Continuous clinicopathological and molecular recognition of very virulent infectious bursal disease virus in commercial broiler chickens.

Gumboro virus is one of the most dangerous immunosuppressant viruses that infect chickens and causes massive financial losses worldwide. The current study aims to conduct a molecular characterization of chicken farms for the infectious bursal disease virus (IBDV). Based on postmortem (PM) lesions, 125 bursal samples from 25 farms were collected from clinically diseased commercial chicken farms with increased mortality and suspected Gumboro virus infection. Pooled bursal samples from suspected IBD-vaccinated flocks were tested for IBDV by reverse transcriptase polymerase chain reaction (RT-PCR). Fifteen out of 25 pooled specimens were found positive for IBDV, with a 60% detection rate, and confirmed positive for very virulent IBDV (vvIBDV) by sequence analysis. Nucleotide phylogenetic analysis of VP1 and VP2 genes was employed to compare the 5 chosen isolates with strains representing different governorates in Egypt during 2022. All strains were clustered with vvIBDV with no evidence of reassortment in the VP1 gene. The VP1 and VP2 genes are divided into groups (I, II). The strains in our study were related to group II, and it acquired a new mutation in the VP2 gene that clustered it into new subgroup B. By mutation analysis, the VP2 gene of all strains had a characteristic mutation to vvIBDV. It acquired new mutations in HVRs compared with HK46 in Y220F, A222T/V in all strains in our study, and Q221K that was found in IBD-EGY-AH5 and AH2 in the loop PBC in addition to G254S in all strains in our study and Q249k that found in IBD-EGY-AH1 and AH3 in the loop PDE. These mutations are important in the virulency and antigenicity of the virus. The VP1 had 242E, 390M, and 393D which were characteristic of vvIBDV and KpnI restriction enzyme (777GGTAC/C782) in addition to a new mutation (F243Y and N383H) in IBD-EGY-AH1 and AH4 strains. According to the current study, the strains were distinct from the vaccinal strain; they could be responsible for the most recent IBDV outbreaks observed in flocks instead of received vaccinations. The current study highlighted the importance of molecular monitoring to keep up to date on the circulating IBDV for regular evaluation of commercial vaccination programs against circulating field viruses.

First Detection and Molecular Characterization of Novel Variant Infectious Bursal Disease Virus (Genotype A2dB1b) in Egypt.

Infectious bursal disease (IBD) is an immunosuppressive disease causing significant damage to the poultry industry worldwide. Its etiological agent is infectious bursal disease virus (IBDV), a highly resistant RNA virus whose genetic variability considerably affects disease manifestation, diagnosis and control, primarily pursued by vaccination. In Egypt, very virulent strains (genotype A3B2), responsible for typical IBD signs and lesions and high mortality, have historically prevailed. The present molecular survey, however, suggests that a major epidemiological shift might be occurring in the country. Out of twenty-four samples collected in twelve governorates in 2022-2023, seven tested positive for IBDV. Two of them were A3B2 strains related to other very virulent Egyptian isolates, whereas the remaining five were novel variant IBDVs (A2dB1b), reported for the first time outside of Eastern and Southern Asia. This emerging genotype spawned a large-scale epidemic in China during the 2010s, characterized by subclinical IBD with severe bursal atrophy and immunosuppression. Its spread to Egypt is even more alarming considering that, contrary to circulating IBDVs, the protection conferred by available commercial vaccines appears suboptimal. These findings are therefore crucial for guiding monitoring and control efforts and helping to track the spread of novel variant IBDVs, possibly limiting their impact.

Infectious bursal disease virus: predicting viral pathotype using machine learning models focused on early changes in total blood cell counts.

