Development of a quadrivalent foot-and-mouth disease vaccine candidate for use in East Africa.

Foot-and-mouth disease (FMD) is a highly contagious viral disease of livestock which is prevalent across Africa, the Middle East, Asia, and South America where it has a severe economic impact on the agriculture industry. Vaccination with inactivated viral vaccines is used as the main control measure in these endemic regions of the world, however the presence of multiple serotypes, subtypes, and the continual emergence of new, antigenically divergent strains limits its effectiveness. East Africa (EA) has been identified as a region that would particularly benefit from updated FMD vaccines, since those currently in use contain older strains which do not provide good protection against contemporary strains. Four serotypes are currently circulating in EA, necessitating the development of a quadrivalent vaccine containing representative strains of each serotype. A key consideration in the selection of vaccine strains is the stability of the virus particle, since the capsids readily dissociate on exposure to elevated temperatures, but only intact capsids induce protective immunity to FMD. Therefore, with a view to producing a more stable, updated quadrivalent vaccine for EA, we recently screened a panel of foot-and-mouth disease virus (FMDV) isolates from the region to select strains with naturally higher thermostabilities and confirmed their immunogenicity in cattle. Herein we describe the formulation and serological assessment of wild-type and recombinant quadrivalent vaccine candidates comprising these stable strains, and demonstrate that both vaccines generate high neutralising antibody titres against the homologous strains and also to heterologous strains from EA. Importantly, the vaccine passed the criteria set by the AgResults vaccine challenge project and offers good cross-protection against a panel of regional FMDV strains.

Effect of mucosal adjuvant IL-1ß on heterotypic immunity in a pig influenza model.

T cell responses directed against highly conserved viral proteins contribute to the clearance of the influenza virus and confer broadly cross-reactive and protective immune responses against a range of influenza viruses in mice and ferrets. We examined the protective efficacy of mucosal delivery of adenoviral vectors expressing hemagglutinin (HA) and nucleoprotein (NP) from the H1N1 virus against heterologous H3N2 challenge in pigs. We also evaluated the effect of mucosal co-delivery of IL-1ß, which significantly increased antibody and T cell responses in inbred Babraham pigs. Another group of outbred pigs was first exposed to pH1N1 as an alternative means of inducing heterosubtypic immunity and were subsequently challenged with H3N2. Although both prior infection and adenoviral vector immunization induced strong T-cell responses against the conserved NP protein, none of the treatment groups demonstrated increased protection against the heterologous H3N2 challenge. Ad-HA/NP+Ad-IL-1ß immunization increased lung pathology, although viral load was unchanged. These data indicate that heterotypic immunity may be difficult to achieve in pigs and the immunological mechanisms may differ from those in small animal models. Caution should be applied in extrapolating from a single model to humans.

OMIP-089: Cattle T-cell phenotyping by an 8-color panel.

This multiplex staining panel was developed to differentiate cattle T cells into conventional (CD4 and CD8) and unconventional (γδ-TCR) subsets as well as their stage of differentiation and activation. The combination of CD45RO and CD62L allows the identification of naïve (TNaïve ), central memory (TCM ), effector memory (TEM ) and terminal effector (TTE ) T cells. Activated cattle T cells (TAV ) can be identified by the cell surface expression of CD25. This panel was developed using cryopreserved cattle peripheral blood mononuclear cells (PBMCs) and tested on fresh as well as stimulated PBMCs. Therefore, this 8-color, 10-parameter flow cytometry panel simultaneously identifies cattle TNaïve , TAV , TCM , TEM , TTE and γδ-TCR cells. This panel will improve our ability to examine T-cell response to pathogens and vaccines in cattle including the potential to identify previously undescribed subpopulations. Furthermore, this panel can be readily optimized for other bovid species as many of these reagents are likely to cross react.

Carbon fibre instrumentation for scoliosis surgery in children with spinal cord intramedullary tumours: a novel technical note.

