Search Results (59)

Avian leukosis virus (ALV), chicken infectious anemia virus (CIAV), and fowl adenovirus (FAdV) are significant immunosuppressive agents that cause huge financial losses to the poultry industry. In this study, specific primers and probes for ALV, CIAV, and FAdV were designed, and the RNA standard for ALV and plasmid standards for CIAV and FAdV were constructed. A triplex reverse transcription–crystal digital PCR (RT-cdPCR) assay was developed for the detection of ALV, CIAV, and FAdV after the optimization of key reaction parameters, including primer and probe concentrations, annealing temperature, and reaction cycle. Subsequently, the assay was assessed for its specificity, sensitivity, and repeatability. The results showed that the assay specifically detected ALV, CIAV, and FAdV, without cross-reactivity with other avian pathogens. The limits of detection (LODs) for sensitivity to ALV, CIAV, and FAdV were 3.75, 3.75, and 5.50 copies/reaction, respectively. Repeatability analysis showed the coefficients of variation (CVs) with 0.38–1.77% for intra-assay and 0.31–3.02% for inter-assay. The assay was applied to evaluate 1211 clinical samples from Guangxi Zhuang Autonomous Region in China and yielded positivity rates of 45.58% (552/1211) for ALV, 10.73% (130/1211) for CIAV, and 5.20% (63/1211) for FAdV. Additionally, these 1211 clinical samples were tested using the reference methods, and the results were compared with those of the established method, showing coincidence rates ≥ 98.18%. In conclusion, a triplex RT-cdPCR was successfully established for the simultaneous and accurate detection of ALV, CIAV, and FAdV. Full article

Marek’s disease (MD), caused by pathogenic Marek’s disease virus serotype 1 (MDV-1), is one of the most important avian immunosuppressive and neoplastic diseases and has led to huge economic losses to the poultry industry worldwide. Rapid and accurate clinical diagnosis is of great significance for efficient control of the disease. Herein, we have established a multiplex PCR (mPCR) method to simply differentiate all of the three types of MDV, using five specific primers targeting to MDV-1 oncogene meq or MDV-2 and MDV-3/HVT gB genes. Simultaneously, it can detect any type of virulent or vaccine MDV strains in one PCR reaction, with amplicons of the short (S) and long (L)-meq of MDV-1 strains, and the gB of MDV-2 and HVT vaccine strains. Non-specific amplifications of avian leukosis virus (ALV), reticuloendotheliosis virus (REV), or fowl adenovirus virus 4 (FAdV-4) were not observed, indicating a good specificity of this method. A total of 522 clinical samples of tumor-bearing or suspected diseased birds collected from 30 poultry farms were detected. The results demonstrated that the newly developed mPCR method accurately detected and differentiated epidemic MDV-1 infections and vaccine strains, and provided nearly 100% consistency for detecting clinical wild-type infections compared with conventional PCR amplification of the meq gene. Collectively, our data has provided a highly efficient method for early differential diagnosis of MD clinical cases, virus identification and future evaluation of vaccination efficacy in healthy chicken flocks, which would be meaningful for efficient control of the disease. Full article

Co-infection with avian leukemia and Pullorum Disease severely compromises poultry health, yet its pathogenic mechanisms remain unclear. We employed transcriptome sequencing to analyze gene expression changes and enriched pathways in kidney, spleen, and liver tissues of Chongqing Chengkou mountain chickens under single-infection (avian leukemia virus or Pullorum Disease) and co-infection conditions. Significant differences were observed in the number and pathways of differentially expressed genes between co-infected and single-infected groups. These genes were predominantly enriched in pathways involving extracellular matrix–receptor interactions, PPAR signaling, and calcium ion signaling. RT-qPCR validation confirmed significant upregulation of MAPK10 and SQLE, alongside downregulation of genes such as FOXG1. This study identifies multiple differentially expressed genes and pathways associated with immunity and tumorigenesis, providing crucial molecular insights into the regulatory mechanisms underlying avian leukemia and Pullorum Disease co-infection. Full article

