transcription factors and viral NS1 protein. The presence of nucleotide deletions or substitutions within ITR regions has been associated with altered pathogenicity. For instance, NGPV strains isolated from ducks frequently exhibit two 14-nucleotide deletions in the ITRs compared to classical GPV, changes that may influence replication efficiency and host adaptation [35]. Additionally, the 287CACATG292 E-box deletion within the left ITR of MDGPV reduces virus replication and virulence, underscoring the functional significance of these non-coding regulatory elements [41].

Host Tropism and Tissue Pathogenesis

The molecular basis for the differential host tropism between cGPV and NGPV is multifactorial, involving capsid-receptor interactions, intracellular replication efficiency, and immune evasion capabilities. cGPV is highly pathogenic in goslings and Muscovy ducklings, causing acute hemorrhagic enteritis and hepatitis with mortality rates approaching 100% in young birds [3, 4]. In contrast, NGPV exhibits reduced pathogenicity in geese but causes SBDS in Pekin ducks, Cherry Valley ducks, and mule ducks, characterized by growth retardation, beak atrophy, tongue protrusion, and skeletal deformities [4, 20, 43]. Comparative pathogenicity studies have established that MGPV (mutated GPV) retains high virulence for goslings, while NGPV is more pathogenic for ducklings, indicating that relatively few genetic changes can dramatically alter host range [4].

Tissue tropism studies using quantitative PCR and immunohistochemistry have demonstrated that GPV disseminates widely in infected hosts, with preferential replication in lymphoid organs, intestinal epithelium, and skeletal tissues. In naturally infected ducks, the highest viral loads are detected in the bursa of Fabricius, intestine, and spleen, followed by skeletal muscle and bone marrow [10, 17, 46]. Immunohistochemical analysis has localized viral antigens to thymic lymphocytes, bursal lymphoid follicles, splenic red pulp, enterocytes, and Purkinje cells in the cerebellum [10, 17]. This broad tissue distribution explains the diverse clinical manifestations of GPV infection, including immunosuppression, enteritis, neurological signs, and locomotor disorders.

The intestinal tract serves as a major portal of entry and replication site for GPV. NGPV infection disrupts intestinal barrier integrity, leading to decreased expression of tight junction proteins and increased intestinal permeability [44, 45]. This is accompanied by dysbiosis of the cecal microbiota, characterized by reduced abundance of beneficial Firmicutes and Bacteroidota and decreased production of short-chain fatty acids (SCFAs) such as butyrate, acetate, and propionate [44, 45]. Importantly, depletion of the gut microbiota via antibiotic treatment alleviates SBDS symptoms, reduces bone resorption, and suppresses systemic inflammation, demonstrating that the intestinal microbiome plays a causal role in GPV pathogenesis [45]. The mechanistic link between microbial dysbiosis and skeletal pathology involves activation of the mucosa-associated lymphoid tissue lymphoma translocation protein 1 (Malt1) and nuclear factor kappa B (NF-κB) signaling pathways, which drive inflammatory bone loss [45].

Co-infection with immunosuppressive viruses, particularly duck circovirus (DuCV), significantly exacerbates GPV pathogenesis. Epidemiological studies have revealed that co-infection rates can exceed 70% in field cases of SBDS [30, 48]. Experimental co-infection models demonstrate synergistic effects: co-infected ducks exhibit more severe growth retardation, more pronounced immunosuppression (evidenced by reduced lymphocyte counts and impaired antibody responses), higher viral loads in tissues and cloacal swabs, and more extensive histopathological lesions compared to singly infected birds [16, 22, 49]. The temporal dynamics of co-infection are notable: in the early stages, DuCV may suppress host immune responses, facilitating GPV replication; in later stages, both viruses replicate to higher titers, and their distribution extends to additional tissues including the liver, kidney, and bursa of Fabricius [49]. This synergism likely explains why SBDS is difficult to reproduce with NGPV alone in experimental settings but occurs naturally with high frequency in co-infected flocks [50].

Genetic Recombination as a Driver of Pathogenicity

Recombination between GPV, MDPV, and NGPV strains is a major mechanism driving the emergence of novel variants with altered pathogenic properties. The single-stranded DNA genome of parvoviruses, combined with the error-prone nature of host DNA polymerases and the ability of multiple strains to co-infect the same host, creates conditions favorable for recombination. Phylogenetic and recombination analyses have identified numerous recombination events, with breakpoints frequently located in the P9 promoter region, NS gene, and VP3 gene [18, 35, 39]. A particularly striking example is the emergence of MDGPV, a natural recombinant between MDPV and GPV. Strain 2022FZ was found to contain three distinct recombination regions: one in the P9 promoter-NS region (nt 425–612), one in NS2 (nt 1,483–1,824), and one in VP3 (nt 3,124–4,248), with GPV sequences replacing the corresponding MDPV regions [13]. This triple-recombinant strain exhibited reduced pathogenicity compared to double-recombinant MDGPV strains, suggesting that the precise combination of parental sequences influences virulence [13].

Recombination is not limited to wild-type strains; vaccine strains have also been implicated. The GPV vaccine strain SYG61v has been identified as a minor parent in recombination events with MDPV, leading to the generation of pathogenic field strains [39]. Similarly, the attenuated GPV vaccine strain 82-0321v was found to serve as a major parent in recombination with a wild GPV strain, producing the NGPV strain SDLY1602 [35]. These findings raise concerns about the safety of live attenuated vaccines and their potential to contribute to the emergence of novel pathogenic variants through recombination with circulating field strains.

The World Organisation for Animal Health (WOAH) classifies GPV as a notifiable pathogen due to its significant economic impact on waterfowl production systems globally. The emergence of NGPV and its rapid spread across Asia, Europe, and Africa underscores the need for enhanced surveillance and molecular characterization of circulating strains to monitor the ongoing evolution of pathogenicity determinants [18, 30]. The Food and Agriculture Organization of the United Nations (FAO) has similarly emphasized the threat posed by emerging waterfowl parvoviruses to food security and rural livelihoods in regions where duck and goose farming are integral to agricultural economies.

Epidemiology of Goose Parvovirus Infections

Global Distribution and Prevalence

Goose parvovirus (GPV) infections represent a formidable and enduring challenge to the global waterfowl industry, exhibiting a cosmopolitan distribution across Asia, Europe, and Africa. The causative agent of Derzsy’s disease, GPV has long been recognized as a pathogen of paramount economic significance, with contemporary epidemiological surveillance revealing a far more complex and dynamic landscape than previously appreciated. The virus is not merely a pathogen of geese; its host range has expanded dramatically, driven by genetic recombination and adaptive evolution, leading to the emergence of novel variants capable of infecting multiple duck species and causing distinct clinical syndromes.

Epidemiological investigations have documented the pervasive nature of GPV infections in commercial settings. A comprehensive two-year observational study in Poland, a European leader in goose production, detected GPV genetic material in 100% of the 27 monitored commercial flocks, with flock sizes ranging from 3,000 to 13,000 birds [1]. This universal prevalence underscores the endemicity of the virus and the inadequacy of current biosecurity and vaccination strategies to eliminate circulation. The situation in China, the epicenter of waterfowl parvovirus evolution, is equally concerning. Surveillance in duck populations has revealed detection rates for GPV reaching 17.93% (71/396 clinical specimens) in samples collected from duck farms between June 2022 and July 2023 [52]. In Muscovy ducks from Fujian Province, a multiplex PCR survey identified GPV positivity rates of 73.53% (75/102) in clinically diseased flocks [53]. These data points illustrate that GPV is not a sporadic threat but a persistent, high-prevalence pathogen circulating extensively within commercial waterfowl populations across disparate geographic regions.

Emergence and Expansion of Novel Goose Parvovirus (NGPV)

Perhaps the most significant epidemiological development in recent decades has been the emergence and global spread of novel goose parvovirus (NGPV). Since its initial recognition in Cherry Valley duck flocks in China in 2015, NGPV has rapidly become the dominant parvovirus lineage in duck populations, causing short beak and dwarfism syndrome (SBDS) [24, 35]. The virus has demonstrated a remarkable capacity for cross-species transmission and geographic expansion. Phylogenetic analyses have confirmed that Chinese NGPV isolates cluster closely with European SBDS-related strains, suggesting a shared common ancestor and pointing to international trade in live birds or contaminated poultry products as a major conduit for viral dissemination [18, 24, 30].

The epidemiological trajectory of NGPV is well-documented. Following the initial Chinese outbreaks, the virus was subsequently detected in Poland in 2019, representing the first documented NGPV outbreak in Pekin ducks in Europe [30]. In these Polish flocks, morbidity ranged from 15% to 40%, with mortality rates of 4%–6%, and the complete coding regions of the Polish isolates shared 98.57%–99.28% nucleotide identity with Chinese NGPV strains [30]. Shortly thereafter, in 2020, NGPV was identified in Egypt, where it caused morbidity rates as high as 70% in mule and Pekin duck farms [27]. More recently, NGPV has been confirmed in duck flocks in Vietnam [47] and in turkeys [26], illustrating a pattern of rapid, intercontinental spread that mirrors the dissemination of other emerging waterfowl pathogens. The World Organisation for Animal Health (WOAH) recognizes the threat posed by such emerging parvovirus variants, and these epidemiological patterns highlight the need for enhanced international surveillance and reporting frameworks to track the global movement of GPV and NGPV lineages.

