Publications

The Pirbright Institute publication directory contains details of selected publications written by our researchers.

There were a total of 2049 results for your search.

Abstract

Recent evidence indicates that local immune responses and tissue resident memory T cells (TRM) are critical for protection against respiratory infections but there is little information on the contributions of upper and lower respiratory tract (URT and LRT) immunity. To provide a rational basis for designing methods for optimal delivery of vaccines to the respiratory tract in a large animal model, we investigated the distribution of droplets generated by a mucosal atomization device (MAD) and two vibrating mesh nebulizers (VMNs) and the immune responses induced by delivery of influenza virus by MAD in pigs. We showed that droplets containing the drug albuterol, a radiolabel (99mTc-DTPA) or a model influenza virus vaccine (S-FLU) have similar aerosol characteristics. 99mTc-DTPA scintigraphy showed that VMNs deliver droplets with uniform distribution throughout the lungs as well as the URT. Surprisingly MAD administration (1ml/nostril) also delivered a high proportion of the dose to the lungs, albeit concentrated in a small area. After MAD administration of influenza virus, antigen specific T cells were found at high frequency in nasal turbinates, trachea, broncho-alveolar lavage, lungs, tracheobronchial nodes and blood. We conclude that the pig is useful for investigating optimal targeting of vaccines to the respiratory tract.Competing Interest StatementMH operates a preclinical CRO and received a fee for his involvement in the pig scintigraphy study. RM and MJ are employed by Aerogen Limited, focused on development of vibrating mesh nebulizer technologies.

Graham S P, McLean R K, Spencer A J, Belij-Rammerstorfer S, Wright D, Ulaszewska M, Edwards J C, Hayes J W P, Martini V, Thakur N, Conceicao C, Dietrich I, Shelton H, Waters R, Ludi A, Wilsden G, Browning C, Bialy D, Bhat S, Stevenson-Leggett P, Hollinghurst P, Gilbride C, Pulido D, Moffat K, Sharpe H, Allen E, Mioulet V, Chiu C, Newman J, Asfor A S, Burman A, Crossley S, Huo J, Owens R J, Carroll M, Hammond J A, Tchilian E, Bailey D, Charleston B, Gilbert S C, Tuthill T J, Lambe T (2020)

Evaluation of the immunogenicity of prime-boost vaccination with the replication-deficient viral vectored COVID-19 vaccine candidate ChAdOx1 nCoV-19

bioRxiv, 159715

Abstract

Clinical development of the COVID-19 vaccine candidate ChAdOx1 nCoV-19, a replication-deficient simian adenoviral vector expressing the full-length SARS-CoV-2 spike (S) protein was initiated in April 2020 following non-human primate studies using a single immunisation. Here, we compared the immunogenicity of one or two doses of ChAdOx1 nCoV-19 in both mice and pigs. Whilst a single dose induced antigen-specific antibody and T cells responses, a booster immunisation enhanced antibody responses, particularly in pigs, with a significant increase in SARS-CoV-2 neutralising titres.

Abstract

Due to the current pandemic, global shortage of reagents has drawn interest in developing alternatives to increase the number coronavirus tests. One such alternative is sample pooling. Here we compared commercial kits that are used in COVID-19 diagnostics, in terms of sensitivity and feasibility for use in pooling. We showed that pooling of up to 60 samples did not affect the efficiency of the kits. Also, the RNA dependent RNA polymerase (RdRp) is a more suitable target in pooled samples than the Envelope (E) protein. This approach could provide an easy method of screening large number of samples and help adjust different government regulations.

Competing Interest Statement: The authors have declared no competing interest.

Abstract

African swine fever (ASF) is a devastating disease in pigs, with no vaccines for control. The genetic manipulation of African swine fever virus (ASFV) is often tedious and time consuming. Here, we describe a method to manipulate the virus genome to produce gene deletion viruses in a much-reduced time. This method combines the conventional homologous recombination with fluorescent-activated cells sorting (FACS), to isolate and purify viruses expressing fluorescent reporter genes. With three rounds of single cell isolation via FACS and two rounds of limiting dilution, we deleted two additional genes, EP153R and EP402R, from Benin 97/1 ASFV lacking the DP148R gene. By combining different fluorescent markers, this method has the potential to greatly facilitate studies on understanding ASFV gene functions and develop candidate live-attenuated vaccines.