Infectious bursal disease (IBD) is an avian viral disease caused in chickens by infectious bursal disease virus (IBDV). IBDV strains (Avibirnavirus genus, Birnaviridae family) exhibit different pathotypes, for which no molecular marker is available yet. The different pathotypes, ranging from sub-clinical to inducing immunosuppression and high mortality, are currently determined through a 10-day-long animal experiment designed to compare mortality and clinical score of the uncharacterized strain with references strains. Limits of this protocol lie within standardization and the extensive use of animal experimentation. The aim of this study was to establish a predictive model of viral pathotype based on a minimum number of early parameters measured during infection, allowing faster pathotyping of IBDV strains with improved ethics. We thus measured, at 2 and 4 days post-infection (dpi), the blood concentrations of various immune and coagulation related cells, the uricemia and the infectious viral load in the bursa of Fabricius of chicken infected under standardized conditions with a panel of viruses encompassing the different pathotypes of IBDV. Machine learning algorithms allowed establishing a predictive model of the pathotype based on early changes of the blood cell formula, whose accuracy reached 84.1%. Its accuracy to predict the attenuated and strictly immunosuppressive pathotypes was above 90%. The key parameters for this model were the blood concentrations of B cells, T cells, monocytes, granulocytes, thrombocytes and erythrocytes of infected chickens at 4 dpi. This predictive model could be a second option to traditional IBDV pathotyping that is faster, and more ethical.

A rapid and affordable amplicon-based method for next-generation genome sequencing of the infectious bursal disease virus.

The infectious bursal disease virus (IBDV) causes a severe immunosuppressive disorder in young chickens. IBDV evolution resulted in the emergence of strains with divergent genetic, antigenic, and pathogenic characteristics. Genetic classification is typically performed by sequencing the coding region of the most immunogenic region of the viral protein 2 (VP2). Sequencing both double-stranded RNA genome segments is essential to achieve a more comprehensive IBDV classification that can detect recombinants and reassortments. Here, we report the development and standardization of a tiled PCR amplicon protocol for the direct and cost-effective genome sequencing of global IBDV strains using next-generation technology. Primers for tiled PCR were designed with adapters to bypass expensive and time-consuming library preparation steps. Sequencing was performed on Illumina MiniSeq equipment, and fourteen complete genomes of field strains were assembled using reference sequences. The PCR-enrichment step was used to obtain genomes from low-titer biological samples that were difficult to amplify using traditional sequencing. Phylogenetic analyses of the obtained genomes confirmed previous strain classification. By combining the enrichment methodology with massive sequencing, it is possible to obtain IBDV genomic sequences in a fast and affordable manner. This procedure can be a valuable tool to better understand virus epidemiology.

Immunization and Host Responses to MB-1, a Live Hatchery Vaccine against Infectious Bursal Disease.

MB-1 is an attenuated infectious bursal disease virus vaccine. Previously, we observed a temporal delay of vaccine virus replication in the bursae of chicks due to maternally derived antibodies (MDAs). The mechanism that allowed its survival despite MDA neutralization remained unclear. We hypothesized that after vaccination at 1 day of age (DOA), the MB-1 virus penetrates and resides in local macrophages that are then distributed to lymphoid organs. Furthermore, MB-1's ability to survive within macrophages ensures its survival during effective MDA protection. PCR analysis of lymphoid organs from chicks with MDA, vaccinated on 1 DOA, demonstrated that the MB-1 virus was identified at low levels solely in the spleen pre-14 days of age. Fourteen days after vaccination, the virus was identified using PCR in the bursa, with viral levels increasing with time. The possible delay in viral colonization of the bursa was attributed to the presence of anti-IBDV capsid VP2 maternal IgA and IgY in the bursa interstitium. These indicate that during the period of high MDA levels, a small but viable MB-1 viral reservoir was maintained in the spleen, which might have served to colonize the bursa after MDA levels declined. Thereafter, individual immunization of chicks against Gumboro disease was achieved.

Evaluación y detección bioinformática de terpenoides con potencial inhibitorio de la proteína viral 1 (VP1) del virus de la bursitis infecciosa / Evaluation and bioinformatic detection of terpenoids with inhibitory potential of viral protein 1 (VP1) of infectious bursitis virus