The avian immunosuppressive and neoplastic diseases are great threats to the poultry industry, causing huge economic losses worldwide. Most recently, the emerging hypervirulent variants of Marek’s disease virus (HV-MDV), partially co-infected with avian leukosis virus (ALV) and/or reticuloendotheliosis virus (REV), have been identified as the key driver of tumour outbreaks in vaccinated chicken flocks, but the role of chicken infectious anemia virus (CIAV) remains unclear. Herein, we have investigated the prevalence and co-infection of CIAV in 71 clinical tumour-bearing flocks collected from central China during 2021–2023, which has shown a CIAV positivity rate of 59.2% (42/71). Notably, the incidence of CIAV mono-infection increased significantly from 0% (0/29) in 2021 to 23.7% (9/38) in 2023, whereas CIAV + MDV co-infection decreased from 65.5% (19/29) to 31.6% (12/38). A total of 20 viral genomes of epidemic CIAV isolates from diverse sources were obtained, and the phylogenetic analysis, including 91 reference isolates were clustered into four major lineages (A–D), with clade C further subdivided into subclades C1 and C2. Clade C1 consisted predominantly of Asian isolates, with 88.5% (46/52) of the isolates originating from mainland China. Among the 20 new isolates, 17 were clustered in subclade C1, two in C2, and one in B. The VP1 gene phylogeny showed a topology largely consistent with that of the whole-genome analysis. Moreover, all newly characterized isolates contained glutamine (Q) at VP1 residue 394, a molecular marker associated with high pathogenicity. Collectively, our data suggest that prevalent HV-MDV variants together with CIAV co-infections are the primary drivers of the ongoing tumour outbreaks in Chinese poultry flocks. Notably, the significantly increased CIAV mono-infections, possibly resulting from an independently evolving lineage among circulating Chinese strains, are likely to pose a new challenge for future control of disease. Full article

As a major retrovirus threatening global poultry farming, Avian Leukosis Virus Subgroup J (ALV-J) has expanded its host range since discovery, extending from conventional broilers to layer chickens and native breeds. Its diverse oncogenic manifestations, including myeloid leukemia, hemangiomas, and tumors of immune and visceral organs, have led to increased mortality, reduced productivity, and substantial economic losses in the poultry industry. Based on the current body of literature, this review summarizes and synthesizes advances in the etiological characteristics, infection and pathogenic mechanisms, host resistance, and research progress in prevention and control of ALV-J. Accumulating evidence indicates that viral evolution driven by mutations and recombination—particularly in the env gene and LTR regions—plays a central role in host range expansion, tumor diversity, and immune evasion. Current studies consistently demonstrate that host resistance to ALV-J is a multifactorial process involving genetic polymorphism, innate immune responses, and cellular autonomous defense systems. In this context, recent advances in disease-resistant breeding highlight CRISPR-Cas9-mediated gene editing as a promising strategy for blocking viral entry or replication. Despite these advances, major gaps remain, including an incomplete understanding of virus–host interaction networks, limited insight into co-infection-mediated synergistic pathogenicity, the absence of effective vaccines, and insufficient large-scale epidemiological surveillance and purification systems. Addressing these challenges will be critical for the development of integrated prevention strategies and the sustainable control of ALV-J in poultry production. Full article

Background/Objectives: Avian leukosis virus subgroup J (ALV-J) harms poultry via tumors and immunosuppression; MDM2-derived circRNA-0172′s role in ALV-J infection is unknown. This study explores its molecular characteristics, circularization mechanism and functions for ALV-J control. Methods: CEF/DF-1/HD11 cells were cultured; RNA/DNA was extracted. circRNA-0172 was validated via PCR, sequencing and RNase R; subcellular localization and circularization were analyzed with fractionation and vectors. miRanda/TargetScan/GO/KEGG, CCK-8 and qRT-PCR assessed targets, pathways and functions. Results: circRNA-0172 (1180 bp, MDM2 exons 3–11) is cytoplasm localized, RNase R resistant, and circularized via flanking ERVL-MaLR repeats. Highly expressed in immune tissues/HD11 cells, it is downregulated by ALV-J. Its overexpression inhibits DF-1 proliferation, downregulates Bcl2 and upregulates CCND1. Conclusions: This is the first confirmation of circRNA-0172′s stability and repeat-dependent circularization; it regulates cell proliferation/apoptosis, providing potential targets for ALV-J-associated avian leukemia. Full article