Abstract

Using the best animal models to study immune responses against specific pathogens or vaccines can dramatically accelerate our understanding. Veterinary species are well studied, particularly livestock, to reduce their disease burden. They have also proven to be powerful models, especially for zoonotic pathogens and novel vaccination strategies. A prerequisite for any model selection is having the right quality and range of species?specific immunological reagents. To help promote the widest possible use of veterinary species, an open access website (https://www.immunologicaltoolbox.co.uk) has been created as a central community annotated hub for veterinary immunological reagents. The website is also the portal into services offered by the UK Immunological Toolbox project that includes antibody generation, sequencing and recombinant expression. The funding for this effort is linked into sustainable sources but ultimate success still relies on community engagement to continually increase the quality and quantity of information. It is hoped that as more users and reagent owners engage it will become an essential resource for researchers, veterinarians and clinicians alike by removing barriers that prevent the use of the most informative animal models.

Flannery J, Frost L, Fay P, Hicks H, Henstock M, Smreczak M, Or?owska A, Rajko-Nenow P, Darpel K, Batten C (2020)

BTV-14 infection in sheep elicits viraemia with mild clinical symptoms

Microorganisms 8 (6), 892

Abstract

In 2011, Bluetongue virus serotype 14 (BTV-14) was detected in Russia during routine surveillance, and was subsequently found in a number of European countries. The strain had high sequence similarity to a BTV-14 vaccine strain. We aimed to determine the risk of this BTV-14 strain causing disease in a UK sheep breed. Four Poll Dorset sheep were infected with a Polish isolate of BTV-14 and infection kinetics were monitored over 28 days. BTV RNA was detected in EDTA blood by 4 days post-infection (dpi) and remained detectable at 28 days post-infection (dpi). Peak viraemia occurred at 6 and 7 dpi with Ct values ranging between 24.6 and 27.3 in all infected animals. BTV antibodies were detected by 10 dpi using a commercial ELISA and neutralising antibodies were detected from 10 dpi. BTV was isolated between 6 and 12 dpi. All infected sheep developed mild clinical signs such as reddening of conjunctiva and mucosal membranes, with one sheep demonstrating more overt clinical signs. Two uninoculated control animals remained clinically healthy and did not have detectable BTV RNA or antibodies. The overall mild clinical symptoms caused by this BTV-14 in this highly susceptible sheep breed were in accordance with the asymptomatic infections observed in the affected countries.

Abstract

IBDV is economically important to the poultry industry. Very virulent (vv) strains cause higher mortality rates than other strains for reasons that remain poorly understood. In order to provide more information on IBDV disease outcome, groups of chickens (n = 18) were inoculated with the vv strain, UK661, or the classical strain, F52/70. Birds infected with UK661 had a lower survival rate (50%) compared to F52/70 (80%). There was no difference in peak viral replication in the bursa of Fabricius (BF), but the expression of chicken IFNα, IFNβ, MX1, and IL-8 was significantly lower in the BF of birds infected with UK661 compared to F52/70 (p < 0.05) as quantified by RTqPCR, and this trend was also observed in DT40 cells infected with UK661 or F52/70 (p < 0.05). The induction of expression of type I IFN in DF-1 cells stimulated with polyI:C (measured by an IFN-β luciferase reporter assay) was significantly reduced in cells expressing ectopic VP4 from UK661 (p < 0.05), but was higher in cells expressing ectopic VP4 from F52/70. Cells infected with a chimeric recombinant IBDV carrying the UK661-VP4 gene in the background of PBG98, an attenuated vaccine strain that induces high levels of innate responses (PBG98-VP4UK661) also showed a reduced level of IFNα and IL-8 compared to cells infected with a chimeric virus carrying the F52/70-VP4 gene (PBG98-VP4F52/70) (p < 0.01), and birds infected with PBG98-VP4UK661 also had a reduced expression of IFNα in the BF compared to birds infected with PBG98-VP4F52/70 (p < 0.05). Taken together, these data demonstrate that UK661 induced the expression of lower levels of anti-viral type I IFN and proinflammatory genes than the classical strain in vitro and in vivo and this was, in part, due to strain-dependent differences in the VP4 protein.