El virus de la bursitis infecciosa (IBDV) es el agente causal de la enfermedad de la bursa, la cual afecta principalmente a poblaciones avícolas jóvenes y genera un impacto económico negativo en la producción. La proteína viral (VP1) es una enzima con funciones clave para la replicación del genoma viral, por lo que puede ser considerada blanco para la búsqueda de compuestos con posibles actividades inhibitorias. El objetivo de esta investigación fue evaluar terpenoides con potencial inhibitorio de la proteína VP1 del IBDV mediante herramientas de aproximaciones bioinformáticas. Se seleccionó un total de 52 terpenoides, cuyas propiedades farmacológicas, farmacocinéticas y tóxicas (ADME-Tox) se evaluaron. Las moléculas sin actividades tóxicas y con aptitudes farmacocinéticas fueron sometidas a pruebas exhaustivas de acoplamiento molecular con el sitio catalítico de la VP1 mediante el uso del algoritmo genético y de Broyden-Fletcher-Goldfarb-Shanno junto con el método de optimización local de gradientes. Los datos obtenidos revelaron que la Giberelina A1 presenta valores de energía libre de unión significativamente (p< 0,05) favorables (ΔG=-7,28±0,06 kcal/mol; Kdcalc= 8,62±0,99 µM) en comparación con los sustratos rCTP y rGTP. El complejo Giberelina A1-VP1 presenta puentes de hidrógeno con los residuos Arg335 y Asp402, los cuales cumplen roles importantes en la actividad catalítica en la replicación viral. Estos hallazgos sugieren que el terpenoide Giberelina A1 puede ser considerado como compuesto candidato para estudios in vitro de inhibición de funciones de la VP1 e in vivode actividades antivirales contra el virus de la bursitis infecciosa.

Infectious bursitis virus (IBDV) is the infectious agent of bursal disease, which mainly affects young poultry populations, generating a negative economic impact on productions. The viral protein 1 (VP1) is an enzyme with important functions for the replication of the viral genome, so it could be considered as a target for searching compounds with possible inhibitory activities. The aim of this research was to evaluate terpenoids with inhibitory potential of the VP1 protein of IBDV using computational approximations tools. A total of 52 terpenoids were selected and evaluated for their pharmacological, pharmacokinetic and toxic properties (ADME-Tox). Molecules without toxic activities and with pharmacokinetic competences were subjected to extensive molecular docking tests with the catalytic site of VP1 using the genetic and Broyden-Fletcher-Goldfarb-Shanno algorithms with a local gradient optimization method. Data obtained revealed that Gibberellin A1 exhibits significantly (P < 0.05) favorable binding free energy values (ΔG=-7.28±0.06 kcal/mol; Kdcalc= 8.62±0.99 µM) compared to rCTP and rGTP subs-trates. The Gibberellin A1-VP1 complex exhibits hydrogen bonds with residues Arg335 and Asp402, which play important roles in catalytic activity in viral replication. These findings suggest that the terpenoid Gibberellin A1 could be considered as a candidate compound for in vitro studies of inhibition of VP1 functions and in vivo antiviral activities against infectious bursitis virus.

Folic acid enhances proinflammatory and antiviral molecular pathways in chicken B-lymphocytes infected with a mild infectious bursal disease virus.

1. This study evaluated the effect of folic acid (FA) supplementation on the proinflammatory and antiviral molecular pathways of B-lymphocytes infected with a modified live IBDV (ST-12) mild vaccine strain during a timed post-infection analysis.2. A chicken B-lymphocytes (DT-40) cell line was cultured in triplicate at a concentration of 5 × 105 cells per well in 24-well plates; and was divided into three groups: 1: No virus, FA; 2: Virus, no FA; 3: Virus + FA at a concentration of 3.96 mM. The experiment was repeated three times.3. Cells in groups 2 and 3 were infected with a modified live IBDV (ST-12) mild vaccine strain at one multiplicity of infection (MOI: 1). After 1 hour of virus adsorption, samples were collected at 0, 3, 6, 12, 24 and 36 hours post-infection (hpi).4. The modified live IBDV (ST-12) mild vaccine strain triggered a B-lymphocyte specific immune response associated with the upregulation of genes involved in virus recognition (Igß), virus sensing (TLR-2, TLR-3, TLR-4 and MDA5), signal transduction and regulation (TRIF, MyD88 and IRF7), and the antiviral effector molecules (IFN-α, OAS, PKR, and viperin).5. FA supplementation modulated IBDV replication and regulated the proinflammatory and antiviral downstream molecular pathways.6. In conclusion, the low virulent pathotype serotype I modified live IBDV (ST-12) mild vaccine strain was able to trigger and mount an immune response in chicken B-lymphocytes without affecting B-cell viability. FA supplementation modulated B lymphocytes response and improved their innate immune proinflammatory and antiviral response molecular pathways.