Host Tropism and Species Susceptibility

The epidemiology of GPV infection is fundamentally shaped by host tropism, which has been profoundly altered by viral evolution. Classical GPV (cGPV) is characterized by a relatively narrow host range, primarily causing severe, acute disease in goslings and Muscovy ducklings, with mortality rates approaching 100% in young birds [4, 6]. In contrast, NGPV exhibits a distinctly different host preference. Experimental infections have demonstrated that NGPV is significantly more pathogenic to ducklings, particularly Cherry Valley and Pekin ducks, while causing less severe disease in goslings [4, 20]. This host shift is a critical epidemiological feature; cGPV and NGPV can be differentiated clinically based on host species, with cGPV infections dominating in geese and Muscovy ducks, and NGPV prevalent in other duck breeds [32].

The molecular basis for this differential host tropism is beginning to be elucidated. Comparative genomic analyses have identified specific amino acid substitutions in the VP1 capsid protein of NGPV that distinguish it from cGPV. Sixteen common amino acid substitutions have been identified in the VP1 proteins of NGPV strains compared with classical Chinese GPV strains, nine of which are identical to those found in the European GPV strain B [24]. Furthermore, structural modeling has revealed that NGPV strains possess identical clusters of receptor-interacting amino acid residues in the VP3 protein, a major determinant of viral receptor binding and host specificity [47]. These findings suggest that relatively few genetic changes were sufficient to facilitate a host species jump, enabling NGPV to exploit new ecological niches in duck populations. This capacity for host switching is a hallmark of parvovirus evolution and has direct implications for the potential emergence of future variants capable of infecting novel avian species.

Recombination and Genetic Diversity Driving Epidemics

The epidemiological landscape of GPV is continuously reshaped by extensive genetic recombination, a process that generates novel viral variants with altered pathogenic potential and host range. Recombination between cGPV, Muscovy duck parvovirus (MDPV), and NGPV is a frequent occurrence, driven by co-infections in susceptible waterfowl populations. A comprehensive phylogeographic analysis identified 11 distinct recombination events among GPV, MDPV, and NGPV, underscoring the dynamic nature of parvovirus evolution [18]. These recombination events are not merely laboratory curiosities; they have direct epidemiological consequences.

Recombinant viruses have been isolated from clinical outbreaks across China. The 2022FZ strain of Muscovy duck-origin goose parvovirus (MDGPV) was identified as a three-region recombinant, with segments originating from both GPV and MDPV in the P9 promoter-NS region, the NS2 region, and the VP3 gene [13]. Another strain, HLJ2023, isolated in Heilongjiang Province, was characterized as a potential recombinant of an NGPV strain and a Muscovy duck-hosted GPV strain, exhibiting 100% mortality in experimentally infected goslings [3]. The HuN18 strain, isolated from domestic Linwu sheldrakes, was found to be a recombinant with the NGPV strain sdlc01 as the major parent and classical GPV strains Y and SYG61v as minor parents, providing evidence that attenuated live vaccines can participate in recombination events with wild-type viruses, potentially generating new emerging variants [33]. These findings are epidemiologically alarming because recombination can restore virulence in attenuated vaccine strains, alter tissue tropism, and facilitate immune evasion, complicating control efforts. The high prevalence of co-infections, with 74.1% of flocks in one study harboring dual viral infections and 22.2% harboring triple infections, creates ideal conditions for recombination to occur [1].

Transmission Dynamics and Risk Factors

Understanding the transmission dynamics of GPV is essential for implementing effective control measures. Horizontal transmission through the fecal-oral route is the predominant mechanism, with infected birds shedding large quantities of virus in feces, contaminating feed, water, and the environment. The virus is remarkably stable and can persist in organic matter and on fomites. A recent study has highlighted the role of transport vehicles as vectors for GPV transmission, demonstrating that insufficiently disinfected vehicles used to transport geese from hatcheries to farms and from farms to slaughterhouses can harbor traces of the virus, serving as a source of infection for naive flocks [11]. This finding has direct implications for biosecurity protocols, emphasizing the need for rigorous cleaning and disinfection of all equipment and vehicles that move between poultry premises.

Vertical transmission is a well-documented and epidemiologically critical feature of GPV infection. The virus can be transmitted from infected breeding birds to their progeny through the egg, enabling the virus to persist across generations and to spread to new geographic regions through the trade of hatching eggs or day-old chicks. Fluorescence-positive signals for NGPV have been detected in Mule duck embryos and newly hatched Mule ducklings, providing direct evidence of vertical transmission from breeding flocks to offspring [32]. Furthermore, co-infection of NGPV and duck circovirus (DuCV) has been detected in duck embryos, indicating that vertical transmission of multiple immunosuppressive viruses can occur simultaneously [50]. This mode of transmission is particularly insidious because it allows the virus to evade detection in adult birds, which may remain subclinical carriers, and to establish infection in immunologically naïve young birds before they are exposed to environmental sources.

Co-infections and Synergistic Pathogenesis

The epidemiology of GPV cannot be understood in isolation; co-infections with other viral and bacterial pathogens are the rule rather than the exception in commercial waterfowl flocks. These polymicrobial interactions significantly alter disease outcomes, exacerbating morbidity, mortality, and economic losses. The most clinically relevant co-infection is between GPV/NGPV and duck circovirus (DuCV). DuCV is a recognized immunosuppressive agent, and its co-occurrence with NGPV is extremely common. In clinical cases of SBDS in ducks, co-infection with DuCV has been observed in 85.7% of birds [30]. In another study examining feather sacs from ducks with feather shedding syndrome, the co-infection rate of NGPV and DuCV was 70.00% [48].

Experimental co-infection studies have conclusively demonstrated synergistic pathogenicity. Co-infection with DuCV and NGPV in specific-pathogen-free (SPF) ducks resulted in more severe clinical signs of short beak, dwarfism, and immunosuppression compared with mono-infection, along with more significant tissue damage and higher viral titers in organs and cloacal swabs [16, 22]. The immunosuppression induced by DuCV appears to permit enhanced NGPV replication, while NGPV-induced tissue damage may facilitate secondary bacterial infections. This synergistic relationship creates a positive feedback loop that amplifies disease severity and viral shedding, thereby increasing the force of infection within flocks and between flocks.

Beyond DuCV, GPV frequently co-infects with goose circovirus (GoCV), goose hemorrhagic polyomavirus (GHPV), goose astrovirus (GAstV), and a range of bacterial pathogens including Escherichia coli, Erysipelothrix rhusiopathiae, Gallibacterium anatis, and Salmonella Typhimurium [1, 56]. In a Polish study, rare was it that a single infectious agent had a clear, unambiguous impact on flock health; typically, mixed viral infections combined with bacterial or fungal complications led to the most severe outcomes, including elevated mortality, growth diversification, and reduced production rates [1]. This polymicrobial reality complicates epidemiological investigations and diagnostic interpretations. It also underscores the necessity of comprehensive diagnostic strategies that can simultaneously detect multiple pathogens, such as multiplex PCR and qPCR assays, which have been developed for the simultaneous detection of GPV alongside DuCV, GAstV, MDPV, and duck adenovirus 3 [52, 53, 55, 57].

Pathogenicity and Virulence Determinants

Epidemiological patterns are also shaped by the inherent pathogenicity of circulating strains, which varies markedly across lineages. Comparative pathogenicity studies have revealed that NGPV and mutated GPV (MGPV) represent the predominant epidemic lineages in China, and they exhibit distinct pathogenic profiles. MGPV induces classical gosling plague pathology in goslings, while NGPV leads to SBDS in ducklings, notably disrupting skeletal development [4]. In vitro, both lineages replicate in duck embryo fibroblasts (DEFs) and goose embryo fibroblasts (GEFs) causing cytopathic effects, but they display distinct levels of intra-embryonic replication capability [4]. Critically, MGPV is more pathogenic to goslings, while NGPV is more pathogenic to ducklings, and each virus elicits stronger antibody responses in its respective preferred host [4].

The molecular determinants of virulence are increasingly well-characterized. The inverted terminal repeat (ITR) regions of the GPV genome play a significant role in modulating viral pathogenicity. Using a reverse genetics system for NGPV, researchers rescued two strains differing exclusively in the ITR region and demonstrated distinct virulence profiles in vivo, providing direct evidence that the ITR regions are not merely structural elements but active contributors to pathogenicity [9]. Furthermore, chimeric virus studies have shown that the non-structural protein Rep1 plays a crucial role in NGPV pathogenicity [20]. Attenuation can be achieved through serial passage in cell culture, as demonstrated with the NMG21 strain, which exhibited a clear decrease in pathogenicity for ducklings after 35 passages in DEFs, with reduced tissue replication rates and lower tissue damage [51]. However, the potential for reversion to virulence and the risk of recombination with wild-type strains remain significant epidemiological concerns when using live attenuated vaccines.

Seasonal, Geographic, and Management-Related Factors

Epidemiological patterns of GPV are modulated by seasonal, geographic, and management-related factors. In China, epidemiological surveys have indicated that epidemics of GPV and other waterfowl viruses are more severe in winter and spring, when cold temperatures may enhance viral environmental stability and when intensive housing practices may facilitate transmission [59]. Significant differences in prevalence have also been observed across different age groups, with younger birds being more susceptible to severe disease [59]. In Poland, the timing of outbreaks correlates with the production cycle, with infections often peaking in young goslings during the early stages of the growing season [1].