Abstract

Bluetongue virus (BTV) causes an economically important disease, bluetongue (BT), in susceptible ruminants and is transmitted primarily by species of Culicoides biting midges (Diptera: Ceratopogonidae). Since 2006, northern Europe has experienced multiple incursions of BTV through a variety of routes of entry, including major outbreaks of strains of BTV serotype 8 (BTV-8) and BTV serotype 1 (BTV-1), which overlapped in distribution within southern Europe. In this paper, we examined the variation in response to coinfection with strains of BTV-1 and BTV-8 using an in vivo transmission model involving Culicoides sonorensis, low passage virus strains, and sheep sourced in the United Kingdom. In the study, four sheep were simultaneously infected using BTV-8 and BTV-1 intrathoracically inoculated C. sonorensis and co-infections of all sheep with both strains were established. However, there were significant variations in both the initiation and peak levels of virus RNA detected throughout the experiment, as well as in the infection rates in the C. sonorensis that were blood-fed on experimentally infected sheep at peak viremia. This is discussed in relation to the potential for reassortment between these strains in the field and the policy implications for detection of BTV strains.

Abstract

Background: The recent reclassification of formaldehyde as a presumed carcinogen prompted the investigation into the comparative efficacy of hydrogen peroxide as a fumigant in microbiological safety cabinets.

Introduction: The aim of the study was to quantify the biocidal efficacy of formaldehyde fumigation, including variables such as exposure time and concentration, and then to compare this to the biocidal efficacy achieved from a hydrogen peroxide vapor fumigation system. The study also investigated the ability of both fumigants to permeate the microbiological safety cabinet (MBSC), including the workspace, under the work tray, and after the HEPA filters. Furthermore, the effect of organic soiling on efficacy was also assessed. Infectious bronchitis virus (IBV) was used as the biological target to develop this study model.

Methods: A model using IBV was developed to determine the efficacy of formaldehyde and hydrogen peroxide as fumigants. Virus was dried on stainless steel discs, and variables including concentration, time, protein soiling, and location within an MBSC were assessed.

Results: It was demonstrated that formaldehyde fumigation could achieve a 6-log reduction in the titer of the virus throughout the cabinet, and high protein soiling in the presentation did not affect efficacy. Appropriate cycle parameters for the hydrogen peroxide system were developed, and when challenged with IBV, it was shown that vaporized hydrogen peroxide could achieve an equal 6-log titer reduction as formaldehyde within the cabinet workspace and overcome the presence of soiling.

Conclusion: Hydrogen peroxide was demonstrated to be a viable alternative to formaldehyde under most situations tested. However, the hydrogen peroxide system did not achieve an equal titer reduction above the cabinet’s first HEPA filter using the cabinet workspace cycle, and further optimization of the hydrogen peroxide cycle parameters, including pulsing of the cabinet fans, may be required to achieve this.

Abstract

Zoonotic coronavirus (CoV) infections, such as those responsible for the current severe acute respiratory syndrome-CoV 2 (SARS-CoV-2) pandemic, cause grave international public health concern. In infected cells, the CoV RNA-synthesizing machinery associates with modified endoplasmic reticulum membranes that are transformed into the viral replication organelle (RO). Although double-membrane vesicles (DMVs) appear to be a pan-CoV RO element, studies to date describe an assortment of additional CoV-induced membrane structures. Despite much speculation, it remains unclear which RO element(s) accommodate viral RNA synthesis. Here we provide detailed 2D and 3D analyses of CoV ROs and show that diverse CoVs essentially induce the same membrane modifications, including the small open double-membrane spherules (DMSs) previously thought to be restricted to gamma- and delta-CoV infections and proposed as sites of replication. Metabolic labeling of newly synthesized viral RNA followed by quantitative electron microscopy (EM) autoradiography revealed abundant viral RNA synthesis associated with DMVs in cells infected with the beta-CoVs Middle East respiratory syndrome-CoV (MERS-CoV) and SARS-CoV and the gamma-CoV infectious bronchitis virus. RNA synthesis could not be linked to DMSs or any other cellular or virus-induced structure. Our results provide a unifying model of the CoV RO and clearly establish DMVs as the central hub for viral RNA synthesis and a potential drug target in CoV infection.

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