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Abstract Background Recent epidemiological investigation of different HA subtypes of avian influenza viruses (AIVs) shows that the H3 subtype is the most predominant among low pathogenic AIVs (LPAIVs), and the seasonal variations in isolation of H3 subtype AIVs are consistent with that of human H3 subtype influenza viruses. Consequently, the development of a rapid, simple, sensitive detection method for H3 subtype AIVs is required. The loop-mediated isothermal amplification (LAMP) assay is a simple, rapid, sensitive and cost-effective nucleic acid amplification method that does not require any specialized equipment. Results A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed to detect the H3 subtype AIVs visually. Specific primer sets target the sequences of the hemagglutinin (HA) gene of H3 subtype AIVs were designed, and assay reaction conditions were optimized. The established assay was performed in a water bath for 50 minutes, and the amplification result was visualized directly as well as under ultraviolet (UV) light reflections. The detection limit of the RT-LAMP assay was 0.1pg total RNA of virus, which was one hundred-fold higher than that of RT-PCR. The results on specificity indicated that the assay had no cross-reactions with other subtype AIVs or avian respiratory pathogens. Furthermore, a total of 176 clinical samples collected from birds at the various live-bird markets (LBMs) were subjected to the H3-subtype-specific RT-LAMP (H3-RT-LAMP). Thirty-eight H3 subtype AIVs were identified from the 176 clinical samples that were consistent with that of virus isolation. Conclusions The newly developed H3-RT-LAMP assay is simple, sensitive, rapid and can identify H3 subtype AIVs visually. Consequently, it will be a very useful screening assay for the surveillance of H3 subtype AIVs in underequipped laboratories as well as in field conditions.

The present study was conducted to determine whether avian reovirus (ARV) activates the phosphatidylinositol 3-kinase-dependent Akt (PI3K/Akt) pathway according to the PXXP or YXXXM motifs of σA and σNS proteins. Gene splicing by overlap extension PCR was used to change the PXXP or YXXXM motifs of the σA and σNS genes. Plasmid constructs that contain mutant σA and σNS genes were generated and transfected into Vero cells, and the expression levels of the corresponding genes were quantified according to immunofluorescence and Western blot analyses. The Akt phosphorylation (P-Akt) profile of the transfected Vero cells was examined by flow cytometry and Western blot. The results showed that the σA and σNS genes were expressed in the Vero cells, and P-Akt expression in the σA mutant groups (amino acids 110–114 and 114–117) was markedly decreased. The results indicated that the σA protein of ARV activates the PI3K/Akt pathway via the PXXP motif. The results of this study reveal the mechanisms by which ARV manipulates the cellular signal transduction pathways, which may provide new ideas for novel drug targets. [ABSTRACT FROM AUTHOR]

`[Objective] The paper was to construct eukaryotic expression vector of Avian reovirus (ARV)σA gene and express σA protein accurately in HKK293T cells. [Methodl The specific primers of ARV σA gene were designed according to the gene sequence of ARV S2 gene in GenBank (accession number KF741763.1). With pMD18-T-σA recombinant vector as the template, the specific sequence of aA gene was amplified by PCR and cloned into pMD18-T vector to construct recombinant plasmid. The cloning vector pMD18-T-σA and eukaryotic expression plasmid pKKla-HA were double digested by restriction enzymes Kpn I and iNol I. The purified

To date, nine neuraminidase (NA) subtypes of avian influenza virus (AIV) have been identified in poultry and wild birds. Rapid and effective methods for differentiating these nine NA subtypes are needed. We developed and validated a rapid, sensitive, and robust method utilizing a GeXP analyzer-based multiplex RT-PCR assay and capillary electrophoresis for the simultaneous differentiation of the N1 to N9 subtypes in a single-tube reaction. Ten pairs of primers–nine subtype-specific pairs and one pan-AIV pair–were screened and used to establish the GeXP multiplex RT-PCR assay. A single subtype was detected using the developed GeXP assay; the N1 to N9 AIV subtypes individually generated two target peaks: the NA subtype-specific peak and the general AIV peak. Different concentrations of multiplexed subtypes were tested with this GeXP assay and the peaks of the corresponding NA subtypes were generated, suggesting that this GeXP assay is useful for identifying NA subtypes in mixed samples. Moreover, no peaks were generated for other important avian viruses, indicating negative results and validating the lack of cross-reactions between AIV subtypes and other avian pathogens. RNA templates synthesized through in vitro transcription were used to analyze the sensitivity of the assay; the limit of detection was 100 copies per reaction mixture. The results obtained from clinical samples using this GeXP method were consistent with the results of the neuraminidase inhibition (NI) test, and the ability of the GeXP assay to identify mixed infections was superior to amplicon sequencing of isolated viruses. In conclusion, this GeXP assay is proposed as a specific, sensitive, rapid, high-throughput, and versatile diagnostic tool for nine NA subtypes of AIV. [ABSTRACT FROM AUTHOR]