Geographic variation in circulating genotypes is evident. The majority of GoCV genomic sequences from Polish flocks exhibited high homology to a previously characterized Polish sequence from domestic geese, while one sequence was closely related to sequences from wild birds, suggesting spillover events between wild and domestic waterfowl [1]. In China, NGPV strains isolated in Shandong and Henan provinces between 2019 and 2023 showed 96.0%–99.9% nucleotide homology with other NGPVs, indicating that no major mutations have occurred in these regions in recent years, suggesting a stable epidemiological situation despite high prevalence [14]. However, the situation in other provinces, such as Heilongjiang, has revealed highly pathogenic recombinant strains with 100% mortality in experimental infections, indicating that virulent variants continue to emerge regionally [3]. The movement of live birds, contaminated equipment, and insufficiently disinfected transport vehicles are major drivers of geographic spread, and biosecurity failures at these points represent critical opportunities for intervention [11].

Economic and Production Impact

The epidemiological significance of GPV is ultimately measured by its economic impact on the waterfowl industry. Derzsy’s disease causes mortality rates of up to 99.6% in young goslings, and even in surviving birds, infection results in stunted growth, dwarfism, and significant weight loss, leading to reduced market value and prolonged time to slaughter [8, 54]. The emergence of NGPV has added a second dimension of economic damage, with SBDS causing high morbidity (up to 70%) and substantial losses due to retarded growth, low performance, and condemnation of carcasses at processing plants [27]. The detection of infectious NGPV in the skin of duck carcasses with residual feather burrs has implications for food safety and product quality, potentially affecting consumer acceptance and trade [7].

In Poland, where goose production is the largest in Europe, GPV infection must be reported to veterinary authorities due to its serious economic and epizootic threat [58]. The costs associated with GPV include mortality, reduced growth rates, increased veterinary and diagnostic expenses, and the costs of vaccination programs. The high prevalence of co-infections complicates treatment and control, often necessitating the use of multiple vaccine types and antimicrobials to manage secondary bacterial complications. The economic burden is amplified by the persistent nature of the virus in flocks, which can lead to recurrent outbreaks in successive production cycles.

Diagnostic Approaches for Goose Parvovirus

The accurate and timely diagnosis of Goose Parvovirus (GPV) infection is paramount for implementing effective control measures, mitigating economic losses, and understanding the complex epidemiology of this pathogen, which affects geese, Muscovy ducks, and an expanding range of waterfowl hosts [1, 4, 27]. The diagnostic landscape for GPV has evolved dramatically from classical pathological assessment and virus isolation to a sophisticated arsenal of molecular, serological, and point-of-care (POC) technologies. The emergence of novel genetic variants, particularly the Novel Goose Parvovirus (NGPV) associated with short beak and dwarfism syndrome (SBDS) in ducks, and the high prevalence of co-infections with agents such as duck circovirus (DuCV), goose circovirus (GoCV), and goose hemorrhagic polyomavirus (GHPV), necessitates diagnostic approaches that are not only highly sensitive and specific but also capable of differentiating between pathotypes and detecting mixed infections [1, 16, 22, 30, 49]. The World Organisation for Animal Health (WOAH) recognizes the significant economic threat posed by GPV, underscoring the need for robust diagnostic protocols in international trade and disease surveillance programs.

Classical Diagnostic Methods: Histopathology, Virus Isolation, and Electron Microscopy

Historically, the diagnosis of Derzsy’s disease relied on a combination of clinical signs, gross pathology, and histopathological examination. Acute GPV infection in goslings is characterized by hemorrhagic enteritis, hepatitis, and myocarditis [3, 8, 72]. Histopathological lesions include extensive lymphocyte necrosis in the thymus, bursa of Fabricius, and spleen, along with the presence of intranuclear inclusion bodies in hepatocytes and enterocytes [36, 72]. While these findings are suggestive, they are not pathognomonic, as co-infections with astroviruses or circoviruses can produce overlapping lesions [1, 56]. Detailed histopathological studies have been instrumental in understanding the immunopathogenesis of NGPV, revealing necrotic thymic cortex, bursal follicle atrophy, and apoptotic B-lymphocytes in the spleen of naturally infected ducks [10, 17].

Virus isolation remains a cornerstone for definitive diagnosis and for generating starting material for vaccines and molecular characterization [3, 37]. The virus is traditionally isolated by inoculating 9- to 12-day-old embryonated goose or Muscovy duck eggs via the allantoic cavity or yolk sac [3, 20, 69]. Successful isolation is indicated by embryo mortality, characteristic lesions including hemorrhages and stunting, and the presence of viral antigens in allantoic fluid. Primary cell cultures derived from goose embryos, such as goose embryo fibroblasts (GEFs) or goose embryo kidney (GEK) cells, are also highly permissive systems [8, 54]. However, the sensitivity of cell culture is limited by the host cell tropism of specific GPV strains. Studies have demonstrated that certain NGPV strains replicate efficiently in duck embryo fibroblasts (DEFs) but poorly in GEFs, highlighting the importance of matching cell substrate to the suspected viral lineage [51, 69, 70]. Furthermore, heterologous cell cultures from cattle, pigs, or monkeys have been shown to be non-permissive for GPV replication, reinforcing the necessity of using waterfowl-derived primary cells for initial isolation [54]. The appearance of a typical cytopathic effect (CPE), characterized by cell rounding, detachment, and syncytia formation, usually requires 3-7 days post-inoculation [27, 69]. Transmission electron microscopy (TEM) of purified virus from infected cell cultures or allantoic fluid provides definitive morphological confirmation, revealing non-enveloped, icosahedral particles with a diameter of approximately 20-30 nm, consistent with the Parvoviridae family [3, 6].

Molecular Diagnostics: The Gold Standard for Detection and Differentiation

Nucleic acid amplification tests (NAATs) have become the gold standard for GPV diagnostics due to their unparalleled sensitivity, specificity, and speed. A wide array of PCR-based methods have been developed, ranging from conventional gel-based PCR to highly sophisticated real-time quantitative PCR (qPCR) and digital PCR (dPCR) platforms.

Conventional and Multiplex PCR: Conventional PCR targeting conserved regions of the viral genome, such as the VP3 or VP1 genes, provides a rapid means of detecting GPV DNA from tissue samples, swabs, or allantoic fluid [28, 72]. However, the high genetic similarity between classical GPV (cGPV), NGPV, and Muscovy duck parvovirus (MDPV) has driven the development of type-specific and multiplex PCR assays. A key advancement is the design of primer sets that allow for the simultaneous detection and differentiation of these pathogens in a single reaction. For instance, a duplex PCR assay using a three-primer system targeting the NS gene can specifically differentiate MDPV and GPV by generating amplicons of distinct sizes [34]. More complex multiplex panels enable the simultaneous detection of GPV, waterfowl reovirus (WRV), and goose astrovirus (GAstV) [53], as well as GPV alongside goose astrovirus 1 and 2 [55]. Given the frequent co-infections observed in field settings, particularly of GPV with DuCV, GoCV, and GHPV, such multiplex panels are invaluable for comprehensive flock health monitoring [1, 59]. The limit of detection (LOD) for these conventional multiplex assays is typically in the range of 1×10³ to 1×10⁴ copies/µL, which is sufficient for clinical diagnosis but less sensitive than real-time methods [53, 55].

Real-Time Quantitative PCR (qPCR): The advent of qPCR has dramatically improved the sensitivity and quantification capability for GPV detection. TaqMan probe-based qPCR assays provide a dynamic range, allowing for absolute quantification of viral genome copies, which is crucial for understanding viral kinetics, pathogenesis, and response to vaccination [32, 68]. Numerous assays have been developed targeting different genomic regions. A pioneering assay targeting the inverted terminal repeat (ITR) region achieved a sensitivity of 10 copies per reaction, vastly outperforming conventional PCR [68]. The specificity of qPCR is further enhanced by the use of locked nucleic acid (LNA)-modified probes, which increase the melting temperature and allow for precise differentiation of virulent and attenuated vaccine strains, a critical tool for disease control programs [61].

The need to differentiate between cGPV and NGPV is a major diagnostic challenge, as their nucleotide homology is often >95% [24, 32]. Wan et al. [32] designed a highly specific TaqMan qPCR targeting a conserved region in the NS gene that is distinct from MDPV, enabling the specific detection and differentiation of cGPV and NGPV, particularly when coupled with host specificity data (e.g., NGPV is associated with SBDS in ducks, while cGPV causes classic Derzsy’s disease in goslings and Muscovy ducks). More recently, a TaqMan-based duplex one-step RT-qPCR has been established for the simultaneous detection of NGPV and novel duck reovirus (NDRV), achieving a LOD of 2.42 copies/µL for NGPV and demonstrating high throughput and accuracy for co-infection studies [60]. For laboratories requiring the highest sensitivity, duplex crystal digital PCR (dPCR) has emerged as a superior alternative to qPCR. By partitioning the sample into thousands of nanoliter-sized reactions, dPCR allows for absolute quantification without the need for a standard curve. In a head-to-head comparison for detecting MDPV and GPV, dPCR demonstrated a LOD of 0.3 copies/µL, which was approximately 38-fold more sensitive than qPCR, making it an ideal tool for detecting low viral loads in clinically healthy carriers or early-stage infections [67].

