OBJECTIVE: Salmonella enterica remains a major cause of food-borne disease in humans, and Salmonella Typhimurium (ST) contamination of poultry products is a worldwide problem. Since macrophages play an essential role in controlling Salmonella infection, the aim of this study was to evaluate the effect of glycyrrhizic acid (GA) on immune function of chicken HD11 macrophages. METHODS: Chicken HD11 macrophages were treated with GA (0, 12.5, 25, 50, 100, 200, 400, or 800 μg/ml) and lipopolysaccharide (LPS, 500 ng/ml) for 3, 6, 12, 24, or 48 h. Evaluated responses included phagocytosis, bacteria-killing, gene expression of cell surface molecules (cluster of differentiation 40 (CD40), CD80, CD83, and CD197) and antimicrobial effectors (inducible nitric oxide synthase (iNOS), NADPH oxidase-1 (NOX-1), interferon-γ (IFN-γ), LPS-induced tumor necrosis factor (TNF)-α factor (LITAF), interleukin-6 (IL-6), and IL-10), and production of nitric oxide (NO) and hydrogen peroxide (H₂O₂). RESULTS: GA increased the internalization of both fluorescein isothiocyanate (FITC)-dextran and ST by HD11 cells and markedly decreased the intracellular survival of ST. We found that the messenger RNA (mRNA) expression of cell surface molecules (CD40, CD80, CD83, and CD197) and cytokines (IFN-γ, IL-6, and IL-10) of HD11 cells was up-regulated following GA exposure. The expression of iNOS and NOX-1 was induced by GA and thereby the productions of NO and H₂O₂ in HD11 cells were enhanced. Notably, it was verified that nuclear factor-κB (NF-κB) and c-Jun N-terminal kinase (JNK) pathways were responsible for GA-induced synthesis of NO and IFN-γ gene expression. CONCLUSIONS Taken together, these results suggested that GA exhibits a potent immune regulatory effect to activate chicken macrophages and enhances Salmonella-killing capacity.
Animals ; Cells, Cultured ; Chickens ; Glycyrrhizic Acid/pharmacology* ; Macrophage Activation/drug effects* ; NF-kappa B/physiology* ; Phagocytosis/drug effects* ; Salmonella/drug effects* ; Signal Transduction/drug effects*

In order to observe the effect of the immune and weight of chickens after use the attenuated vaccine with low dose of chicken infectious anemia virus (CIAV). In this study, the effects of low dose of CIAV on the weight of SPF chickens and NDV antibody production were observed by simulated experiments. The results showed that 10 EID50 and 5 EID50 CIAV per plume attenuated NDV vaccines were used to cause the weight loss of SPF chickens. Compared with the use of the non contaminated vaccine group, it has significant difference. And NDV antibody levels compared with the use of the non contaminated groups also decreased after use the vaccine with two doses of CIAV contaminated. It has significant difference. A certain proportion of CIAV antibody positive was detected at the beginning of the second week after use the NDV vaccine with two doses of CIAV contaminated. The detection of a high proportion of CIAV nucleic acid was detected in the first week after the use of a contaminated vaccine. The results of the study demonstrate the effects of CIAV pollution on the production and immune function of SPF chickens, and it is suggested that increasing the detection of viral nucleic acid can help save time and improve the detection rate in the detection of exogenous virus contamination by SPF chicken test method.
Animals ; Antibodies, Viral ; immunology ; Chicken anemia virus ; genetics ; immunology ; physiology ; Chickens ; Circoviridae Infections ; immunology ; veterinary ; virology ; Poultry Diseases ; immunology ; virology ; Specific Pathogen-Free Organisms ; Vaccines, Attenuated ; administration & dosage ; genetics ; immunology