INTRODUCTION: Scoliosis in children is the most common spinal deformity seen by general practitioners, paediatricians and spinal surgeons. Progressive scoliosis can result in the development of a worsening deformity and cosmesis. Patients usually present with aesthetic concerns. Progressive scoliosis that fails conservative management may require or be offered surgical intervention. Intramedullary tumours may be associated with scoliosis. Management of patients with these dual pathologies can be challenging. Classical scoliosis instrumentation utilising titanium implants impairs post-operative MRI evaluation with metal artefacts. Carbon fibre instrumentations has the potential to reduce the imaging metal artefacts but has not been described in scoliosis correction. METHODS: Surgical technical note describing correction of scoliosis in two adolescents' with intradural tumours utilising carbon fibre implants. RESULTS: We developed a hybrid approach where we initially used titanium implants to manipulate the deformity then replaced the construct with carbon fibre implants in the same setting to maintain the deformity correction with good follow up outlook. CONCLUSION: Our technique is robust, safe and replicable. It enabled appropriate post-operative MRI evaluation of the neural structures with a reduced risk of metal artefacts.

OMIP-085: Cattle B-cell phenotyping by an 8-color panel.

This 8-color panel has been optimized to distinguish between functionally distinct subsets of cattle B cells in both fresh and cryopreserved peripheral blood mononuclear cells (PBMCs). Existing characterized antibodies against cell surface molecules (immunoglobulin light chain (S-Ig[L]), CD20, CD21, CD40, CD71, and CD138) enabled the discrimination of 24 unique populations within the B-cell population. This allows the identification of five putative functionally distinct B-cell subsets critical to infection and vaccination responses: (1) naïve B cells (BNaïve ), (2) regulatory B cells (BReg ), (3) memory B cells (BMem ), (4) plasmablasts (PB), and (5) plasma cells (PC). Although CD3 and CD8α can be included as an additional dump channel, it does not significantly improve the panel's ability to separate "classical" B cells. This panel will promote better characterization and tracking of B-cell responses in cattle as well as other bovid species as the reagents are likely to cross react.

Development and Evaluation of Molecular Pen-Side Assays without Prior RNA Extraction for Peste des Petits Ruminants (PPR) and Foot and Mouth Disease (FMD).

Animal diseases such as peste des petits ruminants (PPR) and foot and mouth disease (FMD) cause significant economic losses in endemic countries and fast, accurate in-field diagnostics would assist with surveillance and outbreak control. The detection of these pathogens is usually performed at reference laboratories, tested using assays that are recommended by The World Organisation for Animal Health (OIE), leading to delays in pathogen detection. This study seeks to demonstrate a proof-of-concept approach for a molecular diagnostic assay that is compatible with material direct from nasal swab sampling, without the need for a prior nucleic acid extraction step, that could potentially be applied at pen-side for both PPR and FMD. The use of such a rapid, low-cost assay without the need for a cold chain could permit testing capacity to be established in remote, resource limited areas and support the surveillance activities necessary to meet the goal of eradication of PPR by 2030. Two individual assays were developed that detect > 99% of PPR and FMD sequences available in GenBank, demonstrating pan-serotype FMD and pan-lineage PPR assays. The ability for the BioGene XF reagent that was used in this study to lyse FMD and PPR viruses and amplify their nucleic acids in the presence of unprocessed nasal swab eluate was evaluated. The reagent was shown to be capable of detecting the viral RNA present in nasal swabs collected from naïve and infected target animals. A study was performed comparing the relative specificity and sensitivity of the new assays to the reference assays. The study used nasal swabs collected from animals before and after infection (12 cattle infected with FMDV and 5 goats infected with PPRV) and both PPR and FMD viral RNA were successfully detected two to four days post-infection in all animals using either the XF or reference assay reagents. These data suggest that the assays are at least as sensitive as the reference assays and support the need for further studies in a field setting.