Isothermal Amplification and CRISPR-Based Diagnostics for Point-of-Care Testing

The reliance on thermocyclers and specialized laboratory infrastructure limits the deployment of PCR-based methods in resource-limited settings or during field outbreaks. To address this, isothermal amplification technologies have been developed, offering rapid, sensitive, and equipment-light alternatives.

Loop-Mediated Isothermal Amplification (LAMP): LAMP assays amplify DNA with high specificity and efficiency under constant temperature conditions (typically 60-65°C) within 60 minutes. A quantitative LAMP (qLAMP) assay targeting the VP3 gene of NGPV demonstrated a LOD of 1.0×10² copies/µL and showed no cross-reactivity with other duck pathogens [40]. The results can be visualized by the naked eye using a colorimetric dye or by measuring turbidity, making it highly suitable for field use. A separate LAMP assay for GPV similarly reported superior sensitivity compared to conventional PCR and a reduced turnaround time [19].

Recombinase Polymerase Amplification (RPA) and CRISPR/Cas Systems: RPA represents a further advancement, operating at a constant low temperature (37-42°C) and achieving amplification in as little as 5-20 minutes. An RPA assay combined with a vertical flow (VF) visualization strip for GPV and NGPV detection successfully amplified the VP3 gene, achieving a LOD of 2×10² copies and 100% concordance with qPCR for field samples [70]. The true game-changer in POC diagnostics, however, is the integration of RPA with CRISPR/Cas systems. The RPA-CRISPR/Cas12a platform exemplifies a major leap forward. In this system, RPA is first used to amplify the target DNA from the VP3 gene. The amplified product is then recognized by a Cas12a-crRNA complex, which activates the nuclease's collateral cleavage activity against a fluorophore-quencher-labeled reporter or a lateral flow assay (LFA) reporter [5, 66].

This dual-readout capability provides exceptional flexibility. Using a fluorescence-based readout, an RPA-CRISPR/AsCas12a system achieved a LOD of 7.8 copies/µL, a 1000-fold improvement over conventional PCR [66]. An RPA-CRISPR/Cas12a assay using a portable blue light transilluminator for visual detection achieved a diagnostic sensitivity of 100% and specificity of 95.5% against qPCR in clinical samples, with a total turnaround time of under one hour [5]. Another elegant approach combines RPA with the Pyrococcus furiosus Argonaute (PfAgo) protein, which cleaves nucleic acids with high specificity and without the need for a protospacer adjacent motif (PAM) sequence. This RPA-PfAgo system also demonstrated a LOD of 10² copies/µL and showed high concordance with qPCR in clinical testing [12]. These CRISPR-based systems are not only rapid and sensitive but are also highly specific, showing no cross-reactivity with related viruses like MDPV, duck plague virus, or duck hepatitis virus [5, 66]. The recent development of CRISPR-based lateral flow assays (CRISPR-LFA) provides a truly instrument-free, paper-strip readout, making it the most promising candidate for widespread field deployment and early outbreak detection.

Serological Assays: Monitoring Immune Status and Vaccine Efficacy

Serological testing plays a vital role in determining flock exposure history, monitoring maternal antibody decay, and evaluating the efficacy of vaccination programs. The gold standard serological test has historically been the virus neutralization test (VNT), which measures functional antibodies capable of blocking viral infectivity [58]. While specific and informative, VNT is labor-intensive, requires live virus and cell culture, and takes several days to complete.

To overcome these limitations, more rapid and scalable enzyme-linked immunosorbent assays (ELISAs) have been developed. A highly effective indirect ELISA uses recombinant VP3 protein subunits expressed in E. coli as the coating antigen. This assay demonstrated high sensitivity and specificity when compared to VNT, making it suitable for large-scale serosurveillance [58]. The VP3 protein is an ideal antigenic target due to its high immunogenicity and structural role in the capsid [58, 64]. Similarly, VP2-based ELISAs and virus-like particle (VLP)-based ELISAs have been developed for NGPV, showing strong correlation with antibody protection levels [63, 65]. However, a significant challenge in serodiagnosis is the antigenic variation between cGPV and NGPV. It has been observed that antibodies raised against cGPV may not effectively neutralize NGPV [43], which underscores the need for variant-specific serological tests to accurately assess immunity against the circulating strain.

For rapid, field-based antibody detection, colloidal gold immunochromatographic strip assays (ICG) have been developed. These strips are simple, require no equipment, and provide a visual result within 10-15 minutes. An ICG assay using a monoclonal antibody against GPV and a polyclonal antibody against VP3 showed good sensitivity (LOD of 1.2 µg/mL) and high specificity with no cross-reactivity to other goose pathogens [71]. While less sensitive than ELISA, ICG strips are invaluable for on-farm screening and rapid assessment of flock immune status.

Integrated Diagnostic Strategies for Surveillance and Control

The diagnostic approach for GPV must be strategic and context-dependent. For initial outbreak investigation in a naive flock, a combination of histopathology, virus isolation with TEM, and broad-spectrum molecular detection (e.g., multiplex PCR or NGPV-specific qPCR) is recommended to confirm the presence of GPV and rule out other common agents like DuCV or astrovirus [1, 16, 56]. For routine surveillance and quantifying vaccine-induced protection, a combination of qPCR for viral genome detection in swabs or organ samples and ELISA for serological monitoring provides a comprehensive picture of infection and immunity [58, 68]. The high sensitivity of duplex dPCR is particularly advantageous for detecting low-level viral carriage in breeder flocks, which is critical for preventing vertical transmission [20, 32].

For large-scale epidemiological studies and monitoring genetic evolution, the isolation of the virus followed by whole-genome sequencing is paramount. Next-generation sequencing (NGS) has been instrumental in identifying novel recombinant strains, such as the three-region recombinant MDGPV strain [13] and NGPV strains with specific mutations in the ITR region that modulate pathogenicity [9, 41]. Phylogenetic analysis of the VP1 and VP3 genes is essential for typing isolates into classical GPV, NGPV, or MDPV clades and for tracking transboundary spread [18, 30, 33]. Furthermore, the development of specific clinical indices, such as a three-dimensional beak development model for ducks infected with NGPV, can aid in the non-invasive phenotyping of pathogenesis and the evaluation of vaccine efficacy in live birds [62]. Looking ahead, the field is moving toward highly integrated POC platforms that combine nucleic acid extraction, isothermal amplification (RPA or LAMP), and CRISPR-based readout on a single microfluidic chip. Such devices could deliver a definitive diagnosis within 30 minutes directly on the farm, facilitating immediate containment measures and reducing reliance on central laboratories. The ongoing genetic evolution of GPV, marked by recombination and cross-species transmission, demands that all diagnostic tools be continuously re-evaluated and updated to maintain their relevance and accuracy against emerging viral lineages [3, 13, 43].

Clinical Manifestations and Disease Impact

Overview and Syndromic Classification of Goose Parvovirus Infection

Goose parvovirus (GPV) is the etiological agent of Derzsy’s disease, an acute, highly contagious, and often fatal septicemic condition of young geese and Muscovy ducklings, recognized globally as one of the most economically devastating viral infections in waterfowl production [1, 2, 6]. The clinical presentation of GPV infection is remarkably dichotomous, determined by host species, viral lineage, and age at exposure. Classical GPV (cGPV) strains induce a fulminant hemorrhagic and necrotic enteritis in goslings, whereas novel GPV (NGPV) variants, which have emerged as dominant lineages since approximately 2015, primarily cause short beak and dwarfism syndrome (SBDS) in Pekin ducks, Cherry Valley ducks, and mule ducks, with a markedly different clinical trajectory [4, 24, 30]. This host-driven tropism and pathogenicity profile is now well-established: MGPV (mutated GPV) is more pathogenic to goslings, eliciting classic gosling plague pathology, while NGPV is distinctly more pathogenic to ducklings, disrupting skeletal development and producing SBDS [4]. The World Organisation for Animal Health (WOAH) recognizes GPV as a significant transboundary pathogen of waterfowl, underscoring its continued threat to global poultry biosecurity.

Disease in Goslings: Derzsy’s Disease (Gosling Plague)

In goslings under one month of age, GPV infection is characterized by an acute, peracute, or subacute course with mortality rates that can approach 100% in naive flocks [3, 6, 64]. The incubation period is short, typically 2–5 days, after which affected goslings exhibit profound depression, anorexia, ruffled feathers, weakness, and a characteristic reluctance to move. Ocular and nasal discharges are common, frequently associated with bilateral periorbital swelling and conjunctivitis [26]. Diarrhea is a hallmark sign, ranging from watery to hemorrhagic, and the feces often contain fibrous casts or necrotic mucosal fragments [3]. The disease progresses rapidly; many birds die within 24–48 hours of clinical onset.

At necropsy, the most striking lesions are confined to the gastrointestinal tract. The intestinal serosa is congested, and the intestinal wall is markedly thinned and translucent. The lumen contains a large quantity of fibrinous exudate, blood-tinged fluid, and necrotic debris, often described as “sausage-like” casts that occlude the intestinal lumen [3, 72]. The liver is swollen, friable, and exhibits multifocal hemorrhages and areas of necrosis. Myocarditis, characterized by pale streaking and serous effusion in the pericardial sac, is consistently observed. Hemorrhages are also noted on the epicardium, spleen, and in the brain [72]. Mortality typically peaks at 7–10 days post-infection, with surviving goslings exhibiting severe growth retardation, feathering abnormalities, and persistent immunosuppression [8].