Foot-and-mouth disease virus (FMDV), Bluetongue virus (BTV), Vesicular stomatitis Virus (VSV), Bovine viral diarrheal (BVDV), Bovine rotavirus (BRV), and Bovine herpesvirus 1 (IBRV) are common cattle infectious viruses that cause a great economic loss every year in many parts of the world. A rapid and high-throughput GenomeLab Gene Expression Profiler (GeXP) analyzer-based multiplex PCR assay was developed for the simultaneous detection and differentiation of these six cattle viruses. Six pairs of chimeric primers consisting of both the gene-specific primer and a universal primer were designed and used for amplification. Then capillary electrophoresis was used to separate the fluorescent labeled PCR products according to the amplicons size. The specificity of GeXP-multiplex PCR assay was examined with samples of the single template and mixed template of six viruses. The sensitivity was evaluated using the GeXP-multiplex PCR assay on serial 10-fold dilutions of ssRNAs obtained via in vitro transcription. To further evaluate the reliability, 305 clinical samples were tested by the GeXP-multiplex PCR assay. The results showed that the corresponding virus specific fragments of genes were amplified. The detection limit of the GeXP-multiplex PCR assay was 100 copies/μL in a mixed sample of ssRNAs containing target genes of six different cattle viruses, whereas the detection limit for the Gexp-mono PCR assay for a single target gene was 10 copies/μL. In detection of viruses in 305 clinical samples, the results of GeXP were consistent with simplex real-time PCR. Analysis of positive samples by sequencing demonstrated that the GeXP-multiplex PCR assay had no false positive samples of nonspecific amplification. In conclusion, this GeXP-multiplex PCR assay is a high throughput, specific, sensitive, rapid and simple method for the detection and differentiation of six cattle viruses. It is an effective tool that can be applied for the rapid differential diagnosis of clinical samples and for epidemiological investigation. [ABSTRACT FROM AUTHOR]

Objectives In order to develop a multiplex RT- PCR assay using the Ge XP analyser for the simultaneous detection of four different NA serotypes of H5-subtype AIVs, effective to control and reduce H5 subtype of avian influenza outbreak. Design Six pairs of primers were designed using conserved and specific sequences of the AIV subtypes H5, N1, N2, N6 and N8 in GenBank. Each gene-specific primer was fused at the 5′ end to a universal sequence to generate six pairs of chimeric primers, and one pair of universal primers was used for RT- PCR, and PCR product separation and detection were performed by capillary electrophoresis using the GenomeLab Ge XP genetic analysis system. Setting Single and mixed avian pathogen cDNA/ DNA templates were employed to evaluate the specificity of a multiplex assay with a Ge XP analyser. Corresponding specific DNA products were amplified for each gene, revealing amplification peaks for M, H5, N1, N2, N6 and N8 genes from four different NA subtypes of influenza A H5 virus. Sample A total of 180 cloacal swabs were collected from poultry at live bird markets. Main outcome measures The multiplex PCR assay demonstrated excellent specificity, with each pair of specific primers generating only products corresponding to the target genes and without cross-amplification with other NA-subtype influenza viruses or other avian pathogens. Using various premixed ss RNAs containing known AIV target genes, the detection limit for the multiplex assay was determined to be 102 copies/μl. The Ge XP assay was further evaluated using 180 clinical specimens and compared with RRT- PCR (real-time reverse transcriptase PCR) and virus isolation. Conclusions This Ge XP analyser-based multiplex assay for four different NA subtypes of H5 HPAI viruses is both highly specific and sensitive and can be used as a rapid and direct diagnostic assay for testing clinical samples. [ABSTRACT FROM AUTHOR]