Sequence diversity and evolution of infectious bursal disease virus in Iraq.

Background: Infectious Bursal Disease (IBD) is a highly infectious disease which causes huge economic losses to the poultry industry due to the direct impact of the illness and indirect consequences such as decreasing the general immunity of the flock, leaving it naive to other diseases. In Iraq, IBD is highly prevalent despite vaccination programs, yet studies on sequence diversity of the causative virus are still rare.  Methods: A sample from Bursa of Fabricius from an IBD outbreak in a flock in the city of Najaf in Iraq was smeared on an FTA card. Amplicons of targeted regions in VP1 and VP2 genes were generated and sequenced. Sequences were then compared with other local and global sequences downloaded from GenBank repositories. Sequence alignment and DNA sequence analyses were achieved using MUSCLE, UGENE and MEGAx software. The molecular clock and sequence evolutionary analyses were applied using MEGAx tools.  Results: The strain sequenced in this study belongs to a very virulent Infectious Bursal Disease Virus (vvIBDV) as the DNA and phylogenetic analysis of VP1 and VP2 gene sequences showed a mutual clustering with similar sequences belonging to vvIBDV genogroup 3. Analyses of the hyper variable region of VP2 gene (hvVP2) of IBDV isolates from Iraq indicates a presence of sequence diversity. Interestingly, the two vaccine strains Ventri IBDV Plus and ABIC MB71 that showed the highest sequence similarity to the local isolates in the hvVP2 region are not used in vaccination routine against IBDV in Iraq.  Conclusion: Sequences of vvIBDV in Iraq are diverse. Remarkably, some of the available vaccine strains show high sequence similarity with local strains in Iraq; however, they are not included in the routine vaccination programs. Analysis of more samples involving more geographical regions is needed to draw a detailed map of antigenic diversity of IBDV in Iraq.

Assessing the antigenicity of different VP3 regions of infectious bursal disease virus in chickens from South Brazil.

BACKGROUND: Infectious bursal disease (IBD), also known as Gumboro disease, is a viral infection that causes mortality and immunosuppression in chickens (Gallus gallus). VP2 and VP3 are the major structural viral capsid components and are the most immunogenic proteins of IBD virus (IBDV). Reliable diagnostic tests using VP2 and VP3 produced in heterologous systems are important tools to control this infection. One advantage of an IBD diagnostic based on VP3, over those that use VP2, is that VP3 has linear epitopes, enabling its production in bacteria. RESULTS: We tested the suitability of recombinant VP3 (rVP3) as a diagnostic reagent in an enzyme-linked immunosorbent assay (ELISA). Compared with a commercial test, rVP3 ELISA showed high sensitivity and specificity as a diagnostic tool for vaccinated animals. In addition, rVP3, but not the commercial ELISA, was able to detect antibodies in nonvaccinated chickens, probably developed against circulating IBDV strains. It was possible the assessment of VP3 regions antigenicity using chicken antisera. CONCLUSIONS: The full-length recombinant VP3 can be used to assess post vaccination immunological status of chickens and its production is feasible and inexpensive. The evaluation of VP3 regions as candidates for general use in the diagnosis of IBD in chickens should be conducted with caution. Our work was the first to identify several regions of VP3 recognized by chicken antibodies.

Safety and Efficacy Profile of Live, Intermediate Plus Vaccine Against Infectious Bursal Disease Based on Strain G6.