The pathophysiological basis for this high mortality involves a combination of direct viral cytopathology and systemic inflammatory dysregulation. Transcriptomic profiling of GPV-infected goslings reveals a marked increase in immunosuppressive factors such as TGF-β and IL-10, while pro-inflammatory cytokines (IL-4, IFN-γ, TNF-α) remain unchanged, indicating a shift toward an anti-inflammatory, pro-apoptotic state that facilitates viral persistence and tissue destruction [8]. GPV infection robustly activates both apoptosis and ferroptosis pathways in goose embryo fibroblasts, evidenced by upregulation of PTGS2, TF, and ASCL1, leading to programmed cell death in critical tissues such as the liver and jejunum [8]. Furthermore, the non-structural protein NS1 induces an apoptosis-inducing factor (AIF)-mediated mitochondrial apoptotic pathway, characterized by mitochondrial depolarization, increased reactive oxygen species, and elevated cathepsin D activity [25]. This NS1-driven apoptosis is accompanied by increasing viral load, creating a vicious cycle of cell death and viral amplification.

Disease in Ducks: Short Beak and Dwarfism Syndrome (SBDS) and Beak Atrophy and Dwarfism Syndrome

The emergence of NGPV in duck populations has fundamentally altered the epidemiological landscape of GPV. SBDS, first recognized in China in 2015 and subsequently reported in Europe (Poland, Egypt) and Vietnam, is characterized by a triad of clinical signs: beak atrophy, tongue protrusion, and severe growth retardation [20, 24, 27, 30]. Morbidity in affected duck flocks ranges from 15% to 80%, while mortality is typically low (4–20%), though the economic losses due to reduced body weight, poor feed conversion, and carcass condemnation are substantial [27, 30].

Affected ducklings present with a shortened, atrophied beak that fails to develop proportionally with the rest of the head. The tongue becomes protruding, swollen, and often necrotic or congested at the tip, particularly in older birds [46]. The tongue histopathology reveals necrosis of the superficial epithelial layer, vacuolar degeneration, and lymphoplasmacytic glossitis, with viral antigen detectable via immunohistochemistry in the propria submucosa [46]. The pathogenesis of the beak deformity is direct: NGPV disrupts the expression of genes critical for skeletal development, including MMP2, MMP9, MMP13, and CCN3, as identified in transcriptomic analyses of infected goslings [8]. The development of a three-dimensional Beak Development Index (BDI), integrating beak length, beak width, and body weight, has provided a quantitative tool to discriminate between primary beak dysplasia and secondary growth delay, confirming that NGPV specifically attacks the growth plates of the beak [62].

Skeletal abnormalities extend beyond the beak. Ducks exhibit shortened tibiae and tarsal bones, brittle bones prone to fracture, and generalized dwarfism [9, 44]. Bone histopathology demonstrates osteoclastic activation, increased bone resorption, and impaired osteogenesis [44, 45]. The virus also induces profound intestinal dysbiosis: 16S rDNA sequencing reveals a significant reduction in beneficial Firmicutes and Bacteroidota, coupled with decreased production of short-chain fatty acids (SCFAs) such as butyrate and propionate [44, 45]. This microbial disruption compromises intestinal barrier integrity, as evidenced by increased intestinal permeability and bacterial translocation, which in turn amplifies systemic inflammation through the Malt1/NF-κB signaling pathway [45]. Importantly, depletion of gut microbiota via antibiotics alleviates SBDS symptoms, improves bone quality, and restores intestinal homeostasis, demonstrating that the gut–bone axis is a central mediator of NGPV pathogenicity [45].

Neurological and muscular involvement is also a feature of NGPV infection. Ducks suffering from locomotor disorders exhibit myofiber atrophy and degeneration, severe enteritis with lymphocytic infiltration, and in the brain, vasculitis, diffuse gliosis, and Purkinje cell degeneration in the cerebellum [17]. Immunohistochemistry confirms NGPV antigen in muscle fibers, enterocytes, and cerebellar Purkinje cells, providing direct evidence of viral replication in tissues responsible for locomotion [17]. The high detection rate of NGPV in skeletal muscle (70.8%) and intestine (91.7%) from affected ducks underscores the multi-organ tropism of the virus [17].

Co-Infections and Their Synergistic Impact

One of the most critical clinical realities in modern waterfowl production is the ubiquity of polymicrobial infections. GPV is rarely encountered as a sole pathogen. Epidemiological surveys in Poland, China, and Egypt consistently demonstrate that GPV circulates concurrently with duck circovirus (DuCV), goose circovirus (GoCV), goose hemorrhagic polyomavirus (GHPV), astroviruses, reoviruses, and bacterial opportunists such as Escherichia coli, Erysipelothrix rhusiopathiae, and Gallibacterium anatis [1, 16, 56].

Co-infection with DuCV is particularly consequential. DuCV is an immunosuppressive agent that impairs T-cell responses and humoral immunity, rendering ducks highly susceptible to NGPV. Experimental co-infection models in Cherry Valley ducks reveal that DuCV and NGPV synergistically potentiate each other’s replication. Co-infected ducks exhibit significantly higher viral loads in serum and tissues, more severe immunosuppression (reduced lymphocyte counts, elevated total bilirubin), and exacerbated SBDS clinical signs compared to single-virus controls [16, 22]. The co-infection also impairs calcium and phosphorus metabolism, further compounding skeletal deformities [22]. Field data corroborate these findings: in Polish Pekin ducks with SBDS, DuCV co-infection was detected in 85.7% of cases, and in Chinese feather shedding syndrome (FSS), the co-detection rate of NGPV and DuCV was 70%, with 82.78% of feather sac samples positive for NGPV [30, 48]. The presence of both viruses in feather follicles and Lieberkühn crypt epithelial cells is associated with feather loss, breakage, and the formation of residual feather burrs in duck carcasses, a significant quality defect in processing plants [7, 31].

Triple or quadruple infections further compound morbidity. In Poland, 22.2% of commercial goose flocks harbored GPV, GoCV, and GHPV simultaneously, and such flocks exhibited the highest mortality rates, growth diversification, and reduced production metrics [1]. Similarly, co-infection with goose astrovirus (GAstV) and GPV in goslings produces a unique syndrome of visceral gout, urate deposition on the heart, liver, and kidneys, accompanied by enteritis, joint swelling, and paralysis, with histologic evidence of heterophil myelocyte infiltration in multiple organs [56]. The presence of multiple enteric viruses (GPV, waterfowl reovirus, astrovirus) in Muscovy ducks is associated with severe viral enteritis and high mortality, complicating differential diagnosis and therapeutic intervention [53].

Impact on Immune Organs and Immunosuppression

GPV exerts a profound and multifaceted immunosuppressive effect, which is a major driver of secondary infections and disease severity. In both natural and experimental infections, the virus targets primary and secondary lymphoid organs, leading to atrophy, lymphocytic depletion, and architectural disruption.

In the thymus, GPV infection causes necrotic changes in the cortex, with disintegration of Hassall’s corpuscles and loss of thymocytes. The bursa of Fabricius undergoes severe follicular atrophy, lymphoid depletion, and obliteration of normal histological morphology, effectively crippling B-cell development [10, 36]. The spleen exhibits diffuse lymphocytic apoptosis and depletion of periarteriolar lymphoid sheaths [10, 36]. Immunohistochemical staining reveals abundant GPV antigen in thymic cortical lymphocytes, bursal medullary lymphoid follicles, and splenic parenchyma, confirming active viral replication within immune cells [10]. Quantitative PCR demonstrates the highest viral loads in the spleen (peaking at 7 days post-infection), followed by bone marrow, peripheral blood lymphocytes, and cecal tonsils [36].

The functional consequence of this lymphoid destruction is a state of profound immunosuppression characterized by reduced circulating lymphocytes, impaired interferon responses, and enhanced susceptibility to opportunistic pathogens. At the molecular level, NGPV VP1 protein directly targets IRF7, a master regulator of type I interferon signaling, thereby blocking the cyclic GMP–AMP synthase (cGAS)–STING pathway and suppressing IFN-β induction [15]. This immune evasion mechanism permits unchecked viral replication and facilitates persistent infection, as demonstrated by the detection of high viral loads in surviving ducks weeks after clinical recovery [20]. The virus also upregulates immunosuppressive cytokines such as TGF-β and IL-10, further dampening the host’s ability to mount an effective adaptive immune response [8].

Feathering Disorders and Integumentary Signs

A growing body of evidence links GPV infection to a spectrum of feathering abnormalities, including feather shedding syndrome (FSS), feather loss disease, and broken feather disease. In Cherry Valley ducks, FSS presents at 4–5 weeks of age with a 20–70% incidence rate; affected birds exhibit spontaneous feather loss, poor feather regrowth, and feather shafts that are brittle and break during processing, resulting in “feather burr” carcasses [7, 48]. The causative role of NGPV in these lesions has been confirmed by the isolation of infectious NGPV from feather sacs and the experimental reproduction of feathering disorders following NGPV inoculation of 8-day-old ducks [7]. In Taiwan, gosling feather loss disease (GFL) and goose broken feather disease (GBF) at 21–60 days of age are highly correlated with dual infection by GoCV and GPV, with viral loads in feather follicles peaking during the third to fifth week of life, coinciding with maximal feather loss [31].

