Co-infection of H9N2 subtype avian influenza virus and QX genotype live attenuated infectious bronchitis virus increase the pathogenicity in SPF chickens.

Avian influenza virus (AIV) infection and vaccination against live attenuated infectious bronchitis virus (aIBV) are frequent in poultry worldwide. Here, we evaluated the clinical effect of H9N2 subtype AIV and QX genotype aIBV co-infection in specific-pathogen-free (SPF) white leghorn chickens and explored the potential mechanisms underlying the observed effects using by 4D-FastDIA-based proteomics. The results showed that co-infection of H9N2 AIV and QX aIBV increased mortality and suppressed the growth of SPF chickens. In particular, severe lesions in the kidneys and slight respiratory signs similar to the symptoms of virulent QX IBV infection were observed in some co-infected chickens, with no such clinical signs observed in single-infected chickens. The replication of H9N2 AIV was significantly enhanced in both the trachea and kidneys, whereas there was only a slight effect on the replication of the QX aIBV. Proteomics analysis showed that the IL-17 signaling pathway was one of the unique pathways enriched in co-infected chickens compared to single infected-chickens. A series of metabolism and immune response-related pathways linked with co-infection were also significantly enriched. Moreover, co-infection of the two pathogens resulted in the enrichment of the negative regulation of telomerase activity. Collectively, our study supports the synergistic effect of the two pathogens, and pointed out that aIBV vaccines might increased IBV-associated lesions due to pathogenic co-infections. Exacerbation of the pathogenicity and mortality in H9N2 AIV and QX aIBV co-infected chickens possibly occurred because of an increase in H9N2 AIV replication, the regulation of telomerase activity, and the disturbance of cell metabolism and the immune system.

Prevalence, Genotype Diversity, and Distinct Pathogenicity of 205 Gammacoronavirus Infectious Bronchitis Virus Isolates in China during 2019-2023.

Gammacoronavirus infectious bronchitis virus (IBV) causes a highly contagious disease in chickens and seriously endangers the poultry industry. The emergence and co-circulation of diverse IBV serotypes and genotypes with distinct pathogenicity worldwide pose a serious challenge to the development of effective intervention measures. In this study, we report the epidemic trends of IBV in China from 2019 to 2023 and a comparative analysis on the antigenic characteristics and pathogenicity of isolates among major prevalent lineages. Phylogenetic and recombination analyses based on the nucleotide sequences of the spike (S) 1 gene clustered a total of 205 isolates into twelve distinct lineages, with GI-19 as a predominant lineage (61.77 ± 4.56%) exhibiting an overall increasing trend over the past five years, and demonstrated that a majority of the variants were derived from gene recombination events. Further characterization of the growth and pathogenic properties of six representative isolates from different lineages classified four out of the six isolates as nephropathogenic types with mortality rates in one-day-old SPF chickens varying from 20-60%, one as a respiratory type with weak virulence, and one as a naturally occurring avirulent strain. Taken together, our findings illuminate the epidemic trends, prevalence, recombination, and pathogenicity of current IBV strains in China, providing key information for further strengthening the surveillance and pathogenicity studies of IBV.

Protection Against Infectious Bronchitis Virus Vaccine Recombinants and Chicken-Selected Vaccine Subpopulations.