Porcine Respiratory Coronavirus as a Model for Acute Respiratory Coronavirus Disease.

In the light of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic, we have developed a porcine respiratory coronavirus (PRCV) model for in depth mechanistic evaluation of the pathogenesis, virology and immune responses of this important family of viruses. Pigs are a large animal with similar physiology and immunology to humans and are a natural host for PRCV. Four PRCV strains were investigated and shown to induce different degrees of lung pathology. Importantly, although all four strains replicated equally well in porcine cell lines in vitro and in the upper respiratory tract in vivo, PRCV strains causing more severe lung pathology were also able to replicate in ex vivo tracheal organ cultures as well as in vivo in the trachea and lung. The time course of infection of PRCV 135, which caused the most severe pulmonary pathology, was investigated. Virus was shed from the upper respiratory tract until day 10 post infection, with infection of the respiratory mucosa, as well as olfactory and sustentacular cells, providing an excellent model to study upper respiratory tract disease in addition to the commonly known lower respiratory tract disease from PRCV. Infected animals made antibody and T cell responses that cross reacted with the four PRCV strains and Transmissible Gastroenteritis Virus. The antibody response was reproduced in vitro in organ cultures. Comparison of mechanisms of infection and immune control in pigs infected with PRCVs of differing pathogenicity with human data from SARS-CoV-2 infection and from our in vitro organ cultures, will enable key events in coronavirus infection and disease pathogenesis to be identified.

Assessment of the Welfare of Experimental Cattle and Pigs Using the Animal Welfare Assessment Grid.

The Animal Welfare Assessment Grid (AWAG) is a method for assessing quality of life, originally designed for experimental primates. This study adapts the AWAG for use in cattle and pigs, by adapting the factors included for these species and including data which had been collected previously as the standard approach to monitoring these species in research. The intention is that the results presented here will allow the future data collected for experimental cattle and pigs to be optimised for inclusion in an AWAG. Data were collected from two vaccine assessment studies at the Pirbright Institute. Factors were scored for every recorded event using retrospective data and CCTV clips. There was a lack of behavioural data recorded in both studies, which limited the accuracy of assessing each animal's welfare. This paper emphasises the importance of including behavioural information when assessing welfare and not simply relying on assessment of physical condition. Scores peaked following an exponential rise as animals reached set humane end points. This demonstrated the potential of using the AWAG to aid the decision-making of when euthanasia should be performed. Our study shows the AWAG to be a useful tool for assessing welfare, which can be used in harm:benefit assessment.

Foot-and-mouth disease virus infection in the domestic dog (Canis lupus familiaris), Iran.

BACKGROUND: Foot-and-mouth disease (FMD) is a highly infectious viral disease, recognised to affect animals in the order Artiodactyla. The disease is rarely fatal in adult animals, however high mortality is associated with neonatal and juvenile infection. CASE PRESENTATION: Five puppies died after being fed lamb carcases, the lambs having died during an outbreak of FMD in Iran. Following a post-mortem examination, cardiac tissue from one of the dead puppies was subjected to virus isolation, antigen ELISA, real-time RT-PCR, sequencing and confocal microscopy to assess the presence and characteristics of any FMD virus. The virological and microscopic examination of the cardiac tissue provided evidence of FMD virus replication in the canine heart. CONCLUSIONS: The data generated in this study demonstrate for the first time that FMD virus can internalise and replicate in dogs and may represent an epidemiologically significant event in FMD transmission, highlighting the dangers of feeding diseased animal carcases to other species. The reporting of this finding may also focus attention on similar disease presentations in dogs in FMD endemic countries allowing a better understanding of the prevalence of such events.

Pathogenesis of non-epithelial foot-and-mouth disease in neonatal animals.