The pathogenesis involves viral replication in the feather follicle epithelium and the dermal papilla, leading to follicular necrosis, dystrophic feather development, and activation of cell death pathways. These integumentary manifestations, while not directly lethal, represent a major source of economic loss due to carcass downgrading and reduced pelt quality.

Angel Wing Syndrome and Locomotor Abnormalities

In a novel and striking clinical finding, GPV has been implicated as an etiological agent of angel wing syndrome (carpal valgus deformity) in Muscovy ducks [21]. First reported in Egyptian Muscovy duck flocks in 2017, affected birds developed bilateral or unilateral outward rotation of the carpal joints, preventing normal wing folding. Isolation and phylogenetic analysis of GPV strains from these flocks confirmed them as GPV variants closely related to both Derzsy’s disease and SBDS strains. Experimental oral inoculation with one isolate (HS1) reproduced both classic GPV signs and angel wing deformity in Muscovy ducks, with less severity in geese [21]. This is the first documented viral cause of angel wing syndrome, which had previously been attributed solely to dietary, environmental, or genetic factors.

Locomotor dysfunction is now recognized as a prominent feature of NGPV infection in ducks. Affected birds display lameness, reluctance to stand, and in severe cases, paralysis. Histopathological examination reveals myofiber atrophy and necrosis, polyneuritis, and cerebellar Purkinje cell degeneration [17]. Viral antigen is present in skeletal muscle, sciatic nerve, and brain tissue, confirming that NGPV directly damages the neural and muscular apparatus of locomotion [17]. These findings underscore the neurotropic potential of GPV and its capacity to cause debilitating motor deficits.

Economic and Production Impact

The disease impact of GPV extends far beyond mortality figures. The virus imposes a staggering economic burden on the waterfowl industry through reduced weight gain, feed conversion inefficiency, increased culling, carcass condemnation, and the costs of biosecurity and vaccination.

In commercial geese, GPV infection, even in subclinical or chronic forms, results in marked growth retardation and weight loss. Surviving goslings weigh significantly less than uninfected cohorts at market age, and the degree of dwarfism correlates with viral load and the severity of histopathological lesions in the liver and jejunum [8]. Flock uniformity is severely compromised, leading to sorting difficulties and reduced market value. In duck operations, the economic impact of SBDS is profound, morbidity can reach 70–80%, and while mortality is low, the affected ducks fail to reach target slaughter weight, and their carcasses are often condemned due to beak deformities, feather abnormalities, or skeletal fractures [27, 30]. The presence of residual feather burrs in processed duck carcasses, a direct consequence of NGPV-induced follicular damage, results in significant downgrading and rejection at slaughterhouses [7].

The role of vertical transmission exacerbates the problem. NGPV is shed in the reproductive tract of breeder ducks and can be transmitted to embryonated eggs, leading to infected day-old ducklings that enter the production cycle already compromised [32, 50]. This vertical route, combined with horizontal transmission through contaminated transport vehicles [11], contaminated feed, and feather dust, ensures that once a farm is infected, eradication is exceedingly difficult.

In Poland, a major European goose producer, GPV is reportable to veterinary authorities, reflecting its epizootic significance. The prevalence of GPV in Polish flocks is 100% at the flock level, with 30.8% of isolates classified as NGPV [1]. Co-infections with GoCV and GHPV were present in 74.1% and 22.2% of flocks, respectively, and these polymicrobial infections consistently produced the poorest health and production outcomes [1]. The cumulative impact of GPV and its co-infecting partners on flock-level mortality, growth performance, and treatment costs underscores the necessity for comprehensive control strategies that address both the primary virus and its immunocompromising synergists.

Genetic Diversity and Evolution of Goose Parvovirus

The genetic landscape of goose parvovirus (GPV) is a dynamic and rapidly evolving tapestry, marked by the emergence of distinct lineages, frequent recombination events, and an expanding host range that challenges traditional virological classifications. The World Organisation for Animal Health (WOAH) recognizes GPV as a significant pathogen of waterfowl, and its genetic diversification has profound implications for disease surveillance, vaccine efficacy, and biosecurity protocols globally. Understanding this diversity is not merely an academic exercise; it is a prerequisite for controlling Derzsy’s disease in geese and the increasingly important short beak and dwarfism syndrome (SBDS) in ducks.

Phylogenetic Architecture: The Classical and Novel Lineages

Phylogenetic analyses have consistently resolved waterfowl parvoviruses into three major clusters: classical GPV (C-GPV), Muscovy duck parvovirus (MDPV), and the more recently recognized novel GPV (N-GPV or NGPV) [18]. This tripartite structure, however, belies a more complex genetic continuum. Within the GPV clade, C-GPV and N-GPV form two distinct sub-branches, each further divisible into three subgroups, underscoring the significant intra-lineage genetic diversity that has accumulated over decades of co-evolution with their hosts [18]. The defining genetic boundary between C-GPV and N-GPV is not arbitrary; comparative genomics of Chinese NGPV strains reveals a nucleotide homology of 95.2%–96.1% with classical GPV strains, a divergence that is accompanied by a suite of signature amino acid substitutions [24, 29]. Specifically, six NGPV isolates analyzed by Li et al. were found to share 16 common amino acid changes in the VP1 capsid protein and 12 in the Rep1 non-structural protein when compared to classical Chinese GPV strains [24, 29]. Crucially, nine of these VP1 substitutions were identical to those found in the European GPV strain B, suggesting a deep ancestral link between European and Chinese NGPVs [24, 29]. This molecular fingerprint is stable enough to allow for the differentiation of lineages using molecular diagnostics, such as TaqMan qPCR assays targeting the NS gene, which can distinguish C-GPV from N-GPV with high specificity and sensitivity [32].

The emergence of N-GPV as a major pathogen is a testament to the power of microevolution. Initially isolated from ducks exhibiting SBDS in China in 2015, this lineage has since been documented in Poland, Egypt, Vietnam, and Turkey, indicating a global dispersal that likely follows trade routes [10, 18, 27, 30]. Polish isolates from the first European outbreak of SBDS in Pekin ducks shared 98.57–99.28% nucleotide identity with Chinese NGPV strains, yet they exhibited a higher rate of amino acid mutations in the Rep protein compared to the VP1 protein, a pattern that hints at adaptive selection pressures distinct from those acting on the capsid [30]. The evolutionary trajectory of these lineages is further illuminated by selection pressure analyses, which have identified specific sites under positive or purifying selection within the structural and non-structural genes, driving the fixation of beneficial mutations and the purging of deleterious ones [18].

Recombination as a Primary Engine of Genetic Novelty

Recombination is arguably the most potent force shaping the genetic diversity of GPV, acting as a molecular crucible that blends the genomic material of co-infecting parvoviruses [18]. Evidence for this process is abundant and mechanistically diverse, reflecting the complex ecology of waterfowl parvoviruses. An exhaustive analysis of 30 global GPV genomes identified no fewer than 11 independent recombination events, with breakpoints distributed across the genome, including the P9 promoter, NS region, and VP structural genes [18]. These events frequently involve C-GPV, N-GPV, and MDPV as parent strains, a consequence of the high rates of co-infection observed in commercial flocks [1, 18].

The recombinant progeny often possess novel pathogenic properties. For instance, the HLJ2023 strain, a highly pathogenic isolate from Heilongjiang, China, was characterized as a potential recombinant of an NGPV strain (JS191021) and a goose parvovirus hosted by Muscovy duck (GMD) PT strain [3]. SimPlot analysis revealed that its VP3 gene region was more closely related to duck parvovirus and NGPV than to classical GPV, and the resultant virus exhibited a 100% mortality rate in experimentally infected goslings [3]. Similarly, the Muscovy duck-origin goose parvovirus (MDGPV) 2022FZ strain represents a groundbreaking discovery: it is the first described waterfowl parvovirus with three distinct recombination regions [13]. These regions were located in the P9 promoter-NS region, the NS2 gene, and the VP3 gene, with the GPV sequences inserted into an MDPV genomic skeleton [13]. The presence of a recombination event within the NS2 gene was a novel finding, expanding the known repertoire of recombination hotspots. Remarkably, despite its complex chimeric structure, the 2022FZ strain was less pathogenic to Muscovy ducklings than previously identified recombinant strains, illustrating that recombination can lead to attenuation as well as increased virulence [13].

Recombination is not confined to interactions between wild-type strains. The involvement of live attenuated vaccine strains in recombination events is a particularly alarming phenomenon. The SDLY1602 NGPV strain was shown to be a recombinant between the classical GPV vaccine strain 82-0321v and a wild GPV strain (GDaGPV) [35]. This finding demonstrates that vaccine-derived genetic material can re-enter the wild population, potentially creating novel variants with unpredictable virulence. The HuN18 strain, isolated from Linwu sheldrakes, also showed evidence of recombination, with the NGPV sdlc01 strain as the major parent and the classical GPV Y strain and the vaccine-related SYG61v strain as minor parents [33]. This underscores the risk that live vaccines, while effective at preventing disease, can serve as a reservoir of genetic material for ongoing recombination.