Outbreaks of infectious bronchitis (IB) continue to occur from novel variants of IB virus (IBV) emerging from selection of vaccine subpopulations and/or naturally occurring recombination events. S1 sequencing of Arkansas (Ark) -type viruses obtained from clinical cases in Alabama broilers and backyard chickens shows both Ark Delmarva Poultry Industry (ArkDPI) vaccine subpopulations as well as Ark vaccine viruses showing recombination with other IB vaccine viruses. IB Ark-type isolates AL5, most similar to an ArkDPI vaccine subpopulation selected in chickens, AL4, showing a cluster of three nonsynonymous changes from ArkDPI subpopulations selected in chickens, and AL9, showing recombination with Massachusetts (Mass) -type IBV, were examined for pathogenicity and ability to break through immunity elicited by vaccination with a commercial ArkDPI vaccine. Analysis of predicted S1 protein structures indicated the changes were in regions previously shown to comprise neutralizing epitopes. Thus, they were expected to contribute to immune escape and possibly virulence. Based on clinical signs, viral load, and histopathology, all three isolates caused disease in naïve chickens, although AL9 and AL5 viral loads in trachea were statistically significantly higher (30- and 40-fold) than AL4. S1 gene sequencing confirmed the stability of the relevant changes in the inoculated viruses in the chickens, although virus in some individual chickens exhibited additional S1 changes. A single amino acid deletion in the S1 NTD was identified in some individual chickens. The location of this deletion in the predicted structure of S1 suggested the possibility that it was a compensatory change for the reduced ability of AL4 to replicate in the trachea of naïve chickens. Chickens vaccinated with a commercial ArkDPI vaccine at day of hatch and challenged at 21 days of age showed that vaccination provided incomplete protection against challenge with these viruses. Moreover, based on viral RNA copy numbers in trachea, differences were detected in the ability of the vaccine to protect against these IBV isolates, with the vaccine protecting the most poorly against AL4. These results provide additional evidence supporting that IBV attenuated vaccines, especially ArkDPI vaccines, contribute to perpetuating the problem of IB in commercial chickens.

Protección contra los virus de la bronquitis infecciosa vacunales recombinantes y las subpoblaciones de vacunas seleccionadas en pollos. Los brotes de la bronquitis infecciosa aviar continúan presentándose a partir de nuevas variantes de dicho virus, que surgen de la selección de subpoblaciones de vacunas y/o eventos de recombinación que ocurren naturalmente. La secuenciación del gene S1 de virus tipo Arkansas (Ark) obtenidos de casos clínicos en pollos de engorde y de traspatio de Alabama muestra que tanto las subpoblaciones de la cepa vacunal Arkansas Delmarva Poultry Industry (ArkDPI) así como los virus de la vacuna Arkansas muestran recombinación con otros virus vacunales de la bronquitis infecciosa. Los aislamientos del virus de la bronquitis infecciosa Arkansas tipo "AL5", más similares a una subpoblación de vacuna ArkDPI seleccionada en pollos, "AL4", que muestra un grupo de tres cambios no sinónimos de subpoblaciones de ArkDPI seleccionadas en pollos y el tipo "AL9", que muestra recombinación con el serotipo Massachusetts, se examinaron para determinar su patogenicidad y capacidad para traspasar la inmunidad generada por la vacunación con una vacuna comercial ArkDPI. El análisis de las estructuras predichas de la proteína S1 indicó que los cambios se produjeron en regiones que previamente se había demostrado comprendían epítopos neutralizantes. Por lo tanto, se esperaba que contribuyeran al escape inmunológico y posiblemente a la virulencia. Con base en los signos clínicos, la carga viral y la histopatología, los tres aislados causaron enfermedad en pollos sin exposición previa, aunque las cargas virales de AL9 y AL5 en la tráquea fueron estadísticamente significativamente mayores (30 y 40 veces) en comparación con AL4. La secuenciación del gene S1 confirmó la estabilidad de los cambios relevantes en los virus inoculados en los pollos, aunque el virus en algunos pollos individuales exhibió cambios adicionales en el gene S1. Se identificó una deleción de un solo aminoácido en el dominio terminal N del gene S1 (NTD S1) en algunos pollos individuales. La ubicación de esta eliminación en la estructura predicha del gene S1 sugirió la posibilidad de que se tratara de un cambio compensatorio por la capacidad reducida de AL4 para replicarse en la tráquea de pollos sin exposición previa. Los pollos vacunados con una vacuna comercial ArkDPI el día de la eclosión y desafiados a los 21 días de edad mostraron que la vacunación proporcionó una protección incompleta contra el desafío con estos virus. Además, basándose en el número de copias del ARN viral en la tráquea, se detectaron diferencias en la capacidad de la vacuna para proteger contra estos aislados del virus de la bronquitis infecciosa, siendo la vacuna con la protección más deficiente contra AL4. Estos resultados proporcionan evidencia adicional que respalda que las vacunas atenuadas contra el virus de la bronquitis infecciosa, especialmente las vacunas ArkDPI, contribuyen a perpetuar esta enfermedad en los pollos comerciales.

Isolation and phylogenetic analysis of avian infectious bronchitis virus from an imported chicken meat product in Malaysia.