We report a novel electrochemical immunosensor that can sensitively detect avian influenza virus H5 subtype (AIV H5) captured by graphene oxide-H5-polychonal antibodies-bovine serum albumin (GO-PAb-BSA) nanocomposite. The graphene oxide (GO) carried H5-polychonal antibody (PAb) were used as signal amplification materials. Upon signal amplification, the immunosensor showed a 256-fold increase in detection sensitivity compared to the immunosensor without GO-PAb-BSA. We designed a PAb labeling GO strategy and signal amplification procedure that allow ultrasensitive and selective detection of AIV H5. The established method responded to 2−15 HA unit/50 µL H5, with a linear calibration range from 2−15 to 2−8 HA unit/50 µL. In summary, we demonstrated that the immunosenser has a high specificity and sensitivity for AIV H5, and the established assay could be potentially applied in the rapid detection of other pathogenic microorganisms. [ABSTRACT FROM AUTHOR]

Antibodies specific to the non-structural proteins of viruses are detected in virus-infected animals and show promise as a reliable diagnostic marker for virus infections. We examined the potential use of two non-structural proteins of fowl adenovirus (FAdV)-based, 100K and 33K, enzyme-linked immunosorbent assays (ELISAs) in the diagnosis of FAdVs. We cloned and expressed the 100K and 33K non-structural protein genes of the FAdVs in the pGEX-4T-1 plasmid vector. Purified 100K and 33K proteins alone or in combination were used as antigens in ELISAs. Antibodies specific to the 100K and 33K non-structural proteins were detected in chickens experimentally infected with FAdVs, but not in chickens vaccinated with inactivated FAdVs. In contrast, the agar gel precipitation (AGP) test detected FAdV-specific antibodies in 70.3% of the vaccinated chickens, suggesting that the non-structural protein-based ELISA could be used in the differential diagnosis of infected and vaccinated chickens. To further validate the 100K and 33K-based ELISA (100K-33K-ELISA) method, we compared its sensitivity and specificity with that of a whole virus-based ELISA and an AGP test in detecting FAdV-specific antibodies in 350 field samples. The results showed that the 100K-33K-ELISA exhibited a higher sensitivity than the AGP test and a comparable sensitivity and specificity to the whole virus ELISA. Overall, the 100K-33K-ELISA method is sensitive, specific and can be used to distinguish an acute FAdV infection from an inactivated virus-based vaccination response. [ABSTRACT FROM PUBLISHER]

Avian reovirus (ARV) is an important pathogen of poultry and causes significant economic losses to the poultry industry. To develop a rapid and sensitive method for the surveillance of ARV, a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was established using a set of six primers specific to the S1 gene segment of ARV. The established assay was performed at 62oC for 60 min in a thermal block, and the result was visualized directly under daylight or ultraviolet light. The detection limit of the RT-LAMP assay was 10 fg total RNA, which was 100-fold higher than that of reverse transcriptase polymerase chain reactions. The specificity of the assay was supported by the lack of cross-reaction with other avian pathogens. Furthermore, viral RNAs of field isolates were successfully detected by the assay. Overall, the newly established RT-LAMP assay is simple, rapid, sensitive, specific, and can visually detect ARV without the use of any specialized equipment. [ABSTRACT FROM PUBLISHER]

The complete mitochondrial genome sequence of the GuangxiPhasianus colchicuswas measured by PCR-based methods and analyzed in detail. Our research findings reveal that the entire mitochondrial genome ofP. colchicusis a circular molecule of length 16,687 bp (GenBank accession number: KT364526). The contents of A, T, C, and G in the mitochondrial genome were found to be 30.64%, 25.29%, 30.81% and 13.26%, respectively. The complete mitochondrial genome of theP. colchicusis a typical structure, including 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, and 1 control region (D-loop region). This complete mitochondrial genome sequence provides essential information in understanding phylogenetic relationships among Galliformes mitochondrial genomes. [ABSTRACT FROM AUTHOR]