Infectious bursal disease (IBD) is an acute, highly contagious, immunosuppressive disease of young chickens that causes considerable economic loss in the poultry industry worldwide. Vaccination with live attenuated vaccines is still the most important method used for the control and prevention of IBD in chickens. Here we present the results of in vitro characterization, as well as efficacy and safety testing of a live, intermediate plus vaccine against IBD based on strain G6. Strain characterization confirmed that G6 strain is an intermediate plus strain, showing a high degree of homology with the existing vaccine strains of the same virulence. Safety studies showed that chickens can be vaccinated from 10 days of age. Onset and duration of immunity in specific pathogen free and maternally derived antibodies (MDA) chickens was proven to be 14 and 35 days after vaccination, respectively. When immunizing MDA-positive chickens, vaccine is capable of breakthrough at a titer of ≤500 ELISA units. The field trial conducted on commercial broilers showed a 95% protection against vvIBDV challenge. Stability of the freeze-dried vaccine after reconstitution was confirmed over a period of 3 h. Overall, IBD G6 vaccine has shown good safety and efficacy profile in accordance with European Pharmacopoeia requirements.

Comparative in vivo pathogenicity study of an ITA genotype isolate (G6) of infectious bursal disease virus.

Recently, a new genotype of infectious bursal disease virus (IBDV), named ITA, was detected in IBD-vaccinated Italian broilers. Genome characterization revealed ITA to be a genetically different IBDV, belonging to genogroup 6 according to a recently proposed IBDV classification. The currently available clinical data do not allow any definition of the degree of pathogenicity of the ITA-IBDV isolates. In the present study, a pathogenicity trial was conducted by the oral inoculation of specific-pathogen-free (SPF) chickens. Birds were housed in poultry isolators and inoculated at 35 days of age with an ITA-IBDV isolate (35 birds) or a strain belonging to the G1a genogroup as a comparison (35 birds). Control birds (25 birds) were contextually mock-inoculated with sterile water. Birds were observed daily for clinical signs and at 0, 7, 14, 21 and 28 days post-inoculation (dpi) were bled for IBDV antibody detection. At 2, 4, 7, 14, 21 and 28 dpi, five birds from each of the inoculated groups, and three from the control group, were euthanized and subjected to a post-mortem examination; the bursa:body weight and thymus:body weight ratios were calculated. Microscopic lesions of the bursa and thymus were scored on the basis of lymphoid necrosis and/or depletion or cortex atrophy, respectively. Both viruses induced a subclinical course of disease, as neither clinical signs nor mortality were recorded during the study, even in the presence of typical IBDV gross and microscopic lesions. Bursal damage, measured by the bursa:body weight ratio, was more noticeable and precocious after ITA-IBDV inoculation. Histopathology scores of the bursa, indicative of rapid lymphoid depletion, confirmed the aggressiveness of the ITA-IBDV strain in this organ. This study showed that, although the ITA-IBDV strain tested causes infection with a subclinical course, it induces severe damage to lymphoid tissues. Therefore, its circulation in birds might be a threat for the poultry industry and may jeopardize the success of the production cycle.

Molecular characterisation of infectious bursal disease virus in Namibia, 2017.

Between July and September 2017, samples collected from six unvaccinated chickens in Namibia were shown to be positive for infectious bursal disease virus (IBDV) by RT-PCR. Partial sequence and phylogenetic analysis of the VP1 and VP2 genes from six viruses revealed that they all belong to the very virulent pathotype (Genogroup 3) and are genetically very similar to IBDVs identified in neighbouring Zambia. This is the first molecular characterisation of IBDV in Namibia and has implications on the control and management of the disease in the country.

Phylogeny and evolution of infectious bursal disease virus circulating in Turkish broiler flocks.