Cross-Species Transmission and Host Adaptation

The ability of GPV to jump between goose and duck hosts is a central theme of its evolutionary biology. Classical GPV is highly pathogenic in goslings and Muscovy ducklings but causes little to no disease in Pekin ducks [32]. Conversely, NGPV is the etiological agent of SBDS in Cherry Valley, Pekin, and Mule ducks, while exhibiting reduced pathogenicity in geese [4, 32]. This host tropism is not absolute but is a gradient, with specific viral lineages demonstrating a clear preference. Comparative pathogenicity studies have confirmed that MGPV (a lineage of mutated GPV) is more pathogenic to goslings, while NGPV is more pathogenic to ducklings [4]. This differential pathogenicity is mirrored by the humoral response: MGPV elicits a stronger antibody response in goslings, whereas NGPV generates a more robust response in ducklings [4].

The molecular determinants of this host switching are being gradually elucidated. The VP3 capsid protein is a major determinant of receptor binding and host specificity. A comparative study of a Vietnamese NGPV isolate (DuPV-BAFU) revealed that its VP3 protein possessed a set of receptor-interacting amino acid residues that were identical to those found in the goose-origin strains GPVa2006 and GPV1995, but distinct from those in MDPV and other GPV strains [47]. This suggests that a specific molecular signature in VP3 is required for successful infection of duck hosts. Furthermore, the non-structural protein Rep1 plays a critical role in pathogenicity. Chimeric virus experiments, in which the Rep1 gene of NGPV was replaced with that from classical GPV, demonstrated that Rep1 is a key determinant of viral virulence in the duck host [20].

The inverted terminal repeats (ITRs) also contribute to host-specific pathogenicity. A reverse genetics system for NGPV allowed researchers to construct and rescue viruses that differed only in their ITR regions, and these viruses exhibited distinct virulence profiles in duck embryos [9]. This discovery provides direct experimental evidence that the ITRs, long considered merely replication origins, can modulate pathogenicity, potentially through their role in viral DNA replication and packaging. The E-box motif (CACATG) within the left ITR has been shown to influence virus replication and cell cycle progression in Muscovy ducks, with deletion of this element leading to reduced virulence [41].

Phylogeography and Global Dissemination

The global spread of GPV, and particularly NGPV, is a recent phenomenon driven largely by international trade in waterfowl and poultry products. Phylogeographic reconstructions have identified China as a major global source of GPV lineages [18]. The virus has subsequently radiated outward, with trade routes serving as the likely conduits for transmission [18]. The emergence of NGPV in Poland in 2019, followed by its detection in Egypt, Turkey, and Vietnam, suggests a pattern of introduction from Chinese sources, likely via the movement of infected breeding stock or contaminated poultry products [18, 27, 28, 30].

The genetic homogeneity of NGPV strains within a given region can be striking. Eight NGPV strains isolated from Shandong and Henan provinces in China in 2023 shared nucleotide homologies of 99.9%–100%, indicating a lack of major mutational drift in recent years [14]. In contrast, the Egyptian NGPV isolates from ducks with locomotor disorders and immune organ atrophy were found to be closely related to Chinese NGPV isolates but exhibited distinct pathological features, including a high detection rate in the intestine (91.7%) and skeletal muscle (70.8%), suggesting that local adaptation or co-infection with other pathogens may influence tissue tropism [10, 17]. The detection of NGPV in the tongue tissue of Pekin ducks with SBDS has further expanded the known cellular tropism of the virus, with immunohistochemical signals localized within the propria submucosa, providing a mechanistic link to the characteristic tongue protrusion [46]. These findings highlight the necessity for continuous genomic surveillance to track the emergence of new variants and monitor their potential for adaptation to new hosts and environments.

Co-infections and Management Strategies

The clinical manifestation and economic impact of goose parvovirus (GPV) infection in waterfowl production are rarely the result of a singular pathogenic event. Contemporary field surveillance and experimental pathology have unequivocally demonstrated that GPV and its genetic variant, the novel goose parvovirus (NGPV), frequently exist within a complex polymicrobial landscape. Mixed infections with other viral, bacterial, and fungal agents are the norm rather than the exception in commercial settings, profoundly influencing disease severity, tissue tropism, immunosuppression, and overall flock productivity. This section provides an exhaustive analysis of the documented co-infections involving GPV and NGPV, delineates the underlying mechanisms of synergistic pathogenicity, and critically evaluates current management and intervention strategies, including advances in differential diagnostics and the development of next-generation vaccines.

The Epidemiological Landscape of Co-infections

Longitudinal observational studies have provided high-resolution data on the prevalence of co-infections in waterfowl. A comprehensive two-year surveillance of commercial goose flocks in Poland revealed a near-universal presence of GPV genetic material (100% of flocks), with goose circovirus (GoCV) and goose hemorrhagic polyomavirus (GHPV) detected in 44.4% and 59.3% of flocks, respectively. Critically, dual-virus co-infections were found in 74.1% of flocks, while triple-virus co-infections involving GPV, GoCV, and GHPV were present in 22.2% of flocks [1]. This study highlighted that viral and bacterial co-infections are a significant problem; rarely did a single factor have a clear impact on health status. Typically, mixed viral infections, complicated by bacterial pathogens (predominantly Escherichia coli, and less frequently Erysipelothrix rhusiopathiae, Gallibacterium anatis, and Salmonella Typhimurium) or fungal complications, lead to increased mortality, growth diversification, and reduced production rates [1]. This high prevalence of polymicrobial involvement underscores the necessity of shifting diagnostic and therapeutic paradigms from a single-agent focus to a holistic, systems-level approach.

Synergistic Pathogenicity of Viral Co-infections

The interplay between GPV/NGPV and other immunosuppressive viruses is of particular concern, as it creates a vicious cycle of heightened susceptibility and exacerbated pathology.

Duck Circovirus and NGPV/GPV Co-infection

Duck circovirus (DuCV) is a ubiquitous pathogen known to induce immunosuppression in ducks [16]. The co-infection of DuCV with NGPV or GPV is arguably the most clinically significant viral synergy documented in the literature. In Cherry Valley ducks, co-infection with DuCV and NGPV produces a more severe form of short beak and dwarfism syndrome (SBDS) than NGPV alone. Clinical signs of short beak, dwarfism, and immunosuppression are more pronounced, and tissue damage in target organs (thymus, spleen, bursa of Fabricius) is markedly more severe [16]. Quantitative analysis reveals a dynamic synergistic effect on viral replication. In the early stages of co-infection, viral loads of both DuCV and GPV were significantly lower than in mono-infected birds, suggesting an initial immunological interference. However, as the infection progressed, the viral loads of both pathogens in co-infected ducks became significantly higher than those in mono-infected groups, with extended viral distribution in the liver, kidney, duodenum, spleen, and bursa of Fabricius [22, 49]. This temporal switch underscores a sophisticated pathogenic strategy where initial immune evasion paves the way for explosive, synergistic replication and dissemination.

Detailed experiments have demonstrated that co-infection selectively disrupts metabolic and immune functions. Ducks co-infected with DuCV and NGPV exhibited significantly reduced levels of serum calcium and phosphorus, alongside elevated total bilirubin, indicating impaired metabolic homeostasis. The lymphocyte count was also significantly reduced, confirming a state of profound immunosuppression [22]. This synergistic pathogenicity is likely driven by DuCV’s established ability to induce B-lymphocyte apoptosis and disrupt antigen presentation, thereby impairing the host’s ability to control NGPV replication. The resulting unbridled NGPV replication then leads to more severe damage to the skeletal system (as evidenced by altered bone metabolism markers) and the intestinal barrier [22, 45]. The high co-detection rate of NGPV and DuCV in feather sacs of ducks with feather shedding syndrome further implicates this synergy in novel clinical presentations, highlighting the viruses’ capacity to infect ectodermal tissues and cause significant economic losses due to carcass defects [48].

Goose Circovirus and GPV Co-infection

Similarly, the role of goose circovirus (GoCV) in exacerbating GPV pathology has been established in geese. In southern Taiwan, a high prevalence of GoCV and GPV co-infection was strongly correlated with gosling feather loss disease and goose broke feather disease. Quantitative PCR analysis demonstrated a high correlation between feather loss severity and the viral loads of both GoCV and GPV during the third to fifth weeks of age. Histopathological examination revealed inclusion bodies in feather follicles and Lieberkühn crypt epithelial cells, suggesting direct viral involvement in feather development pathology [31]. This synergistic effect on feathering disorders mirrors the DuCV-NGPV synergy in ducks and suggests a common mechanism where circovirus-induced immunosuppression allows for more robust GPV replication in epithelial tissues.

Waterfowl Astrovirus and GPV Co-infection

Co-infection of GPV with goose astrovirus (GAstV) represents another complex pathological scenario. Clinical cases of co-infection present with a severe, overlapping syndrome characterized by high mortality, depression, anorexia, enteritis, joint swelling, and paralysis. Postmortem findings reveal a combination of pathologies, including urate deposits covering internal organs (typical of astrovirus-induced visceral gout) alongside duodenal and ileal swelling (typical of GPV enteritis). Histologically, this co-infection induces extensive infiltration of heterophil myelocytes into the kidney, spleen, liver, lung, bursa of Fabricius, and pancreas, a finding unique to the co-infected state and not seen in mono-infections [56]. This suggests that the two viruses synergistically disrupt hematopoiesis and myeloid cell migration, leading to a dysregulated inflammatory response that amplifies tissue damage. The development of duplex diagnostic assays capable of simultaneously detecting GPV and GAstV has revealed clinical co-infection rates of 15% [75], highlighting the frequent occurrence of this dangerous combination.

