Biology subjects

Brodrick, A. J.

Publications and source records attributed to Brodrick, A. J..

4 recordsLinked to original sources

bioRxiv2025

The C terminus of infectious bursal disease virus (IBDV) VP3 encodes a predicted intrinsically disordered region (IDR), which promotes the formation of cytoplasmic puncta and modulates their physical properties.

The virus factories (VFs) of infectious bursal disease virus (IBDV) form through liquid-liquid phase separation (LLPS). A major component of the IBDV VF is the nonstructural protein VP3. Here, we predicted the full-length structure of the VP3 monomer and homodimer and employed molecular dynamics simulations to characterize their behavior. We identified the 36 amino acid carboxy(C)-terminus as a highly dynamic intrinsically disordered region (IDR). We then compared the cytoplasmic puncta that were made in the presence of the wild type (wt) VP3 with those made with a VP3 that lacked the C-terminus (VP3{Delta}C). Using live-cell imaging with fluorescent reporter tagged proteins, we found that VP3{Delta}C puncta were significantly less numerous (p<0.0001), smaller (p<0.0001), and more irregular in shape than puncta formed in the presence of wt VP3, demonstrating that the VP3 C terminal IDR promoted their formation. Moreover, by fluorescence recovery after photobleaching (FRAP), the VP3{Delta}C puncta had a significantly reduced mobile fraction (0.29) as compared to full-length VP3 puncta (0.70) (p<0.001), demonstrating that the VP3 C terminal IDR modulated their physical properties. However, the VP3{Delta}C puncta still exhibited liquid-like fusion events in the cytoplasm and were sensitive to treatment with aliphatic diols. Moreover, VP3 did not form puncta when expressed alone, and the removal of the C terminus did not abolish puncta formation completely. We propose that VP3 forms part of a higher order complex with other biomolecules to drive LLPS, and that the VP3 C terminal IDR modulates the physical properties of the resultant LLPS structures. ImportanceLLPS is a phenomenon of growing interest in cell biology. It is a part of the replication cycles of diverse viruses, but our understanding of the molecular basis that underpins the mechanism of phase separation is incomplete. We previously demonstrated that the birnavirus IBDV, a major agricultural pathogen, exploits LLPS in the formation of its VFs. Here, we have characterized the C-terminal 36 amino acid region of IBDV VP3 bioinformatically and by molecular dynamics simulations and found that it encodes a highly dynamic intrinsically disordered region (IDR). Furthermore, we found this region to promote the formation of cytoplasmic puncta and modulate their physical properties. This work contributes to a more detailed understanding of birnavirus replication at the molecular level, and to the study of LLPS as a phenomenon.

molecular biology↗

bioRxiv2025

Prior infection with IBDV prolonged the shedding of a mallard H3N8 influenza A virus (IAV) challenge from the oropharyngeal cavity of some chickens and increased the number of amino acid substitutions in the IAV samples

Infectious bursal disease virus (IBDV) is endemic worldwide and causes immunosuppression in chickens. We hypothesized that a previous history of IBDV in chickens would render them more susceptible to infection by influenza A viruses (IAVs) from aquatic waterfowl reservoirs. To model this, we inoculated 14 day old specific pathogen free (SPF) chickens with a low pathogenicity avian influenza (LPAI) virus strain from a mallard (A/Mallard/Alberta/156/01 (H3N8)) and compared replication and shedding between immunocompetent chickens and chickens that had immune dysregulation due to a prior IBDV infection with strain F52/70 (genogroup A1B1) at 2 days of age. The mallard IAV strain replicated in the upper respiratory tract of the chickens, and virus was shed from the oropharyngeal cavity, but there was no shedding from the cloaca, and no transmission to sentinel chickens. Replication of the mallard IAV in the chicken host was associated with amino acid substitutions in the polymerase complex and HA. IBDV infection increased the average fold change of IAV replication in the trachea of chickens, prolonged the shedding of infectious IAV from 5 to 6 days in some chickens, increased the number of amino acid substitutions detected in the IAV population from 13 to 30, and significantly increased the number of mutations per IAV sample from 2.50 (SD +/- 1.83) in the Mock/IAV group to 4.75 (SD +/- 1.81) in the IBDV/IAV group (p < 0.01). Taken together, IBDV infection prolonged the shedding of the mallard IAV in some chickens and changed IAV intra-host evolution. Author summarySpillover of IAVs from wild aquatic waterfowl into poultry populations occur frequently, which increases the risk of human infection as people have more contact with poultry than wild birds. Poultry flocks may have other co-morbidities that may influence the spread of IAV. Our data demonstrate that prior IBDV infection increased the average fold change of a mallard H3N8 LPAI virus in the trachea of inoculated chickens, prolonged the shedding of infectious IAV from the oropharyngeal cavity, and significantly increased the average number of amino acid substitutions per IAV sample. We hypothesize that IBDV infection could increase the amount of IAV shed into the environment and broaden the diversity of the IAV population shed. We conclude that controlling the spread of wild aquatic waterfowl strains of IAV in chickens should involve a holistic approach, including the control of co-morbidities and immunosuppressive diseases that could exacerbate their spread.

microbiology↗

bioRxiv2024

Reassortant strains of infectious bursal disease virus (IBDV) belonging to genogroup A3B1 predominate in British broiler chicken flocks.