Avian infectious bronchitis (IB), a Gammacoronavirus, is a highly contagious upper respiratory disease, affecting chickens of all ages with a significant economic threat to the poultry industry. In February 2020, a specimen of imported chicken meat product was received and requested for coronavirus testing. The result was positive for the avian coronavirus, the IB virus (IBV) by molecular detection in the pre-screening test. Thus, this study aimed to isolate and characterize the IBV from the specimen. Virus isolation via egg inoculation was attempted and IBV was successfully isolated. The S1 subunit of the spike (S) gene of the IBV was amplified, sequenced, and the Basic Local Alignment Search Tool (BLAST) analysis showed that the IBV has 99% and 98% nucleotide similarity with the Malaysian and China IBVs, respectively. The phylogenetic analysis indicated that the virus belongs to the GI-19 lineage (also known as the QX strain) and is grouped with other IBVs from Malaysia and China. The GI-19 lineage is one of the primary IB strains that circulate in Malaysia. The recovery of the virus may be due to the persistence characteristic of the virus on meat; and the cold chain practices in the imported food product prolong the survival of this coronavirus. Though IBV is not identified as a hazard in chicken meat or meat products, raw food should be cooked thoroughly before being consumed. With the increase in international trade in poultry and poultry products, disease screening at the entry point and import risk analysis is crucial to ensure food safety and prevent the introduction of new viruses into Malaysia.

Infectious bronchitis virus: an overview of the "chicken coronavirus".

Infectious bronchitis virus (IBV) is a highly contagious avian Gammacoronavirus that affects mainly chickens (Gallus gallus) but can circulate in other avian species. IBV constitutes a significant threat to the poultry industry, causing reduced egg yield, growth and mortality levels that can vary in impact. The virus can be transmitted horizontally by inhalation or direct/indirect contact with infected birds or contaminated fomites, vehicles, farm personnel and litter (Figure 1). The error-prone viral polymerase and recombination mechanisms mean diverse viral population results, with multiple genotypes, serotypes, pathotypes and protectotypes. This significantly complicates control and mitigation strategies based on vigilance in biosecurity and the deployment of vaccination.

Two mutations on S2 subunit were critical for Vero cell tropism expansion of infectious bronchitis virus HV80.

Infectious bronchitis virus (IBV) restricts cell tropism. Except for the Beaudette strain, other IBVs cannot infect mammalian cell lines. The limited cell tropism of other IBVs has hindered IBV vaccine development and research on the mechanisms of IBV infection. A novel Vero cell-adapted strain, HV80, has been previously reported. In this study, we constructed recombinants expressing the chimeric S glycoprotein, S1 or S2 subunit of strain H120 and demonstrated that mutations on S2 subunit are associated with the strain HV80 Vero cell adaptation. R687P or P687R substitution recombinants were constructed with the genome backbone of strains HV80 or H120. We found that the RRRR690/S motif at the S2' cleavage site is crucial to the Vero cell adaptation of strain HV80. Another six amino acid substitutions in the S2 subunit of the recombinants showed that the Q855H mutation induced syncytium formation. A transient transfection assay demonstrated the S glycoprotein with the PRRR690/S motif at the S2' cleavage site induced low-level cell-cell fusion, while H855Q substitution hindered cell-cell fusion and blocked cleavage event with S20 product. This study provides a basis for the construction of IBV recombinants capable of replicating in Vero cells, thus contributing to the advancement in the development of genetically engineered cell-based IBV vaccines.

Establishment of a rapid real-time fluorescence-based recombinase-aided amplification method for detection of avian infectious bronchitis virus.

Infectious bronchitis (IB) is an acute, highly contagious contact respiratory disease of chickens caused by infectious bronchitis virus (IBV). IBV is very prone to mutation, which brings great difficulties to the prevention and control of the disease. Therefore, there is a pressing need for a method that is fast, sensitive, specific, and convenient for detecting IBV. In this study, a real-time fluorescence-based recombinase-aided amplification (RF-RAA) method was established. Primers and probe were designed based on the conserved regions of the IBV M gene and the reaction concentrations were optimized, then the specificity, sensitivity, and reproducibility of this assay were tested. The results showed that the RF-RAA method could be completed at 39℃ within 20 min, during which the results could be interpreted visually in real-time. The RF-RAA method had good specificity, no cross-reaction with common poultry pathogens, and it detected a minimum concentration of template of 2 copies/µL for IBV. Besides, its reproducibility was stable. A total of 144 clinical samples were tested by RF-RAA and real-time quantitative PCR (qPCR), 132 samples of which were positive and 12 samples were negative, and the coincidence rate of the two methods was 100 %. In conclusion, the developed RF-RAA detection method is rapid, specific, sensitive, reproducible, and convenient, which can be utilized for laboratory detection and clinical diagnosis of IBV.