The complete mitochondrial genome sequence of the Cenxi classical three-buff chicken was measured by PCR-based methods and analyzed in detail. Our research findings reveal that the entire mitochondrial genome of the Cenxi classical three-buff chicken is a circular molecule consisting of 16,786 bp (GenBank accession number: KM433666). The contents of A, T, C, and G in the mitochondrial genome were found to be 30.27%, 23.75%, 32.49% and 13.49%, respectively. The complete mitochondrial genome of the Cenxi classical three-buff chicken contains a typical structure, including 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes and 1 control region (D-loop region). This complete mitochondrial genome sequence provides essential information in understanding phylogenetic relationships amongGallus gallus domesticusmitochondrial genomes. [ABSTRACT FROM AUTHOR]

The complete mitochondrial genome sequence of the Luchuan pig was measured by PCR-based methods, primer-walking sequencing, and fragment cloning. The entire mitochondrial genome of the Luchuan pig was identified as a circular molecule consisting of 16,730 bp (GenBank accession number: KP126954). The contents of A, T, C, and G in the mitochondrial genome were found to be 34.65%, 25.80%, 26.19%, and 13.36%, respectively. The complete mitochondrial genome of the Luchuan pig contains a typical structure, including 13 protein-coding genes, two rRNA genes, 22 tRNA genes, and one control region (D-loop region). This complete mitochondrial genome sequence provides essential information in understanding phylogenetic relationships amongSus scrofadomestic mitochondrial genomes. [ABSTRACT FROM AUTHOR]

The objective of this study was to obtain the complete mitochondrial DNA sequence of chukar partridge, and to provide reference data for protection and utilization of these resources of chukar partridge. The complete mitochondrial genome sequence of the China chukar partridge was measured by PCR-based methods and analysed in detail. Our research findings reveal that the entire mitochondrial genome of the chukar partridge is a circular molecule consisting of 16,688 bp (GenBank accession number: KY829450). The contents of A, T, C, and G in the mitochondrial genome were found to be 30.44%, 24.43%, 31.57%, and 13.56%, respectively. The complete mitochondrial genome of the chukar partridge has a typical structure, including 13 protein-coding genes, two rRNA genes, 22 tRNA genes, and one control region (D-loop region). This complete mitochondrial genome sequence provides essential information in understanding phylogenetic relationships among Galliformes mitochondrial genomes. [ABSTRACT FROM AUTHOR]

The complete mitochondrial genome sequence of the Rongshui Xiang duck was measured by PCR-based methods. Our research findings reveal that the entire mitochondrial genome of the Rongshui Xiang duck is a circular molecule consisting of 16,605 bp (GenBank accession number: KJ833587). The contents of A, T, C, and G in the mitochondrial genome were found to be 29.20%, 22.20%, 32.82% and 15.79%, respectively, similar to the majority of avian species. The complete mitochondrial genome of the Rongshui Xiang duck contains 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, and 1 control region. The characteristics of the mitochondrial genome were analyzed in detail. Our complete mitochondrial genome sequence should provide essential information for understanding phylogenetic relationships among duck mitochondrial genomes. [ABSTRACT FROM PUBLISHER]

The complete mitochondrial genome sequence of the Xilin small partridge duck was measured by PCR-based methods. Our research findings revealed that the entire mitochondrial genome of the Xilin small partridge duck was 16,604 bp (GenBank accession number: KJ833586). The contents of A, T, C, and G in the mitochondrial genome were 29.20%, 22.19%, 32.82% and 15.79%, respectively, which were similar to the majority of most avian species. The complete mitochondrial genome of the Xilin small partridge duck contains 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, and 1 control region. Components of the Xilin small partridge duck’s mitochondrial genome were similar to those of otherAnatidaein gene arrangement and composition. The complete mitochondrial genome of the Xilin small partridge duck should provide essential information for understanding phylogenetic relationships of duck mitochondrial genome. [ABSTRACT FROM PUBLISHER]