Foot-and-mouth disease virus (FMDV), which causes a highly contagious viral disease of cloven-hoofed animals, is notable for epithelial cell tropism, resulting in the appearance of vesicles on the feet and in and around the mouth in infected animals, while FMDV infection in neonatal animals is also associated with not only epithelial lesions, but also muscle-associated lesions, which leads to myocarditis, resulting in high-mortality. However, critical knowledge about the non-epithelial tropism of FMDV is still lacking. In this paper, the current progress of the FMDV non-epithelial tropisms is summarized and the possible role of the key viral and cellular components involved is discussed.

Paediatric spinal trauma presenting to a UK major trauma centre.

PURPOSE: Paediatric spine trauma is uncommon and is managed differently from adults due to the anatomical differences of the paediatric spine. The paediatric spine is less ossified, with lax ligaments and a higher fulcrum in the c-spine which results in a different pattern of injuries. The aim of this study is to provide a contemporary audit of paediatric spinal trauma. METHODS: A retrospective review was conducted using the Trauma and Audit Research Network database at a major trauma centre (2011-2018). All patients < 18 years old with a spine injury underwent case note and radiology review. RESULTS: A total of 72 patients (37, 51.4% male with an average age of 13.3 (± 5.9) years old) were identified. The most common mechanism of injury was road traffic collisions (n = 39, 54.2%). The most common sporting cause was motocross accidents (n = 6, 8.3%), and a further 6 (8.3%) patients had a suspected inflicted injury. Eight patients (11.1%) sustained a spinal cord injury. Twenty-seven (37.5%) patients underwent surgical intervention to treat their spinal injury. CONCLUSION: This series demonstrates the profile of injury mechanisms causing paediatric spinal injuries. Paediatric spine injuries continue to have the potential for lifelong disability and require careful, specialist management. This series also highlights certain causes such as motocross accidents and suspected inflicted injury which are more frequent than expected and raise potential public health concerns.

Ruptured intra-cranial arachnoid cysts: a case series from a single UK institution.

INTRODUCTION: Intracranial arachnoid cysts are a common incidental finding; however presentation with cyst rupture is a recognised complication. Patients are advised to avoid contact sports to reduce the risk of cyst rupture but the risk of rupture is not currently known. The aim of this study is to describe a single institution's experience of managing ruptured intra-cranial arachnoid cysts. METHOD: A retrospective case note review of all patients admitted to a single institution with a ruptured intra-cranial arachnoid cyst between 2005 and 2016 (inclusive). Medical records were reviewed for demographics, history of trauma, surgical treatment and radiological evidence of cyst rupture. RESULTS: Fourteen patients were identified for inclusion with an average age of 23.4 years (range 7-57) and 10 (71%) were male. Nine patients (64%) had a documented history of head trauma. Eleven patients (78.6%) required neurosurgical intervention which included hygroma/haematoma evacuation only (n = 5), haematoma evacuation with cyst fenestration (n = 4) and cyst fenestration/shunting only (n = 2). Twelve patients (85%) experienced full recovery and the remaining two had persisting headaches or neurological symptoms. DISCUSSION: Cyst rupture remains an uncommon cause for presentation with arachnoid cysts. However the majority are associated with head trauma and thus current advice to avoid contact sports seems justified.

A COVID-19 vaccine candidate using SpyCatcher multimerization of the SARS-CoV-2 spike protein receptor-binding domain induces potent neutralising antibody responses.

There is need for effective and affordable vaccines against SARS-CoV-2 to tackle the ongoing pandemic. In this study, we describe a protein nanoparticle vaccine against SARS-CoV-2. The vaccine is based on the display of coronavirus spike glycoprotein receptor-binding domain (RBD) on a synthetic virus-like particle (VLP) platform, SpyCatcher003-mi3, using SpyTag/SpyCatcher technology. Low doses of RBD-SpyVLP in a prime-boost regimen induce a strong neutralising antibody response in mice and pigs that is superior to convalescent human sera. We evaluate antibody quality using ACE2 blocking and neutralisation of cell infection by pseudovirus or wild-type SARS-CoV-2. Using competition assays with a monoclonal antibody panel, we show that RBD-SpyVLP induces a polyclonal antibody response that recognises key epitopes on the RBD, reducing the likelihood of selecting neutralisation-escape mutants. Moreover, RBD-SpyVLP is thermostable and can be lyophilised without losing immunogenicity, to facilitate global distribution and reduce cold-chain dependence. The data suggests that RBD-SpyVLP provides strong potential to address clinical and logistic challenges of the COVID-19 pandemic.