Characterisation of the tracheal transcriptional response of chickens to chronic infection with Mycoplasma synoviae.

Mycoplasma synoviae causes infectious synovitis and respiratory tract infections in chickens and is responsible for significant economic losses in the poultry industry. Effective attachment and colonisation of the trachea is critical for the persistence of the organism and progression of the disease it causes. The respiratory tract infection is usually sub-clinical, but concurrent infection with infectious bronchitis virus (IBV) is known to enhance the pathogenicity of M. synoviae. This study aimed to explore differentially expressed genes in the tracheal mucosa, and their functional categories, during chronic infection with M. synoviae, using a M. synoviae-IBV infection model. The transcriptional profiles of the trachea were assessed 2 weeks after infection using RNA sequencing. In chickens infected with M. synoviae or IBV, only 1 or 8 genes were differentially expressed compared to uninfected chickens, respectively. In contrast, the M. synoviae-IBV infected chickens had 621 upregulated and 206 downregulated genes compared to uninfected chickens. Upregulated genes and their functional categories were suggestive of uncontrolled lymphoid cell proliferation and an ongoing pro-inflammatory response. Genes associated with anti-inflammatory effects, pathogen removal, apoptosis, regulation of the immune response, airway homoeostasis, cell adhesion and tissue regeneration were downregulated. Overall, transcriptional changes in the trachea, 2 weeks after infection with M. synoviae and IBV, indicate immune dysregulation, robust inflammation and a lack of cytotoxic damage during chronic infection. This model provides insights into the pathogenesis of chronic infection with M. synoviae.

Deciphering infected cell types, hub gene networks and cell-cell communication in infectious bronchitis virus via single-cell RNA sequencing.

Infectious bronchitis virus (IBV) is a coronavirus that infects chickens, which exhibits a broad tropism for epithelial cells, infecting the tracheal mucosal epithelium, intestinal mucosal epithelium, and renal tubular epithelial cells. Utilizing single-cell RNA sequencing (scRNA-seq), we systematically examined cells in renal, bursal, and tracheal tissues following IBV infection and identified tissue-specific molecular markers expressed in distinct cell types. We evaluated the expression of viral RNA in diverse cellular populations and subsequently ascertained that distal tubules and collecting ducts within the kidney, bursal mucosal epithelial cells, and follicle-associated epithelial cells exhibit susceptibility to IBV infection through immunofluorescence. Furthermore, our findings revealed an upregulation in the transcription of proinflammatory cytokines IL18 and IL1B in renal macrophages as well as increased expression of apoptosis-related gene STAT in distal tubules and collecting duct cells upon IBV infection leading to renal damage. Cell-to-cell communication unveiled potential interactions between diverse cell types, as well as upregulated signaling pathways and key sender-receiver cell populations after IBV infection. Integrating single-cell data from all tissues, we applied weighted gene co-expression network analysis (WGCNA) to identify gene modules that are specifically expressed in different cell populations. Based on the WGCNA results, we identified seven immune-related gene modules and determined the differential expression pattern of module genes, as well as the hub genes within these modules. Our comprehensive data provides valuable insights into the pathogenesis of IBV as well as avian antiviral immunology.

The efficacy of simultaneous successive classic and variant infectious bronchitis virus vaccines versus circulating variant II Egyptian field virus.