The emergence of new infectious bursal disease virus (IBDV) variants can threaten poultry health and production all over the world causing significant economic losses. Therefore, this study was performed to determine IBDV molecular epidemilogy, VP2 gene variation, and corresponding pathological lesions in IBDV infected chickens in Turkey. For this, 1855 bursa of Fabricius samples were collected from 371 vaccinated broiler flocks. Atrophia and haemorrhages were seen in the bursa Fabricius of very virulent IBDV (vvIBDV) infected chickens. Partial VP2 gene was sequenced and phylogenetic, recombination, and evolutionary analyses were performed. 1548 (83.5%) out of 1855 of bursa of Fabricius samples were IBDV positive and 1525 of those could be sequenced. The recombination analysis did not detect occurrence of any recombination event among the Turkish strains. Among 1525 sequenced samples, 1380 of them were found to be classical strains. Among 1380 classical strains, 1317 were similar to IBDV 2512, 11 to Faragher 52/70, 40 to 228 E, and 12 to Lukert strain. Out of 1525 reverse transcriptase ploymerase chain reaction positive samples, 144 of them were found to be similar to vvIBDV-VP2 gene reported to GenBank previously. The phylogenetic tree performed on a broad sequence dataset demonstrated grouping of vvIBDV Turkish strains in three different clusters, including sequences collected also from Iraq and Kuwait (Cluster 1), Indian (Cluster 2), and a distinct Turkish-only cluster (Cluster 3). The evolutionary rate estimation on branches/clades including Turkish strain mirrored the expected one for RNA viruses and no significant differences were found among different considered branches. In conclusion, results of this study indicate that vvIBDV strains similar to those circulating in various countries in the Middle East are present and undergoing evolution in chickens from Turkish broiler flocks. This point needs to be taken into account in planning adequate control strategies.

A proposed nomenclature for infectious bursal disease virus isolates.

Infectious bursal disease virus (IBDV) was initially identified in the USA. For decades, these viruses were not categorized using a typing system because they were considered to be antigenically and pathogenically similar. In the 1980s, a second major serotype, serotype 2, was found in turkeys. Classification of IBDV became more complex with the discovery of antigenic variant strains called "variants" in the United States and a highly virulent strain known as "very virulent" or vvIBDV identified in Europe. To distinguish the IBDV strains identified prior to this time from the antigenic variant viruses, the term "classic viruses" was adopted. Studies over the next three decades produced a wealth of information on the antigenicity, pathogenicity and molecular structure of IBDV isolates. These data made it clear that the descriptive nomenclature used for IBDV was inadequate. For example, not all viruses identified as vvIBDV by genotyping are highly pathogenic; some have reassorted genome segments that result in lower virulence. Furthermore, variant viruses are not an antigenically homogeneous group and the term "classic virus" has been used interchangeably to describe antigenic and pathogenic types of IBDV. These and other issues make the current naming system for strains of IBDV archaic. The lack of uniform testing and standards for antigenicity and pathogenicity makes it difficult to categorize IBDV strains on a global basis. A new nomenclature that includes a genotyping system that can easily be applied worldwide is proposed and serves as a platform to begin discussions on its value to the scientific community.

Circulation of very virulent avian infectious bursal disease virus in Finland.

In the spring of 2014 infectious bursal disease (IBD) was confirmed in a Finnish layer flock exhibiting clinical signs and increased mortality. Organ and blood samples were sent for diagnosis to the Finnish Food Safety Authority Evira. IBD virus (IBDV) was detected in RT-PCR studies. Altogether hens from six layer farms associated with increased mortality (7-10%, worst case 30%) were diagnosed with IBD during 2014. Antibodies were also detected with IBD-ELISA tests in hens on two farms. Phylogenetic analysis showed that the causative agent of the 2014 IBD outbreak was a non-reassortant very virulent type IBDV. The representative virus strains from previous IBD outbreaks in 1978, 1987 and 1993 were also included in the analysis. The strains isolated in 2014 and 1993 were very similar indicating circulation of a very virulent IBDV for over 20 years in the country. In spite of the comprehensive phylogenetic analysis, the definitive origin of the viruses from 2014 and previous outbreaks remains unclear.

Infectious Bursal Disease Virus Hijacks Endosomal Membranes as the Scaffolding Structure for Viral Replication.