Avian leukosis virus subgroup J (ALV-J) is an avian retrovirus that can induce myelocytomas. A high-frequency mutation in gene envelope endows ALV-J with the potential for cross-species transmission. We wished to ascertain if the ALV-J can spread across species under selection pressure in susceptible and resistant hosts. First, we inoculated (in turn) two susceptible host birds (specific pathogen-free (SPF) chickens and turkeys). Then, we inoculated three resistant hosts (pheasants, quails and ducks) to detect the viral shedding, pathologic changes, and genetic evolution of different isolates. We found that pheasants and quails were infected under the selective pressure that accumulates stepwise in different hosts, and that ducks were not infected. Infection rates for SPF chickens and turkeys were 100% (16/16), whereas those for pheasants and quails were 37.5% (6/16) and 11.1% (3/27). Infected hosts showed immune tolerance, and inflammation and tissue damage could be seen in the liver, spleen, kidneys and cardiovascular system. Non-synonymous mutation and synonymous ratio (NS/S) analyses revealed the NS/S in hypervariable region (hr) 2 of pheasants and quails was 2.5. That finding suggested that mutation of isolates in pheasants and quails was induced by selective pressure from the resistant host, and that the hr2 region is a critical domain in cross-species transmission of ALV-J. Sequencing showed that ALV-J isolates from turkeys, pheasants and quails had moved away from the original virus, and were closer to the ALV-J prototype strain HPRS-103. However, the HPRS-103 strain cannot infect pheasants and quails, so further studies are needed.
Amino Acid Sequence ; Animals ; Avian Leukosis ; transmission ; virology ; Avian Leukosis Virus ; classification ; genetics ; physiology ; Chickens ; Ducks ; virology ; Galliformes ; virology ; Host Specificity ; Molecular Sequence Data ; Poultry Diseases ; transmission ; virology ; Quail ; virology ; Sequence Alignment ; Turkeys ; virology ; Viral Envelope Proteins ; chemistry ; genetics ; metabolism

Small interfering ribonucleic acid (siRNA)-induced RNA degradation can inhibit viral infection, and has been investigated extensively for its efficacy as antiviral therapy. The potential therapeutic role of lentiviral-mediated short hairpin ribonucleic acid (shRNA) to Newcastle disease virus (NDV) replication in vivo has been explored less often. We constructed two recombinant lentiviral vectors containing shRNA against the phosphoprotein (P) of the NDV, RNAi-341 and RNAi-671. Recombinant shRNA lentivirus vectors were co-transfected into 293T cells, along with helper plasmids, to package the recombinant shRNA lentivirus. Lentivirus-based shRNAs were titrated and transduced into NDV-susceptible chicken embryo fibroblasts (CEFs) and chick embryos. Antiviral activity against the NDV strain was evaluated by virus titration and real-time reverse transcription-polymerase chain reaction. RNAi-341 and RNAi-671 strongly suppressed transient expression of a FLAG-tagged P fusion protein in 293T cells. RNAi-341 and RNAi-671 NDV reduced virus titers by 66.6-fold and 30.6-fold, respectively, in CEFs 16 h after infection. RNAi-341 and RNAi-671 reduced virus titers in specific pathogen-free chick embryos by 99% and 98%, respectively, 48 h after infection. Both shRNAs inhibited accumulation of not only P-gene mRNA, but also nucleocapsid, M-, F-, HN-, and L-gene mRNA. RNAi-341 silenced P-gene mRNA more potently than RNAi-671. These results suggest that shRNAs silencing the P gene had substantial antiviral properties and inhibited NDV replication in CEFs and chick embryos.
Animals ; Chick Embryo ; Chickens ; Down-Regulation ; Fibroblasts ; virology ; Gene Targeting ; Lentivirus ; genetics ; metabolism ; Newcastle Disease ; virology ; Newcastle disease virus ; genetics ; physiology ; Phosphoproteins ; genetics ; metabolism ; Poultry Diseases ; virology ; RNA Interference ; RNA, Small Interfering ; genetics ; metabolism ; Viral Proteins ; genetics ; metabolism ; Virus Replication