Background: Being a ubiquitous, highly contagious virus with a continuous mutation and a large number of evolutions worldwide, the infectious bronchitis virus (IBV) continues to wreak problems among Egyptian chickens and generate economic losses. The commonly applied IBV vaccination protocols in broilers include alternatives to classic and/or variant attenuated live virus vaccines. Aim: The current study targeted to assess the protective efficacy of concurrent and successive Ma5 and 4/91 vaccine strain regimens against the field variant II IBV strain (IBV-EGY-ZU/Ck-127/2021) in chickens. Methods: Commercial broiler chickens were vaccinated with Ma5 and 4/91 strains simultaneously at 1 and 14 days of age. The evaluation parameters included clinical protection and humoral and early innate immunity aspects in the renal tissues of vaccinated and infected birds. Results: The vaccine regimen ameliorated the clinical and histopathological lesions against variant II IBV and enhanced body gain as well as succeeded in preventing tracheal shedding and minimizing cloacal shedding of the field virus. The IL-1ß mRNA gene expression was evident as early as 24 hours, with highly significant upregulation at 48 hours post vaccination and 24 hours post challenge (PC) in vaccinated birds. Remarkable upregulation was observed in oligoadenylate synthetases (OAS) expression 48 hours PC in vaccinated and unvaccinated infected birds. The vaccinated birds developed a significant antibody titer of 704.0 ± 111.98 at 28 days of age, with a consistent antibody titer increase after the challenge. Conclusion: Overall, a combination of heterologous protectotype commercial vaccines achieved good protection against the Egyptian variant II IBV strain. This vaccine program could be an effective protocol against the threat posed by IBV viruses circulating in the Egyptian field.

An Amplicon-Based Application for the Whole-Genome Sequencing of GI-19 Lineage Infectious Bronchitis Virus Directly from Clinical Samples.

Infectious bronchitis virus (IBV) causes a highly contagious respiratory disease in chickens, leading to significant economic losses in the poultry industry worldwide. IBV exhibits a high mutation rate, resulting in the continuous emergence of new variants and strains. A complete genome analysis of IBV is crucial for understanding its characteristics. However, it is challenging to obtain whole-genome sequences from IBV-infected clinical samples due to the low abundance of IBV relative to the host genome. Here, we present a novel approach employing next-generation sequencing (NGS) to directly sequence the complete genome of IBV. Through in silico analysis, six primer pairs were designed to match various genotypes, including the GI-19 lineage of IBV. The primer sets successfully amplified six overlapping fragments by long-range PCR and the size of the amplicons ranged from 3.7 to 6.4 kb, resulting in full coverage of the IBV genome. Furthermore, utilizing Illumina sequencing, we obtained the complete genome sequences of two strains belonging to the GI-19 lineage (QX genotype) from clinical samples, with 100% coverage rates, over 1000 × mean depth coverage, and a high percentage of mapped reads to the reference genomes (96.63% and 97.66%). The reported method significantly improves the whole-genome sequencing of IBVs from clinical samples; thus, it can improve understanding of the epidemiology and evolution of IBVs.

Differential Impact of Massachusetts, Canadian 4/91, and California (Cal) 1737 Genotypes of Infectious Bronchitis Virus Infection on Lymphoid Organs of Chickens.

Infectious bronchitis virus (IBV) induces severe economic losses in chicken farms due to the emergence of new variants leading to vaccine breaks. The studied IBV strains belong to Massachusetts (Mass), Canadian 4/91, and California (Cal) 1737 genotypes that are prevalent globally. This study was designed to compare the impact of these three IBV genotypes on primary and secondary lymphoid organs. For this purpose, one-week-old specific pathogen-free chickens were inoculated with Mass, Canadian 4/91, or Cal 1737 IBV variants, keeping a mock-infected control. We examined the IBV replication in primary and secondary lymphoid organs. The molecular, histopathological, and immunohistochemical examinations revealed significant differences in lesion scores and viral distribution in these immune organs. In addition, we observed B-cell depletion in the bursa of Fabricius and the spleen with a significant elevation of T cells in these organs. Further studies are required to determine the functional consequences of IBV replication in lymphoid organs.

The Effect of Global Spread, Epidemiology, and Control Strategies on the Evolution of the GI-19 Lineage of Infectious Bronchitis Virus.