Birnaviruses are unconventional members of the group of double-stranded RNA (dsRNA) viruses that are characterized by the lack of a transcriptionally active inner core. Instead, the birnaviral particles organize their genome in ribonucleoprotein complexes (RNPs) composed by dsRNA segments, the dsRNA-binding VP3 protein, and the virally encoded RNA-dependent RNA polymerase (RdRp). This and other structural features suggest that birnaviruses may follow a completely different replication program from that followed by members of the Reoviridae family, supporting the hypothesis that birnaviruses are the evolutionary link between single-stranded positive RNA (+ssRNA) and dsRNA viruses. Here we demonstrate that infectious bursal disease virus (IBDV), a prototypical member of the Birnaviridae family, hijacks endosomal membranes of infected cells through the interaction of a viral protein, VP3, with the phospholipids on the cytosolic leaflet of these compartments for replication. Employing a mutagenesis approach, we demonstrated that VP3 domain PATCH 2 (P2) mediates the association of VP3 with the endosomal membranes. To determine the role of VP3 P2 in the context of the virus replication cycle, we used avian cells stably overexpressing VP3 P2 for IBDV infection. Importantly, the intra- and extracellular virus yields, as well as the intracellular levels of VP2 viral capsid protein, were significantly diminished in cells stably overexpressing VP3 P2. Together, our results indicate that the association of VP3 with endosomes has a relevant role in the IBDV replication cycle. This report provides direct experimental evidence for membranous compartments such as endosomes being required by a dsRNA virus for its replication. The results also support the previously proposed role of birnaviruses as an evolutionary link between +ssRNA and dsRNA viruses.IMPORTANCE Infectious bursal disease (IBD; also called Gumboro disease) is an acute, highly contagious immunosuppressive disease that affects young chickens and spreads worldwide. The etiological agent of IBD is infectious bursal disease virus (IBDV). This virus destroys the central immune organ (bursa of Fabricius), resulting in immunosuppression and reduced responses of chickens to vaccines, which increase their susceptibility to other pathogens. IBDV is a member of Birnaviridae family, which comprises unconventional members of dsRNA viruses, whose replication strategy has been scarcely studied. In this report we show that IBDV hijacks the endosomes of the infected cells for establishing viral replication complexes via the association of the ribonucleoprotein complex component VP3 with the phospholipids in the cytosolic leaflet of endosomal membranes. We show that this interaction is mediated by the VP3 PATCH 2 domain and demonstrate its relevant role in the context of viral infection.

Propagation and titration of infectious bursal disease virus, including non-cell-culture-adapted strains, using ex vivo-stimulated chicken bursal cells.

Infectious bursal disease virus (IBDV) is a Birnaviridae family member of economic importance for poultry. This virus infects and destroys developing B lymphocytes in the cloacal bursa, resulting in a potentially fatal or immunosuppressive disease in chickens. Naturally occurring viruses and many vaccine strains are not able to grow in in vitro systems without prior adaptation, which often affects viral properties such as virulence. Primary bursal cells, which are the main target cells of lymphotropic IBDV in vivo, may represent an attractive system for the study of IBDV. Unfortunately, bursal cells isolated from bursal follicles undergo apoptosis within hours following their isolation. Here, we demonstrate that ex vivo stimulation of bursal cells with phorbol 12-myristate 13-acetate maintains their viability long enough to allow IBDV replication to high titres. A wide range of field-derived or vaccine serotype 1 IBDV strains could be titrated in these phorbol 12-myristate 13-acetate -stimulated bursal cells and furthermore were permissive for replication of non-cell-culture-adapted viruses. These cells also supported multistep replication experiments and flow cytometry analysis of infection. Ex vivo-stimulated bursal cells therefore offer a promising tool in the study of IBDV.

Induction of apoptosis in the bursa of Fabricius by vaccination against Gumboro disease.

Infectious bursal disease is a severe acute viral disease of young chickens, affecting mainly the B-lymphocytes in the bursa of Fabricius, leading to severe immunosuppression as a result of the death of lymphoid cells. In the bursa infected with infectious bursal disease virus, viral replication is associated with apoptosis of lymphoid cells, inflammatory change and atrophy. Vaccination has appeared to be a crucial factor for control, with live attenuated vaccines being the most used. However, the apoptotic effect of these vaccines on the bursa has not been tested. We determined the apoptotic effect caused by the most used vaccines in local production on the bursa of Fabricius cells and the correlation with histological changes. In this study, it was demonstrated that apoptosis levels in the vaccinated groups were higher than those observed in the non-vaccinated birds leading to the conclusion that the action of the live virus vaccine strains modifies the boundary of the bursa and shapes processes of cell death by apoptosis. In contrast to other studies, the vaccine strains used did not show the phenomenon of bursal atrophy during the experimental period.