As part of ongoing epidemiological surveillance for infectious bursal disease virus (IBDV), the hypervariable region (HVR) of the VP2 capsid gene encoded by segment A, and a region of the VP1 polymerase gene, encoded by segment B, were sequenced from 20 IBDV-positive bursal samples obtained in 2020 and 2021, from 16 commercial British broiler farms. Of the 16 farms, none contained very virulent (vv) strains belonging to genogroup A3B2, but 5/16 (31%) contained strains of genogroup A3B1, demonstrating birds were infected with reassortant strains containing a vv segment A and a non-vv segment B. In addition, 3/16 (19%) farms contained vaccine or classical strains belonging to genogroup A1B1, and 8/16 (50%) were co-infected with both genogroup A1B1 and A3B1 strains. Therefore, a total of 13/16 (81%) of the farms contained genogroup A3B1 reassortant viruses, the majority of which 8/13 (62%)) were found to be co-infected with genogroup A1B1 strains. Moreover, of the flocks containing reassortant strains, 5/13 (38%) had HVR mutations Q219L, G254D, D279N, and N280T, consistent with a recently described Western European clade, but 8/13 had other mutations or no mutations, demonstrating that multiple clades were present in the samples. Taken together, vv strains were not detected in the British broiler flocks we sampled, whereas reassortant strains predominated, which belonged to different clades, and were frequently found in samples that were also infected with genogroup A1B1 strains.

microbiology↗

bioRxiv2022

Evaluating the breadth of neutralizing antibody responses elicited by infectious bursal disease virus (IBDV) genogroup A1 strains using a novel chicken B-cell rescue system and neutralization assay

Eight infectious bursal disease virus (IBDV) genogroups have been identified based on the sequence of the capsid hypervariable region (HVR) (A1-8), yet many vaccines are based on A1 strains. Given reported vaccine failures, there is a need to evaluate the ability of vaccines to neutralize the different genogroups. To address this, we used a reverse genetics system and the chicken B-cell line DT40 to rescue a panel of chimeric IBDVs and perform neutralization assays. Chimeric viruses had the backbone of a lab-adapted strain (PBG98) and the HVRs from diverse field strains: classical F52-70 (A1), US-variant Del-E (A2), Chinese-variant SHG19 (A2), very-virulent UK661 (A3), M04/09 distinct (A4), Italian ITA-04 (A6), and Australian-variant Vic-01/94 (A8). Rescued viruses showed no substitutions at amino-acid positions 253, 284, or 330, previously associated with cell-culture adaptation. Sera from chickens inoculated with wt (F52-70) or vaccine (228E) A1 strains had the highest mean virus neutralization (VN) titers against the A1 virus (log2 15.41 and 12.66), and the lowest against A2 viruses (log2 7.41-7.91, p=0.0001-0.0274), consistent with A1 viruses being most antigenically distant from A2 strains, which correlated with the extent of differences in the predicted HVR structure. VN titers against the other genogroups ranged from log2 9.32-13.32, and A1 strains were likely more closely antigenically related to genogroups A3 and A4 than A6 and A8. Our data are consistent with field observations, validating our method, which can used to screen future vaccine candidates for breadth of neutralizing antibodies, and evaluate the antigenic relatedness of different genogroups. ImportanceThere is a need to evaluate the ability of vaccines to neutralize diverse IBDV genogroups, and to better understand the relationship between HVR sequence, structure, and antigenicity. Here, we used a chicken B cell-line to rescue a panel of chimeric IBDVs with the HVR from seven diverse IBDV field strains, and conduct neutralization assays and protein modelling. We evaluated the ability of serum from vaccinated or infected birds to neutralize the different genogroups. Our novel chicken B-cell rescue system and neutralization assay can be used to screen IBDV vaccine candidates, platforms, and regimens for the breadth of neutralizing antibody responses elicited, evaluate the antigenic relatedness of diverse IBDV strains, and when coupled with structural modelling, elucidate immunodominant and conserved epitopes to strategically design novel IBDV vaccines in the future.

microbiology↗