The GI-19 lineage of infectious bronchitis virus (IBV) has emerged as one of the most impactful, particularly in the "Old World". Originating in China several decades ago, it has consistently spread and evolved, often forming independent clades in various areas and countries, each with distinct production systems and control strategies. This study leverages this scenario to explore how different environments may influence virus evolution. Through the analysis of the complete S1 sequence, four datasets were identified, comprising strains of monophyletic clades circulating in different continents or countries (e.g., Asia vs. Europe and China vs. Thailand), indicative of single introduction events and independent evolution. The population dynamics and evolutionary rate variation over time, as well as the presence and intensity of selective pressures, were estimated and compared across these datasets. Since the lineage origin (approximately in the mid-20th century), a more persistent and stable viral population was estimated in Asia and China, while in Europe and Thailand, a sharp increase following the introduction (i.e., 2005 and 2007, respectively) of GI-19 was observed, succeeded by a rapid decline. Although a greater number of sites on the S1 subunit were under diversifying selection in the Asian and Chinese datasets, more focused and stronger pressures were evident in both the European (positions 2, 52, 54, 222, and 379 and Thai (i.e., positions 10, 12, 32, 56, 62, 64, 65, 78, 95, 96, 119, 128, 140, 182, 292, 304, 320, and 323) strains, likely reflecting a more intense and uniform application of vaccines in these regions. This evidence, along with the analysis of control strategies implemented in different areas, suggests a strong link between effective, systematic vaccine implementation and infection control. However, while the overall evolutionary rate was estimated at approximately 10-3 to 10-4, a significant inverse correlation was found between viral population size and the rate of viral evolution over time. Therefore, despite the stronger selective pressure imposed by vaccination, effectively constraining the former through adequate control strategies can efficiently prevent viral evolution and the emergence of vaccine-escaping variants.

Differential analysis of IBV-infected primary chicken embryonic fibroblasts and immortalized DF-1.

Infectious bronchitis virus (IBV), the causative agent of infectious bronchitis, is responsible for major economic losses in the poultry industry worldwide. While IBVs can usually be passaged in primary chicken embryonic fibroblasts (CEFs), most of the wild ones cannot adapt to passaged cell lines. In this study, the wild strain CK/CH/MY/2020 was used to infect primary CEF and immortalize DF-1 CEF cells. Results indicated that IBV was able to cause lesions and pass onto CEF, but not DF-1. Indeed, the virus could enter DF-1 cells and synthesize the associated structural gene but could not assemble into complete viral particles for release. Furthermore, transcriptome sequencing analysis showed significant differences in gene expression between CEF and DF-1 cells after viral infection, although the corresponding antiviral responses could be activated in both cell types. The biggest difference was in terms of the amino acid biosynthesis pathway and the cytokine receptor interaction pathway, which were significantly and specifically activated in CEF. This could actually explain why intact viruses can be assembled but not in DF-1. In addition, SLBP and P2RX7 affect the replication of IBV's structural genes to some extent. Overall, IBV can enter CEF and DF-1 cells, but the complex intracellular cytokine interactions affect the assembly and release of viral particles. The insight will be useful for the study of IBV through in vitro transmission and pathogenesis. IMPORTANCE: Infectious bronchitis virus (IBV) is responsible for high morbidity and mortality as well as substantial economic losses worldwide. Transcriptome sequencing of IBV-infected chicken embryonic fibroblast and DF-1 cells revealed that the virus elicits antiviral immunity in cells after viral infection, but IBV cannot activate DF-1 cells to produce sufficient amounts of viral structures to assemble into complete virions, which may be caused by the interactions between cytokines. The study of IBV cellular adaptations is important for vaccine development and investigation of the pathogenesis of IBV.

Avian infectious bronchitis virus (AIBV) review by continent.

Infectious bronchitis virus (IBV) is a positive-sense, single-stranded, enveloped RNA virus responsible for substantial economic losses to the poultry industry worldwide by causing a highly contagious respiratory disease. The virus can spread quickly through contact, contaminated equipment, aerosols, and personal-to-person contact. We highlight the prevalence and geographic distribution of all nine genotypes, as well as the relevant symptoms and economic impact, by extensively analyzing the current literature. Moreover, phylogenetic analysis was performed using Molecular Evolutionary Genetics Analysis (MEGA-6), which provided insights into the global molecular diversity and evolution of IBV strains. This review highlights that IBV genotype I (GI) is prevalent worldwide because sporadic cases have been found on many continents. Conversely, GII was identified as a European strain that subsequently dispersed throughout Europe and South America. GIII and GV are predominant in Australia, with very few reports from Asia. GIV, GVIII, and GIX originate from North America. GIV was found to circulate in Asia, and GVII was identified in Europe and China. Geographically, the GVI-1 lineage is thought to be restricted to Asia. This review highlights that IBV still often arises in commercial chicken flocks despite immunization and biosecurity measures because of the ongoing introduction of novel IBV variants and inadequate cross-protection provided by the presently available vaccines. Consequently, IB consistently jeopardizes the ability of the poultry industry to grow and prosper. Identifying these domains will aid in discerning the pathogenicity and prevalence of IBV genotypes, potentially enhancing disease prevention and management tactics.