Avian leukosis virus subgroup J (ALV-J), an α-retrovirus, mediates infection by binding to the host-specific receptor chNHE1 (chicken sodium–hydrogen exchanger type 1), leading to immunosuppression and tumorigenesis, which severely threatens the sustainable development of the poultry industry. Studies have shown that the tryptophan residue at position 38 (W38) of the chNHE1 protein is the critical site for ALV-J infection. In this study, we employed the CRISPR/Cas9 system to construct a lentiviral vector targeting the W38 site of chNHE1, transfected it into chicken primordial germ cells (PGCs), and validated its antiviral efficacy through ALV-J infection assays, successfully establishing an in vitro gene-editing system for chicken PGCs. The constructed dual lentiviral vector efficiently targeted the W38 site. PGCs isolated from 5.5- to 7-day-old chicken embryos were suitable for in vitro gene editing. Stable fluorescence expression was observed within 24–72 h post-transfection, confirming high transfection efficiency. ALV-J challenge tests demonstrated that no viral env gene expression was detected in transfected PGCs at 48 h or 72 h post-infection, while high env expression was observed in control groups. After 7 days of infection, p27 antigen ELISA tests were negative in transfected groups but positive in controls, indicating that W38-deleted PGCs exhibited strong resistance to ALV-J. This study successfully generated ALV-J-resistant gene-edited PGCs using CRISPR/Cas9 technology, providing a novel strategy for disease-resistant poultry breeding and advancing avian gene-editing applications. Full article

Circular RNAs (circRNAs), a class of covalently closed non-coding RNAs, are pivotal regulators of gene expression and contributors to disease pathogenesis. This study elucidated the biogenesis, functional significance, and regulatory network of circRNA_4083, a novel exon-derived circRNA originating from exons 22 and 23 of the MSH3 gene in chicken. Through comprehensive molecular characterization—including Sanger sequencing, RNase R digestion assays, and subcellular localization—we confirmed the robust stability and predominant cytoplasmic localization of circRNA_4083 across diverse chicken tissues. Mechanistic investigations revealed that reverse complementary sequences within flanking intronic regions are indispensable for its circularization, as demonstrated by overexpression plasmids (#1–#4) in DF-1 cells. Functional analyses demonstrated that circRNA_4083 significantly inhibited cell apoptosis and increased cellular viability. Integrative bioinformatics approaches predicted a competing endogenous RNA (ceRNA) network comprising 12 miRNAs and 2132 target genes (FDR < 0.05), with significant enrichment in pathways critical to genomic stability, including non-homologous end joining (NHEJ) and ubiquitin-mediated proteolysis. These findings position circRNA_4083 as a key modulator of cellular viability and genomic integrity, with potential implications for avian leukosis virus-J (ALV-J) pathogenesis and resistance breeding strategies. This work advances our understanding of circRNA-driven regulatory mechanisms in avian species and underscores their relevance in poultry health. Full article

Avian leukosis virus (ALV), the prototypical alpharetrovirus, causes tumorigenesis, immunosuppression, and wasting disease in poultry. The ALV genus is classified into ten subgroups, which differ in their host range, cell tropism, and receptor usage. The subgroups A, B, K, and J cause significant economic losses worldwide. The most recently discovered subgroup, ALV-K, which is now widespread in China, has been shown to use the tva cell receptor and share it with ALV-A. However, the specific amino acid residues crucial for ALV-K host cell entry remain unknown. Using precise tva expression and chimeric tva receptors, we further elucidated the significance of the cysteine-rich domain in mediating interactions with both ALV-A and ALV-K. Through a comprehensive analysis of mutated tva receptor variants, we pinpointed tryptophan at position 33 (W33) as a pivotal amino acid residue essential for ALV-K virus binding and entry. Of note is the finding that the substitution of W33 induced resistance to ALV-K while preserving sensitivity to ALV-A. This study not only represents an advance in the understanding of the specificity of the tva receptor for ALV-K, but also offers a biotechnological strategy for the prevention of ALV-K infections in poultry. Full article