Newcastle disease viruses (NDVs) cause systemic diseases in chickens with high mortality. However, little is known about persistence of NDVs in contaminated tissues from infected birds. In this study, we examined viral replication in the feather pulp of chickens inoculated with viscerotropic velogenic NDV (vvNDV) genotype VII. Reverse transcription real-time PCR and immunohistochemistry were used to investigate viral persistence in the samples. vvNDV was detected in the oropharynx and cloaca and viral antigens were detected in the feathers, suggesting that feathers act as sources of viral transmission.
Animals ; Antigens, Viral/analysis ; Chickens ; Cloaca/virology ; Feathers/*virology ; Microbial Viability ; Newcastle Disease/transmission/*virology ; Newcastle disease virus/isolation & purification/*physiology ; Oropharynx/virology ; Poultry Diseases/transmission/*virology ; Virus Replication/*physiology

We used a meq-deleted attenuated MDV-I strain GX0101Δmeq as a vector to construct a recombinant virus expressing the exogenous gene NDV-F. The ORF of exogenous gene NDV-F was inserted into the eukaryotic expression vector pcDNA3.1(-). Then, the expression cassette of NDV-F which contains the CMV promoter was amplified. Simultaneously, we amplified the selected gene Kan+ expression cassette and inserted them into the PMD18-T vector. Tandem expression cassettes were amplified using primers containing the 50-bp homologous arm of MDV-US2. The PCR product was electroporated into EL250 host bacteria containing GX0101Δmeq. Then, the Kan+ expression cassette was deleted from the recombinant virus genome using 1% arabinose. The plasmid of the positive clone which the Kan+ expression cassette was deleted was extracted and transfected into CEFs to rescue the recombinant virus. The recombinant virus was injected into chickens to observe its growth and replication. The recombinant virus rMDV-F containing the exogenous gene NDV-F was rescued successfully. The recombinant virus could duplicate and express well in CEFs, and grow and replicate well in chickens. Using GX0101Δmeq as a vector, combined with a recombinant system of Red E/T and FLP/FRT, we constructed a recombinant virus that expressed the exogenous gene NDV-F. This study could lay the foundation for further study of recombinant viruses.
Animals ; Cell Line ; Chickens ; virology ; DNA, Recombinant ; genetics ; Gene Expression ; Genetic Engineering ; Genetic Vectors ; genetics ; Mardivirus ; genetics ; physiology ; Plasmids ; genetics ; Viral Proteins ; genetics ; Virus Replication

Infectious bursal disease virus (IBDV) is an important member of the Birnaviridae family. IBUV mainly targets the bursa of Fabricius, the central immune organ of chicken, resulting in chicken infectious bursal disease (IBD). IBD represents one of the great challenges for ongoing development of the poultry industry. Reverse genetics for IBDV emerged over twenty years ago. Since then, the technologies behind virus rescue have continually improved leading to a deep understanding of IBDV gene function and tailored vaccine development. Our lab has also been instrumental in the field of IBDV research. Here we review studies on the pathogenic mechanism and the effective prevention and control of IBD.
Animals ; Birnaviridae Infections ; virology ; Chickens ; Infectious bursal disease virus ; genetics ; physiology ; Poultry Products ; virology ; Reverse Genetics

OBJECTIVETo study the changes in the levels of authophagy-related proteins ATG4A and p-ATG4A in nervous tissue after treated with tri-ortho-cresyl phosphate and explore the possible pathogenesis of OPIDN.

METHODSIn the first experiment, thirty hens were randomly divided into control group and 1 d, 5 d, 10 d and 21d treated groups, hens in treated groups were treated with TOCP by gavage at a single dosage of 600 mg/kg. In the second experiment, other thirty hens were also randomly divided into control group and 1 d, 5 d, 10 d and 21 d treated groups, hens in treated group were pretreated with PMSF by subcutaneous at a single dosage of 90 mg/kg. 24 h later, hens in intervention group was treated with TOCP by gavage at a single dosage of 600 mg/kg. The hens were killed at the corresponding time points, and collected their tibial nerves. The levels of ATG4A and p-ATG4A were measured by immunoblotting.