Detection of multiple recombinations of avian coronavirus in South Korea by whole-genome analysis.

Infectious bronchitis virus (IBV), an avian coronavirus, has caused considerable damage to the poultry industry. In Korea, indigenous KM91-like and newly introduced QX-like lineages belonging to the GI-19 lineage have been prevalent despite constant vaccination. In this study, complete genome sequences of 23 IBV isolates in Korea from 2010 to 2020 were obtained using next-generation sequencing, and their phylogenetic relationship and recombination events were analyzed. Phylogenetic analysis based on the S1 gene showed that all isolates belonged to the GI-19 lineage and were divided into five subgroups (KM91-like, K40/09-like, and QX-like II to IV). Among the 23 isolates, 14 recombinants were found, including frequent recombination between KM91-like and QX-like strains. In addition, it was observed that other lineages, such as GI-1, GI-13, and GI-16, were involved in recombination. Most recombination breakpoints were detected in the ORF1ab gene, particularly nsp3. However, when considering the size of each genome, recombination occurred more frequently in the 3a, E and 5a genes. Taken together, genetic recombination frequently occurred throughout the entire genome between various IBV strains in Korea, including live attenuated vaccine strain. Our study suggests the necessity of further research on the contribution of recombination of genomes outside the spike region to the biological characteristics of IBV.

Molecular characterization of a novel variant of infectious bronchitis virus from field outbreaks in backyard chicken population of North East India.

Infectious bronchitis virus (IBV) causes considerable economic impacts on global poultry production. Since its emergence in early 1930, IBV continues to evolve and now exists in a wide range of antigenically and genetically distinct variants, that makes the prevention and the control of the disease both complex and challenging. Although IBV has been reported regularly from different corner of India, information about the molecular epidemiology of circulating strain in relation to clinical form of the disease is not available. We have studied the clinico-pathology and confirmed eight distinct field outbreaks of the disease from poultry population of Mizoram, India. The clinical disease in affected birds resulted sever pathological lesions involving respiratory, gastrointestinal, and urinary system together. The complete S1 nucleotide sequences and protein analyses have revealed a distinct variant of genotype I-IBV (GI), designated as GI-24 circulating in India. The S1 protein of the field strains displayed unique additional eighteen amino acids at C terminal end when compared with M41strain. Comparison of the S1 protein among all the 27 lineages of GI revealed five mutations that are exclusive to only the Indian strains. All the field strains have also possessed the amino acid mutations at highly variable region 2 (HVR2) of S1 receptor-binding domain (RBD) that are considered characteristic of nephropathogenic strains. The circulating GI-24 strains displayed potency for a wide range of tropism from respiratory epithelium to GIT and urinary system. This study provides insight on recently emerging IBV outbreaks in NER, India, which might be causing huge economic losses to the poultry farmers in the region.

Antiviral effect of palmatine against infectious bronchitis virus through regulation of NF-κB/IRF7/JAK-STAT signalling pathway and apoptosis.