The complete mitochondrial genome sequence of the Longsheng duck was measured by PCR-based methods. Our research findings revealed that the entire mitochondrial genome of the Longsheng duck was 16,603 bp (GenBank accession number: KJ739616). The contents of A, T, C, and G in the mitochondrial genome were 29.22%, 22.21%, 32.79% and 15.77%, respectively, which were similar to the majority of most avian species. The complete mitochondrial genome of the Longsheng duck contains 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, and one control region. Components of the Longsheng duck’s mitochondrial genome were similar to those of otherAnas platyrhynchosin gene arrangement and composition. The complete mitochondrial genome of the Longsheng duck should provide essential information for understanding phylogenetic relationships of duck mitochondrial genome. [ABSTRACT FROM AUTHOR]

The entire mitochondrial genome of Jingxi duck from China was 16,603 bp in length, and has been analyzed for gene locations, length, start codons and stop codons. With the base composition of 29.20% for A, 22.19% for T, 32.82% for C, 15.79% for G, so the percentage of A and T (51.39%) was slightly higher than those of G and C. The Jingxi duck mitochondrial genome contained two ribosomal RNA genes, 13 protein-coding genes, 22 transfer RNA genes and one non-coding control region (D-loop region). The arrangement of these genes was the same as most birds. The complete mitochondrial genome sequence of the Jingxi duck will be useful for phylogenetics, and provide an important data set for further study on the germplasm resources. [ABSTRACT FROM AUTHOR]

• Four host proteins interacting with ARV σA protein were screened by a yeast two-hybrid system. • NME2 was further validated as a σA-binding protein through co-immunoprecipitation. • The key interaction domain was identified at aa 121–416 in NME2 and at aa 71–139 in σA, respectively. • Overexpression of NME2 substantially inhibited ARV replication. Our present study aimed to identify host cell proteins that may interact with avian reovirus (ARV) σA protein and their potential effect on ARV replication. The ARV structural protein σA has been demonstrated to suppress interferon production and confirmed to activate the PI3K/Akt pathway. However, host cell factors interacting with σA to affect ARV replication remain unknown. In current study, a cDNA library of chicken embryo fibroblasts (CEFs) was constructed, and host cell proteins interacting with σA were screened by a yeast two-hybrid system. We identified four candidate cellular proteins that interact with ARV σA protein. Among them, Gallus NME/NM23 nucleoside diphosphate kinase 2 (NME2) was further validated as a σA-binding protein through co-immunoprecipitation. The key interaction domain was identified at amino acids (aa) 121–416 in NME2 and at aa 71–139 in σA, respectively. We demonstrated that overexpression of NME2 substantially inhibited ARV replication. In addition silencing NME2 by small interfering RNAs (siRNAs) resulted in marked enhancement of ARV replication. Our work has demonstrated that NME2 is a σA-binding protein that may affect ARV replication in CEF cells. [ABSTRACT FROM AUTHOR]

Starting in 2015, the widespread prevalence of hydropericardium-hepatitis syndrome (HHS) has led to considerable financial losses within China's poultry farming industry. In this study, pathogenicity assessments, whole-genome sequencing, and analyses were conducted on 10 new isolates of the novel genotype FAdV-4 during a HHS outbreak in Guangxi Province, China, from 2019 to 2020. The results indicated that strains GX2019-010 to GX2019-013 and GX2019-015 to GX2019-018 were highly virulent, while strain GX2020-019 exhibited moderate virulence. Strain GX2019-014 was characterized as a wild-type strain with low virulence, displaying no pathogenic effects when 0.5 mL containing 10 6 TCID 50 virus was inoculated into the muscle of specific pathogen-free (SPF) chickens at 4 weeks of age, while 10 7 TCID 50 and 10 8 TCID 50 resulted in mortality rates of 80 and 100%, respectively. The whole genomes of strains GX2019-010 to GX2019-013, GX2019-015 to GX2019-018, and GX2020-019 showed high homology with other Chinese newly emerging highly pathogenic FAdV-4 strains, whereas GX2019-014 was closer to nonmutant strains and shared the same residues with known nonpathogenic strains (B1-7, KR5, and ON1) at positions 219AA and 380AA of the Fiber-2 protein. Our work enriches the research on prevalent strains of FAdV-4 in China, expands the knowledge on the virulence diversity of the novel genotype FAdV-4, and provides valuable reference material for further investigations into the key virulence-associated genetic loci of FAdV-4., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Wei, Xie, Xie, Deng, Li, Xie, Fan, Zhang, Wang, Ren, Wan, Luo and Li.)