Federation of European Laboratory Animal Science Associations recommendations of best practices for the health management of ruminants and pigs used for scientific and educational purposes.

Most ruminants and pigs used for scientific and educational aims are bred not for these purposes but in a farm environment. Given the wide range of diseases that these species might have, ensuring that the animals' health status is appropriate can be complex and challenging. The Federation of European Laboratory Animal Science Associations has previously published recommendations for the health monitoring of experimental colonies of pigs (1998) and, respectively, calves, sheep and goats (2000). Unfortunately, the uptake of those recommendations was poor and insufficiently reported in scientific publications. These new recommendations for best practice focus on the main species of ruminants (cattle, sheep and goats) and pigs. They provide general and specific information helpful for designing a health management programme for the suppliers and for the user establishments, as well as guidance on animal procurement. Critical thinking based on the fields of use of the animals is promoted, aiming to help in taking informed decisions rather than establishing an exhaustive exclusion list for pathogens. Implementing the best health and welfare management practices should be done under the guidance of a competent attending veterinarian, with expertise and sufficient authority to take the appropriate action, doubled by excellent communication skills. It is strongly recommended that the user establishment's veterinarian works in close collaboration with the supplier's veterinarian.

A Systematic Evaluation of High-Throughput Sequencing Approaches to Identify Low-Frequency Single Nucleotide Variants in Viral Populations.

High-throughput sequencing such as those provided by Illumina are an efficient way to understand sequence variation within viral populations. However, challenges exist in distinguishing process-introduced error from biological variance, which significantly impacts our ability to identify sub-consensus single-nucleotide variants (SNVs). Here we have taken a systematic approach to evaluate laboratory and bioinformatic pipelines to accurately identify low-frequency SNVs in viral populations. Artificial DNA and RNA "populations" were created by introducing known SNVs at predetermined frequencies into template nucleic acid before being sequenced on an Illumina MiSeq platform. These were used to assess the effects of abundance and starting input material type, technical replicates, read length and quality, short-read aligner, and percentage frequency thresholds on the ability to accurately call variants. Analyses revealed that the abundance and type of input nucleic acid had the greatest impact on the accuracy of SNV calling as measured by a micro-averaged Matthews correlation coefficient score, with DNA and high RNA inputs (107 copies) allowing for variants to be called at a 0.2% frequency. Reduced input RNA (105 copies) required more technical replicates to maintain accuracy, while low RNA inputs (103 copies) suffered from consensus-level errors. Base errors identified at specific motifs identified in all technical replicates were also identified which can be excluded to further increase SNV calling accuracy. These findings indicate that samples with low RNA inputs should be excluded for SNV calling and reinforce the importance of optimising the technical and bioinformatics steps in pipelines that are used to accurately identify sequence variants.

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

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.

Prenatal diagnosis of arachnoid cysts: a case series and systematic review.