Avian leukosis (AL), a major vertically transmitted infectious disease, poses a significant challenge to the conservation and industrial development of indigenous chicken breeds in China. In this study, Chengkou mountain chickens were used as a model to systematically identify genetic markers associated with resistance to avian leukosis virus subgroup J (ALV-J) through a genome-wide association study (GWAS). Genomic DNA was extracted from 500 hens at 300 days of age, and cloacal swabs, plasma, and egg white samples were collected to assess the ALV-J infection status. A total of 325 ALV-positive (ALV+) and 175 ALV-negative (ALV−) individuals were identified. Based on 10× whole-genome resequencing and stringent quality control, 12,644,463 high-quality SNPs were obtained. GWAS revealed a significant enrichment of SNPs on chromosome 6 (Chr6), from which 218 SNPs significantly associated with ALV-J resistance and 49 candidate genes were identified. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that many of these genes, including PTPN13, TTF2, TIAL1, DLG2, FBXL7, CDH5, and CDH11, are involved in tumorigenesis and immunosuppression through the JAK/STAT signaling pathway and cell adhesion molecule pathways. Additionally, candidate genes, such as ANKH, SLC4A7, and SLC5A1, were found to potentially regulate ALV-J infection by modulating membrane transport and inflammatory responses. This study is the first to identify ALV-J resistance-associated genetic markers in Chengkou mountain chickens, revealing key genes related to immune regulation, membrane function, and tumor development. The findings provide a foundational molecular basis for disease-resistant breeding in poultry. Full article

This study investigates the immunological factors underlying the differential susceptibility of two chicken strains, E- and M-lines, to avian leukosis virus subgroup J (ALV-J). During the eradication of avian leukosis at a chicken breeder farm in Guangdong, we observed strain-specific differences in susceptibility to ALV-J. Moreover, E-line chickens exhibited a slower antibody response to ALV-J compared to M-line chickens. As the T cell receptor (TCR) and B cell receptor (BCR) are critical for antigen recognition, their activation triggers specific immune responses, including antibody production. Using high-throughput sequencing, we characterized the T cell receptor beta (TCRβ) and immunoglobulin heavy chain (IGH) repertoires in spleen tissues from both chicken strains. The M-line demonstrated higher clonal diversity in both TCRβ and IGH repertoires under normal conditions compared to the E-line, suggesting a broader baseline antigen recognition capacity. Following ALV-J infection, the TCRβ repertoire diversity remained unchanged, while the IGH repertoire displayed distinct clonal expansion patterns and complementarity-determining region 3 (CDR3) length distributions between the two lines, potentially affecting their ability to recognize ALV-J antigens. Our study provides the first comprehensive comparison of TCRβ and IGH repertoire dynamics in chickens with different ALV-J susceptibilities, offering new insights into the molecular and immunological mechanisms underlying resistance to ALV-J. Full article

Mycoplasma, reticuloendotheliosis virus (REV), avian leukosis virus (ALV), chicken infectious anemia virus (CIAV), bovine polyomavirus (BPV), bovine viral diarrhea virus (BVDV), and porcine circovirus (PCV) are considered common contaminants in live veterinary vaccines against Newcastle disease virus (NDV), fowlpox virus (FPV), infectious bursal disease virus (IBDV), classical swine fever virus (CSFV), pseudorabies virus (PRV), and porcine reproductive and respiratory syndrome virus (PRRSV). In the past five years, Getah virus (GETV), an arbovirus affecting many farming mammals, was reported as a new contaminant in live PRRSV vaccines in two previous studies, which arouses our considerable interest. Therefore, in this paper, we aim to analyze and discuss the source, biological hazard, and genomic characteristics of these contaminating GETV strains further. Full article