The Role of Inverted Terminal Repeat Regions in Recombination and Emergence

The high frequency of co-infection creates an environment ripe for genetic recombination, a fundamental mechanism driving GPV evolution and the emergence of novel strains. The GPV genome, like other parvoviruses, features complex inverted terminal repeat (ITR) sequences at both ends. Using a reverse genetics system, researchers have demonstrated that the ITR regions play a significant role in modulating viral pathogenicity [9]. During co-infections, the replicative machinery of one parvovirus can recombine with the genome of another, leading to chimeric viruses. This process has been documented extensively. For instance, the Muscovy duck-origin goose parvovirus (MDGPV) strain 2022FZ was identified as having three recombination regions, in the P9 promoter-NS region, the NS2 region, and the VP3 gene, derived from both GPV and Muscovy duck parvovirus (MDPV) [13]. Similarly, novel GPV strains have been identified as potential recombinants of NGPV and classical GPV, with recombination events detected in the VP3 gene [3]. The global phylogeographic analysis of GPV indicates that China is a major exporter of these recombinant strains, with trade serving as a potential transmission conduit [18]. These recombination events are not merely academic; they can alter host range, virulence, and antigenicity, rendering existing vaccines partially or wholly ineffective and complicating management strategies.

Management Strategies: Diagnostics, Vaccination, and Immune Modulation

The management of GPV-related disease, particularly in the context of co-infections, requires a multi-pronged approach centered on sensitive differential diagnosis, strategic vaccination, and innovative immune-based interventions.

Advanced Differential Diagnostics

Given that clinical signs of GPV, NGPV, DuCV, GoCV, GAstV, and other pathogens overlap considerably, accurate diagnosis cannot rely on clinical observation alone. The development of high-throughput, multiplex molecular assays is therefore a cornerstone of modern management. Several advanced platforms have been developed to address this need:

1. Multiplex Quantitative PCR (qPCR): Quadruplex TaqMan-based qPCR assays now allow for the simultaneous detection and differentiation of MDPV, GPV, DuCV, and Duck adenovirus 3 (DAdV-3) with high sensitivity (detection limits of 1-10 copies/µL) and no cross-reactivity with other common avian pathogens [52]. Similarly, duplex assays have been developed for the simultaneous detection of NGPV and novel duck reovirus (NDRV), revealing a co-infection rate of 13.3% in clinical settings [60].

2. Duplex Crystal Digital PCR (dPCR): For samples containing extremely low viral loads, such as those from subclinical carriers or environmental samples, duplex crystal dPCR offers a significantly higher sensitivity (0.3 copies/µL) compared to conventional qPCR, which is critical for early detection and surveillance [67].

3. Isothermal Amplification with CRISPR: The integration of recombinase polymerase amplification (RPA) with CRISPR/Cas12a or AsCas12a systems represents a paradigm shift in field-deployable diagnostics. These assays achieve detection limits as low as 7.8 copies/µL within one hour, offer visual readouts using portable blue light transilluminators or lateral flow strips, and demonstrate high specificity without cross-reactivity to other waterfowl viruses [5, 66]. The RPA-CRISPR/Cas12a assay for GPV has shown 100% diagnostic sensitivity and 95.5% specificity in field validation, surpassing standard qPCR [5].

Vaccination Strategies

Vaccination remains the most effective long-term management strategy. However, the antigenic diversity between classical GPV and NGPV, along with the presence of multiple recombinants, poses a significant challenge. A single vaccine may not provide cross-protection against all circulating strains.

1. Live Attenuated Vaccines: Attenuation of virulent NGPV strains via serial passage in duck embryo fibroblast cells (DEFs) has yielded promising live vaccine candidates. For example, the NMG21 strain, passaged 35 times (NMG21-35), showed a clear decrease in pathogenicity for ducklings, with less tissue damage, lower tissue replication rates, and higher antibody levels compared to the parent strain [51].

2. Virus-Like Particle (VLP) Vaccines: VLPs represent a highly safe and immunogenic vaccine platform, as they lack viral nucleic acid and cannot replicate. Recombinant VP2 protein of NGPV, expressed via baculovirus in insect cells, self-assembles into VLPs morphologically identical to the native virion. These VLPs induce significant serum antibody responses in ducks and, crucially, provide complete protection against clinical signs, mortality, and viral shedding after NGPV challenge. No viral shedding was detected in the immunized group, whereas shedding was present in controls, indicating that VLPs can induce sterilizing immunity in the vaccinated host and potentially reduce horizontal transmission within the flock [65]. Similarly, VP2 VLPs produced in Saccharomyces cerevisiae have shown efficacy in geese, triggering effective protection against GPV infection [6].

3. Recombinant Subunit Vaccines: Expression of the VP2 protein in baculovirus, when mixed with an adjuvant like ISA 78-VG, stimulates strong antibody responses in breeding ducks. Maternal antibodies derived from vaccinated breeders provide sufficient protection to ducklings, a critical advantage for preventing early-life mortality [63]. The VP3 protein has also been exploited, with recombinant fragments used in ELISA for serological monitoring of antibody titers [58].

4. Oral Vaccine Vectors: Oral vaccines offer practical advantages for mass administration to large waterfowl flocks. Recombinant Lactobacillus casei expressing the GPV VP3 gene has been shown to colonize the intestine for approximately 34 days and induce both humoral and mucosal immune responses in goslings. Although the protection rate (30%) was comparable to a commercial vaccine, it highlights the potential for probiotic-based vaccination [64]. A more advanced approach using recombinant Lactococcus lactis expressing NGPV VP3 fused with duck interferon-alpha (IFNα) has shown superior efficacy. This construct induces robust mucosal (intestinal sIgA), humoral (serum IgG), and cellular (IFN-γ, IL-10) immune responses. Following challenge, vaccinated ducklings exhibited significantly reduced viral shedding and protection from severe intestinal damage, with the fusion protein group showing the lowest viral load [74].

5. DNA Vaccines and Adjuvants: Oral DNA vaccines encoding the VP2 protein, adjuvanted with cyclic peptide nanotubes (cPNTs), have demonstrated efficacy in inducing virus-specific IgA antibodies in the serum and intestinal tract of ducklings. The cPNTs act as both a delivery vehicle and an adjuvant, protecting the DNA from degradation and enhancing uptake by intestinal epithelial cells [73].

Immune Modulation and Microbiome-Based Interventions

Recent research has illuminated the critical role of the host's intestinal microbiota in modulating the severity of NGPV-induced SBDS. It has been demonstrated that NGPV infection induces dysbiosis in the cecal microbiota, characterized by a decreased abundance of Firmicutes and Bacteroidota, and a corresponding reduction in beneficial short-chain fatty acids (SCFAs) such as butyrate [44, 45]. This dysbiosis is not a passive consequence of infection; rather, it actively contributes to pathogenesis. The depletion of intestinal microbiota using broad-spectrum antibiotics (ABX) in an experimental model of SBDS resulted in significant alleviation of clinical signs, including improved body weight, beak length, and tarsal length. ABX-treated NGPV-infected ducks exhibited improved bone quality, reduced bone resorption, mitigated histopathological lesions in the intestine, and a marked reduction in systemic inflammation [45].

Mechanistically, the benefits of microbiota depletion were linked to reduced activation of the mucosa-associated lymphoid tissue lymphoma translocation protein 1 (Malt1) and nuclear factor κB (NF-κB) pathways, which are central drivers of inflammation in SBDS [45]. Conversely, transplantation of fecal microbiota (FMT) from healthy ducks into NGPV-infected ducks did not alleviate symptoms, indicating that the dysbiotic state is a driver of pathology [45]. This has profound implications for management: interventions aimed at restoring eubiosis, such as the use of specific probiotics, prebiotics, or SCFA supplementation, could serve as novel, non-antibiotic strategies to mitigate the impact of NGPV infections, particularly given global concerns over antimicrobial resistance. The modulation of the gut-liver-bone axis through microbiome manipulation represents a frontier for managing the chronic growth retardation and skeletal abnormalities characteristic of SBDS.

Biosecurity and Transmission Control

Beyond medical interventions, rigorous biosecurity is paramount. GPV is highly stable in the environment, and mechanical vectors play a critical role in its dissemination. A study evaluating the role of transport vehicles in GPV transmission found that insufficiently or inadequately disinfected vehicles, those transporting geese from hatcheries to farms and from farms to slaughterhouses, serve as a significant vector for GPV infections, even in flocks that have been preventively vaccinated [11]. This finding necessitates the implementation of stringent, validated cleaning and disinfection protocols for all equipment, vehicles, and personnel entering or exiting production facilities. Furthermore, the detection of infectious NGPV in the skin of duck carcasses with residual feather burrs at slaughterhouses indicates that contaminated carcasses can serve as a reservoir for the virus, potentially contaminating the processing environment and subsequent product batches [7]. This has implications for food safety and processing plant hygiene, aligning with the One Health approach advocated by the World Organisation for Animal Health (WOAH). The role of trade as a conduit for GPV spread, with phylogeographic data identifying China as a major exporter of the virus [18], underscores the need for international cooperation and standardized surveillance protocols to limit transboundary spread, a core tenet of the standards set by WOAH.

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