The GX2020-019 strain of fowl adenovirus serotype 4 (FAdV-4) was isolated from the liver of chickens with hydropericardium hepatitis syndrome in Guangxi Province, China, and was purified by plaque assay three times. Pathogenicity studies showed that GX2020-019 can cause typical FAdV-4 pathology, such as hydropericardium syndrome and liver yellowing and swelling. Four-week-old specific pathogen-free (SPF) chickens inoculated with the virus at doses of 10 3 median tissue culture infectious dose (TCID 50 ), 10 4 TCID 50 , 10 5 TCID 50 , 10 6 TCID 50 , and 10 7 TCID 50 had mortality rates of 0, 20, 60, 100, and 100%, respectively, which were lower than those of chickens inoculated with other highly pathogenic Chinese isolates, indicating that GX2020-019 is a moderately virulent strain. Persistent shedding occurred through the oral and cloacal routes for up to 35 days postinfection. The viral infection caused severe pathological damage to the liver, kidney, lung, bursa of Fabricius, thymus, and spleen. The damage to the liver and immune organs could not be fully restored 21 days after infection, which continued to affect the immune function of chickens. Whole genome analysis indicated that the strain belonged to the FAdV-C group, serotype 4, and had 99.7-100% homology with recent FAdV-4 strains isolated from China. However, the amino acid sequences encoded by ORF30 and ORF49 are identical to the sequences found in nonpathogenic strains, and none of the 32 amino acid mutation sites that appeared in other Chinese isolates were found. Our research expands understanding of the pathogenicity of FAdV-4 and provides a reference for further studies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Wei, Xie, Fan, Xie, Deng, Luo, Li, Zhang, Zeng, Huang, Ruan and Wang.)

Avian reovirus (ARV) infection can lead to severe immunosuppression, complications, and secondary diseases, causing immense economic losses to the poultry industry. In-depth study of the mechanism by which the innate immune system combats ARV infection, especially the antiviral effect mediated by interferon, is needed to prevent and contain ARV infection. In this study, ARV strain S1133 was used to artificially infect 7-day-old specific pathogen-free chickens. The results indicated that ARV rapidly proliferated in the immune organs, including the spleen, bursa of Fabricius, and thymus. The viral load peaked early in the infection and led to varying degrees of pathological damage to tissues and organs. Real-time quantitative PCR revealed that the mRNA levels of interferon and multiple interferon-stimulated genes (ISGs) in the spleen, bursa of Fabricius, and thymus were upregulated to varying degrees in the early stage of infection. Among the ISGs, IFIT5, and Mx were the most upregulated in various tissues and organs, suggesting that they are important ISGs for host resistance to ARV infection. Further investigation of the role of IFIT5 in ARV infection showed that overexpression of the IFIT5 gene inhibited ARV replication, whereas inhibition of the endogenously expressed IFIT5 gene by siRNA promoted ARV replication. IFIT5 may be a positive feedback regulator of the innate immune signaling pathways during ARV infection and may induce IFN-α production by promoting the expression of MAD5 and MAVS to exert its antiviral effect. The results of this study help explain the innate immune regulatory mechanism of ARV infection and reveal the important role of IFIT5 in inhibiting ARV replication, which has important theoretical significance and practical application value for the prevention and control of ARV infection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wang, Wan, Ren, Xie, Xie, Huang, Deng, Xie, Luo, Li, Zeng, Zhang and Zhang.)

Background: Chicken anemia virus (CAV) causes chicken infectious anemia, which results in immunosuppression; the virus has spread widely in chicken flocks in China., Objectives: The aim of this study was to understand recent CAV genetic evolution in chicken flocks in Guangxi Province, southern China., Methods: In total, 350 liver samples were collected from eight commercial broiler chicken farms in Guangxi Province in southern China from 2018 to 2020. CAV was detected by conventional PCR, and twenty CAV complete genomes were amplified and used for the phylogenetic analysis and recombination analysis., Results: The overall CAV-positive rate was 17.1%. The genetic analysis revealed that 84 CAVs were distributed in groups A, B, C (subgroups C1-C3) and D. In total, 30 of 47 Chinese CAV sequences from 2005-2020 belong to subgroup C3, including 15 CAVs from this study. There were some specific mutation sites among the intergenotypes in the VP1 protein. The amino acids at position 394Q in the VP1 protein of 20 CAV strains were consistent with the characteristics of a highly pathogenic strain. GX1904B was a putative recombinant., Conclusions: Subgroup C3 was the dominant genotype in Guangxi Province from 2018-2020. The 20 CAV strains in this study might be virulent according to the amino acid residue analysis. These data help improve our understanding of the epidemiological trends of CAV in southern China., Competing Interests: The authors declare no conflicts of interest., (© 2022 The Korean Society of Veterinary Science.)