The Characteristics of an Ovine Lymphoid Cell-Line sensitive to Vaccinal Infectious Bursal Disease Virus Strain.

Infectious bursal disease (IBD), also known as Gumboro disease, is a globally well-known disease with a significant socio-economic effect. For control of IBD, several commercial egg- and cell-based vaccines are prepared. The cell-based IBD vaccines are significantly cost-effective; however, it is essential to confirm their safety and efficacy. The main cell line used to product the cell-based IBD vaccines, is a primary chicken embryo fibroblast (CEF). Nevertheless, manipulation of CEF is extremely challenging and time-consuming. This study aimed to characterize a sensitive suspension cell culture from ovine lymphoid, according to WHO technical report series; No. 978, Annex III. This authentication covered the growth curves, sensitivity, stability, karyotyping and identifying the adventitious agents. This cell line passed all defined tests and was considered as a suitable one for IBD vaccine preparation in a large scale.

Exploratory serosurvey for antibodies to avian pathogens in backyard chickens from a satellite community of Jalapa City, Guatemala

An exploratory serosurvey was conducted to determine the presence of circulating antibodies to avian patho-gens in backyard chickens from Los Achiotes (LAC), a satellite community of Jalapa City, located in eastern Guatemala. Blood samples from 51 adult chickens belonging to 51 households were taken and investigated for the presence of antibodies to Avian Influenza (AI), Newcastle Disease (ND), Infectious Bronchitis (IB), Infectious Bursal Disease (IBD), Mycoplasma gallisepticum (MG) and M. synoviae (MS). Antibodies for AI, ND, were investigated by Hemagglutination Inhibition, for IB and IBD by ELISA (BioChek®) and for MG and MS by a rapid serum plate agglutination test. The cut-off point for positive titers was 1:4 for AI and ND and a 0.2 S/P ratio for IB and IBD. All sampled chickens were positive for concomitant antibodies to various pathogens. Over half of the chickens were positive reactors to antibodies to all six tested pathogens; about a third carried antibodies to five and the rest to four or three. The frequencies of positive reactors were: AI = 27 (53%); ND = 49 (96.1%); IB = 50 (98%); IBD = 51 (100%); MG = 45 (88%) and MS = 48 (94%). The results show that the dynamic population of backyard chickens in LAC could be a potential threat to backyard poultry, farm poultry, wild birds and human population. The need to develop interventions and policies following the One Health approach (animal health to achieve human health) is stressed.

Se realizó un estudio serológico exploratorio buscando anticuerpos contra patógenos aviares en gallinas de traspatio de la comunidad Los Achiotes –una comunidad satélite de la Ciudad de Jalapa, en el oriente de Guatemala−. Se tomaron muestras de sangre de 51 gallinas provenientes de sendas casas. Se buscaron anticuerpos contra influenza aviar (IA), enfermedad de Newcastle (ENC), bronquitis infecciosa (BI), enfermedad de Gumboro (EG), Mycoplasma gallisepticum (MG) y M. synoviae (MS). Para investigar la presencia de anticuerpos contra IA y ENC se utilizó la prueba de inhibición de hemoaglutinación; para los anticuerpos contra BI la prueba de ELISA BioChek® y para los anticuerpos contra MG y MS la prueba rápida en placa. El punto de corte para títulos positivos fue de 1:4 para IA y ENC y de una razón S/P de 0.2 para BI y EG. Todas las gallinas muestreadas portaban concomitantemente anticuerpos contra varios patógenos aviares. Más de la mitad de las gallinas portaban anticuerpos contra los seis patógenos estudiados. Las frecuencias de reactores positivos a anticuerpos fueron: IA = 27 (53%); ENC = 49 (96.1%); BI = 50 (98%); EG = 51 (100%); MG = 45 (88%) y MS = 48 (94%). Se concluye que la población dinámica de gallinas de traspatio de Los Achiotes podría ser una potencial amenaza para la avicultura artesanal, la avicultura tecnificada, las aves silvestres y la población humana. Se señala la necesidad de generar intervenciones y políticas desde la corriente denominada Una salud (salud animal para lograr la salud humana).