RESULTScompared with the control group, the levels of ATG4A decreased by36%, 43.7% and 41% at 1d, 5d and 10d in the intoxication groups (P < 0.05), the levels of p-ATG4A decreased by 22.5%, 25%and 21%at 1d, 5d and 10d in the intoxication group (P < 0.05). However, compared with the control group, there is no significant change in the levels of ATG4A and p-ATG4A in PMSF-pretreated groups.

CONCLUSIONThe intoxication of TOCP influence the levels of autophagy-related proteins ATG4A and p-ATG4A, which might be associated with the inhibition of autophagy activity in neurons of OPIDN.

Animals ; Apoptosis Regulatory Proteins ; metabolism ; Autophagy ; drug effects ; Chickens ; Female ; Nerve Tissue ; physiology ; Phosphorylation ; drug effects ; Tibial Nerve ; Tritolyl Phosphates ; toxicity

In 2014, two genetically distinct H5N8 highly pathogenic avian influenza (HPAI) viruses were isolated from poultry and wild birds in Korea. The intravenous pathogenicity indices for the two representative viruses were both 3.0. Mortality of chickens intranasally inoculated with the two H5N8 viruses was 100% with a mean death times of 2.5 and 4.5 days. Mortality rates of the contact groups for the two H5N8 viruses were 33.3% and 66.6%. Our study showed that transmissibility of the novel H5N8 viruses was different from that of previously identified H5N1 HPAI viruses, possibly due to genetic changes.
Animals ; Chickens ; Disease Outbreaks/veterinary ; Influenza A virus/*pathogenicity/*physiology ; Influenza in Birds/*mortality/transmission/virology ; Poultry Diseases/*mortality/transmission/virology ; Republic of Korea/epidemiology ; Virulence

Salmonella enterica Gallinarum (SG) causes fowl typhoid (FT), a septicemic disease in avian species. We constructed deletion mutants lacking the stress sigma factor RpoS, the nitric oxide (NO)-detoxifying flavohemoglobin Hmp, and the SsrA/SsrB regulator to confirm the functions of these factors in SG. All gene products were fully functional in wild-type (WT) SG whereas mutants harboring single mutations or a combination of rpoS, hmp, and ssrAB mutations showed hypersusceptibility to H2O2, loss of NO metabolism, and absence of Salmonella pathogenicity island (SPI)-2 expression, respectively. A triple-deletion mutant, SGDelta3 (SGDeltarpoSDeltahmpDeltassrAB), was evaluated for attenuated virulence and protection efficacy in two-week-old Lohmann layer chickens. The SGDelta3 mutant did not cause any mortality after inoculation with either 1 x 10(6) or 1 x 10(8) colony-forming units (CFUs) of bacteria. Significantly lower numbers of salmonellae were recovered from the liver and spleen of chickens inoculated with the SGDelta3 mutant compared to chickens inoculated with WT SG. Vaccination with the SGDelta3 mutant conferred complete protection against challenge with virulent SG on the chickens comparable to the group vaccinated with a conventional vaccine strain, SG9R. Overall, these results indicate that SGDelta3 could be a promising candidate for a live Salmonella vaccine against FT.
Administration, Oral ; Animals ; Bacterial Proteins/*genetics/immunology ; *Chickens ; Female ; Poultry Diseases/*immunology/microbiology ; Salmonella Infections, Animal/*immunology/microbiology ; Salmonella Vaccines/administration & dosage/genetics/*immunology ; Salmonella enterica/immunology/*physiology ; Vaccines, Attenuated/administration & dosage/genetics/immunology ; Virulence