Avian reovirus (ARV) can induce many diseases as well as immunosuppression in chickens, severely endangering the poultry industry. Interferons (IFNs) play an antiviral role by inducing the expression of interferon-stimulated genes (ISGs). The effect of ARV infection on the expression of host ISGs is unclear. Specific-pathogen-free (SPF) chickens were infected with ARV strain S1133 in this study, and real time quantitative PCR was used to detect changes in the dynamic expression of IFNs and common ISGs in joints of SPF chickens. The results showed that the transcription levels of IFN A, IFNB , and several ISGs, including myxovirus resistance ( MX ), interferon-induced transmembrane protein 3 ( IFITM3 ), protein kinase R ( PKR ), oligoadenylate synthase ( OAS ), interferon-induced protein with tetratricopeptide repeats 5 ( IFIT5 ), interferon-stimulated gene 12 ( ISG12 ), virus inhibitory protein ( VIPERIN ), interferon-alpha-inducible protein 6 ( IFI6 ), and integrin-associated protein ( CD47 ), were upregulated in joints on days 1-7 of infection (the levels of increase of MX, IFIT5, OAS, VIPERIN, ISG12 , and IFI6 were the most significant, at hundreds-fold). In addition, the expression levels of the ISGs encoding zinc finger protein 313 ( ZFP313 ), and DNA damage-inducible transcript 4 ( DDIT4 ) increased suddenly on the 1st or 2nd day, then decreased to control levels. The ARV viral load in chicken joints rapidly increased after 1 day of viral challenge, and the viral load remained high within 6 days of viral challenge. The ARV viral load sharply decreased starting on day 7. These results indicate that in SPF chicken joints, many ISGs have mRNA expression patterns that are basically consistent with the viral load in joints. IFNA, IFNB, and the ISGs MX, IFITM3, PKR, OAS, IFIT5, ISG12, VIPERIN, IFI6, and CD47 play important roles in defending against ARV invasion, inhibiting ARV replication and proliferation, and promoting virus clearance. These results enrich our understanding of the innate immune response mechanisms of hosts against ARV infection and provide a theoretical basis for prevention and control of ARV infection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Wang, Xie, Xie, Wan, Huang, Deng, Xie, Luo, Zeng, Zhang, Zhang and Zhou.)

A previously unidentified chicken parvovirus (ChPV) and turkey parvovirus (TuPV) strain, associated with runting-stunting syndrome (RSS) and poultry enteritis and mortality syndrome (PEMS) in turkeys, is now prevalent among chickens in China. In this study, a large-scale surveillance of parvoviruses in chickens and turkeys using conserved PCR assays was performed. We assessed the prevalence of ChPV/TuPV in commercial chicken and turkey farms in China between 2014 and 2019. Parvoviruses were prevalent in 51.73% (1,795/3,470) of commercial chicken and turkey farms in Guangxi, China. The highest frequency of ChPV positive samples tested by PCR occurred in chickens that were broiler chickens 64.18% (1,041/1,622) compared with breeder chickens 38.75% (572/1,476) and layer hens 38.89% (112/288), and TuPV was detected in 70/84 (83.33%). Native and exotic chicken species were both prevalent in commercial farms in southern China, and exotic broiler chickens had a higher positive rate with 88.10% (148/168), while native chickens were 50.00% (1,465/2,930). The environmental samples from poultry houses tested positive for ChPV and TuPV were 47.05% (415/874). Samples from open house flocks had higher prevalence rates of ChPV than those of closed house flocks ( Table 5 ), among which those from the open house showed 84.16% (85/101) positivity, those from litter showed 62.86% (44/70) positivity, and those from drinking water showed 50.00% (56/112) positivity, whereas those from the closed house litter were 53.57% (60/112), those from swabs were 50.18% (138/275), and those from drinking water were 15.69% (32/204). Samples collected during spring were more frequently ChPV/ TuPV positive than those collected during other seasons. This study is the first report regarding the epidemiological surveillance of ChPV and TuPV in chicken/turkey flocks in Guangxi, China. Our results suggest that ChPV and TuPV are widely distributed in commercial fowl in Guangxi. These findings highlight the need for further epidemiological and genetic research on ChPV and TuPV in this area., (Copyright © 2020 Zhang, Feng, Xie, Deng, Zhang, Xie, Xie, Fan, Luo, Zeng, Huang and Wang.)