1. Infectious bronchitis virus (IBV), a gamma-coronavirus, can infect chickens of all ages and leads to an acute contact respiratory infection. This study evaluated the anti-viral activity of palmatine, a natural non-flavonoid alkaloid, against IBV in chicken embryo kidney (CEK) cells.2. The half toxic concentration (CC50) of palmatine was 672.92 µM, the half inhibitory concentration (IC50) of palmatine against IBV was 7.76 µM and the selection index (SI) was 86.74.3. Mode of action assay showed that palmatine was able to directly inactivate IBV and inhibited the adsorption, penetration and intracellular replication of IBV.4. Palmatine significantly upregulated TRAF6, TAB1 and IKK-ß compared with the IBV-infected group, leading to the increased expressions of pro-inflammatory cytokines IL-1ß and TNF-α in the downstream NF-κB signalling pathway.5. Palmatine significantly up-regulated the levels of MDA5, MAVS, IRF7, IFN-α and IFN-ß in the IRF7 pathway, inducing type I interferon production. It up-regulated the expression of 2'5'-oligoadenylate synthase (OAS) in the JAK-STAT pathway.6. IBV infection induced cell apoptosis and palmatine-treatment delayed the process of apoptosis by regulation of the expression of apoptosis-related genes (BAX, BCL-2, CASPASE-3 and CASPASE-8).7. Palmatine could exert anti-IBV activity through regulation of NF-κB/IRF7/JAK-STAT signalling pathways and apoptosis, providing a theoretical basis for the utilisation of palmatine to treat IBV infection.

Salmonella typhimurium Vaccine Candidate Delivering Infectious Bronchitis Virus S1 Protein to Induce Protection.

Infectious bronchitis (IB) is a highly infectious viral disease of chickens which causes significant economic losses in the poultry industry worldwide. An effective vaccine against IB is urgently needed to provide both biosafety and high-efficiency immune protection. In this study, the S1 protein of the infectious bronchitis virus was delivered by a recombinant attenuated Salmonella typhimurium vector to form the vaccine candidate χ11246(pYA4545-S1). S. typhimurium χ11246 carried a sifA- mutation with regulated delayed systems, striking a balance between host safety and immunogenicity. Here, we demonstrated that S1 protein is highly expressed in HD11 cells. Immunization with χ11246(pYA4545-S1) induced the production of antibody and cytokine, leading to an effective immune response against IB. Oral immunization with χ11246(pYA4545-S1) provided 72%, 56%, and 56% protection in the lacrimal gland, trachea, and cloaca against infectious bronchitis virus infection, respectively. Furthermore, it significantly reduced histopathological lesions in chickens. Together, this study provides a new idea for the prevention of IB.

The spike protein of the apathogenic Beaudette strain of avian coronavirus can elicit a protective immune response against a virulent M41 challenge.

The avian Gammacoronavirus infectious bronchitis virus (IBV) causes major economic losses in the poultry industry as the aetiological agent of infectious bronchitis, a highly contagious respiratory disease in chickens. IBV causes major economic losses to poultry industries across the globe and is a concern for global food security. IBV vaccines are currently produced by serial passage, typically 80 to 100 times in chicken embryonated eggs (CEE) to achieve attenuation by unknown molecular mechanisms. Vaccines produced in this manner present a risk of reversion as often few consensus level changes are acquired. The process of serial passage is cumbersome, time consuming, solely dependent on the supply of CEE and does not allow for rapid vaccine development in response to newly emerging IBV strains. Both alternative rational attenuation and cell culture-based propagation methods would therefore be highly beneficial. The majority of IBV strains are however unable to be propagated in cell culture proving a significant barrier to the development of cell-based vaccines. In this study we demonstrate the incorporation of a heterologous Spike (S) gene derived from the apathogenic Beaudette strain of IBV into a pathogenic M41 genomic backbone generated a recombinant IBV denoted M41K-Beau(S) that exhibits Beaudette's unique ability to replicate in Vero cells, a cell line licenced for vaccine production. The rIBV M41K-Beau(S) additionally exhibited an attenuated in vivo phenotype which was not the consequence of the presence of a large heterologous gene demonstrating that the Beaudette S not only offers a method for virus propagation in cell culture but also a mechanism for rational attenuation. Although historical research suggested that Beaudette, and by extension the Beaudette S protein was poorly immunogenic, vaccination of chickens with M41K-Beau(S) induced a complete cross protective immune response in terms of clinical disease and tracheal ciliary activity against challenge with a virulent IBV, M41-CK, belonging to the same serogroup as Beaudette. This implies that the amino acid sequence differences between the Beaudette and M41 S proteins have not distorted important protective epitopes. The Beaudette S protein therefore offers a significant avenue for vaccine development, with the advantage of a propagation platform less reliant on CEE.