Viral co-infections pose significant challenges, causing substantial economic losses worldwide in the poultry industry. Among these, avian lLeukosis virus subgroup J (ALV-J) and chicken infectious anemia virus (CIAV) are particularly concerning, as they frequently lead to co-infections in chickens, further compromising their immune defenses, increasing susceptibility to secondary infections and diminishing vaccine efficacy. While our previous studies have examined the pathogenicity and immunosuppressive effects of these co-infections in vitro and in vivo, the key genes and molecular pathways involved remain largely unexplored. This study investigates the synergistic effects of co-infection with ALV-J and CIAV through comprehensive transcriptome analysis using high-throughput sequencing. We identified 1007 differentially expressed mRNAs (DEmRNAs) and 62 differentially expressed miRNAs (DEmiRNAs) associated with the synergistic activation effects of co-infection, along with 331 DEmRNAs and 62 DEmiRNAs linked to specific activation processes. Notably, the immune suppression observed in co-infected chickens may be influenced by the enhanced utilization of reactive oxygen species (ROS) and oxidative stress pathways, which impact host immune responses. Furthermore, co-infection appears to employ distinct immune evasion strategies through the modulation of rRNA metabolism, differing from single infections. These insights provide a deeper understanding of the molecular mechanisms underlying immune suppression during viral co-infections and help develop targeted therapies and improve disease control in poultry, reducing economic losses. Full article

In the realm of poultry production, viral superinfections pose significant challenges, causing substantial economic losses worldwide. Among these, avian leukosis virus subgroup J (ALV-J) and infectious bursal disease virus (IBDV) are particularly concerning as they frequently lead to superinfections in chicken, further exacerbating production losses and health complications. Our previous research delved into the pathogenicity and immunosuppressive effects of these superinfections through in vitro and in vivo analyses. Yet, the underlying key genes and pathways governing this phenomenon remained elusive. In this study, we randomly selected three chickens at 21 days post infection from each treatment group (ALV-J, IBDV, ALV-J+IBDV, and control group) to collect the bursa of Fabricius samples for full transcriptome analysis. Utilizing these data, we constructed a comprehensive circRNA/lncRNA-miRNA-mRNA network which elucidated both synergistic and specific activations during the superinfection. Notably, three pivotal genes (FILIP1L, DCX, and MYPN) were pinpointed in datasets reflecting synergistic activations. Conversely, four other genes (STAP, HKR6, XKR4, and TLR5) emerged in datasets associated with specific activations. Further exploration revealed diverse significant GO terms and pathways associated with both synergistic and distinct activation processes. These ceRNA network and core genes potentially wield substantial influence over the synergistic or specific activation of tumorigenesis and pathogenesis induced by ALV-J and IBDV. These findings could help develop targeted therapies and improve disease control in poultry, reducing economic losses. Full article

Avian Leukosis Virus (ALV) is a retrovirus that induces immunosuppression and tumor formation in poultry, posing a significant threat to the poultry industry. Currently, there are no effective vaccines or treatments for ALV. Therefore, the early diagnosis of infected flocks and farm sanitation are crucial for controlling outbreaks of this disease. To address the limitations of traditional diagnostic methods, which require sophisticated equipment and skilled personnel, a dual-tube detection method for ALV-J based on reverse transcription isothermal amplification (RAA) and the CRISPR-Cas13a system has been developed. This method offers the advantages of high sensitivity, specificity, and rapidity; it is capable of detecting virus concentrations as low as 5.4 × 10⁰ copies/μL without cross-reactivity with other avian viruses, with a total testing time not exceeding 85 min. The system was applied to 429 clinical samples, resulting in a positivity rate of 15.2% for CRISPR-Cas13a, which was higher than the 14.7% detected by PCR and 14.2% by ELISA, indicating superior detection capability and consistency. Furthermore, the dual-tube RAA-CRISPR detection system provides visually interpretable results, making it suitable for on-site diagnosis in remote farms lacking laboratory facilities. In conclusion, the proposed ALV-J detection method, characterized by its high sensitivity, specificity, and convenience, is expected to be a vital technology for purification efforts against ALV-J. Full article