Objective To prepare the monoclonal antibodies (mAb) against hemagglutinin of H4 subtype avian influenza virus (AIV), and develop a sandwich ELISA for the detection of H4 subtype AIV. Methods The BALB/c mice were immunized with inactive H4 subtype AIV. A mAb against H4 subtype AIV, designated as 6G4, was obtained by cell fusion, hemagglutination inhibition (HI) screening and subcloing. Immuofluorescence cytochemistry and Western blotting were used to detect the reactivity of 6G4 with H4 subtype AIV, and the specificity, broad spectrum and stability of 6G4 were characterized by HI assay. Subclass of 6G4 was determined by kit. With chicken polyclonal antibody against H4 subtype AIV as coated antibody, 6G4 mAb as capture antibody and HRP-labeled goat anti-mouse IgG as the enzyme-labeled antibody, a sandwich ELISA for the detection of H4 subtype AIV was established by optimization of the reaction conditions and serial verification. Results 6G4 belonged to IgG1 subclass, and the light chain belonged to κ. It could secrete antibody stably and had good reactivity, specificity, broad spectrum and stability. ELISA based on 6G4 was specific, sensitive, accurate and suitable for the detection of a large number of samples. Conclusion We successfully achieved the anti-H4 subtype AIV mAb, and developed the sandwich ELISA for the detection of H4 subtype AIV.

The aim of the current study was to determine the genomic sequence of parvovirus strain GX-Tu-PV-1, which was isolated from a turkey in Guangxi Province, South China. The analysis showed that the genome sequence of GX-Tu-PV-1 was 81.3% to ∼99.3% similar to those of other turkey parvoviruses (TuPVs) and 79.8% to ∼92.1% related to chicken parvovirus (ChPV). This study will help in understanding the epidemiology and molecular characteristics of parvovirus in turkeys., (Copyright © 2019 Zhang et al.)

We report here the complete genome sequence of strain H9N2, an avian influenza virus (AIV) isolated from dove in Guangxi, China. Phylogenetic analysis showed that it was a novel reassortant AIV derived from chicken, duck, and wild bird. This finding provides useful information for understanding the H9N2 subtype of AIV circulating in southern China., (Copyright © 2018 Li et al.)

Recent studies have demonstrated that at least eight subtypes of avian influenza virus (AIV) can infect humans, including H1, H2, H3, H5, H6, H7, H9 and H10. A GeXP analyser-based multiplex reverse transcription (RT)-PCR (GeXP-multiplex RT-PCR) assay was developed in our recent studies to simultaneously detect these eight AIV subtypes using the haemagglutinin (HA) gene. The assay consists of chimeric primer-based PCR amplification with fluorescent labelling and capillary electrophoresis separation. RNA was extracted from chick embryo allantoic fluid or liquid cultures of viral isolates. In addition, RNA synthesised via in vitro transcription was used to determine the specificity and sensitivity of the assay. After selecting the primer pairs, their concentrations and GeXP-multiplex RT-PCR conditions were optimised. The established GeXP-multiplex RT-PCR assay can detect as few as 100 copies of premixed RNA templates. In the present study, 120 clinical specimens collected from domestic poultry at live bird markets and from wild birds were used to evaluate the performance of the assay. The GeXP-multiplex RT-PCR assay specificity was the same as that of conventional RT-PCR. Thus, the GeXP-multiplex RT-PCR assay is a rapid and relatively high-throughput method for detecting and identifying eight AIV subtypes that may infect humans.