INTRODUCTION: Primary cysts are believed to arise from the splitting of the arachnoid membrane during prenatal development and can be diagnosed in utero. Prenatal diagnosis is uncommon; therefore, the evidence base for counselling expectant mothers is limited. The purpose of this article is to present a case series and review the current literature on prenatally diagnosed arachnoid cysts. METHOD: A keyword search of hospital electronic records was performed for all patients with a prenatally diagnosed arachnoid cyst at a tertiary neurosurgical centre. Case notes were reviewed for all patients diagnosed between 2005 and 2017. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines were used to structure a systematic review of all English language articles published up to May 2018. RESULTS: A total of eight eligible patients were identified from our own records and 123 from the literature. Sixty-eight per cent of patients had a normal outcome. Sixty-three per cent of patients underwent surgical intervention which was not associated with abnormal outcome. The diagnosis of syndromic/genetic diagnosis (p < 0.001) and the presence of other intra-cranial anatomical abnormalities (p = 0.05) were significant predictors of abnormal outcome. CONCLUSION: The pathogenesis and prognosis of a prenatal arachnoid cyst diagnosis remain unclear. These results suggest favourable outcomes from simple cysts without associated abnormalities and expectant mothers should be counselled accordingly. A wider prospective review is required to better established evidence-based practice.

Neurosurgical management of head injuries incurred during sports: a single centre experience.

Introduction: Accidents during sporting activities are a common cause of head injury, particularly in children and young adults. Whilst most sporting head injuries are minor, there remains a proportion which is associated with high morbidity and mortality. The epidemiology of sports associated head injuries is variable based on geographical region so the aim of this study was to review the management and outcomes of sporting head injuries managed by a single neurosurgical unit in the South of England.Method: A retrospective review of the Trauma Audit and Research Network database was conducted for all patients admitted to a tertiary neurosurgical centre over a six-year period (January 2011-December 2016). Case notes were reviewed for demographics, mechanism of injury, injury severity score, intensive care admission, surgical interventions and Glasgow Outcome Score at discharge.Results: Seventy-six patients (mean age: 37.6 ± 18.4 years, male gender n = 43; 56.6%) were eligible for inclusion in this series. Horse riding accidents were identified as the most common cause of head injury (n = 31; 40.8%). Fifteen patients (19.7%) in this series had a severe head injury (GCS 3-8 on admission). Twenty-eight (36.8%) patients required admission to an intensive care unit and 26 (34.2%) patients underwent neurosurgical intervention. At discharge, 68 (89.5%) patients had a Glasgow Outcome Score 4-5.Conclusion: The majority of patients with head injuries admitted to a neurosurgical unit can expect a good functional outcome despite the need for intensive care or neurosurgical intervention. The range of sports resulting in head injury is likely influenced by geographic location; however, further national study is required for wider comparison.

Paediatric brain abscesses: a single centre experience.

Introduction: Brain parenchymal abscesses are relatively infrequent but potentially serious infections in the paediatric population. Surgical intervention in addition to a prolonged administration of antibiotics is generally appropriate management. This study presents our centre's experience of managing such patients in the context of relevant literature. Method: A single-centre retrospective case note review was conducted over a 15 year period (2003-2017). Patients were selected from electronic hospital records using ICD10 code G06.0. Patients < 18 years of age with a confirmed intra-parenchymal abscess were included. Patient records were reviewed for abscess location, microbiology results, surgical intervention, and outcome using the Glasgow Outcome Score at 3 months. Results: Twenty-four patients were identified (mean age: 7.4 ± 5.3 years, male n = 11). Twelve (50.0%) patients had an abscess in the frontal lobe and Streptococcus was the most common causative microorganism (n = 15). Nineteen patients (79.2%) had an identifiable source which included: ENT infections, congenital cardiac malformations, recent dental surgery and meningitis. All 24 patients underwent surgery with 20 patients having a total of 32 aspirations between them and the other 4 having craniotomy and excision. Twenty patients had 3 month follow-up data of which 18 patients scored GOS: 5, one was GOS: 4 and one was GOS: 3. Conclusions: Brain parenchymal abscess remains an uncommon pathology in the paediatric population. The majority of patients have a preceding infection with Streptococcus as the most common causative organism. Antimicrobial therapy should be selected accordingly. All of our patients underwent surgical intervention and received intravenous antibiotics with favourable outcome and no mortality.

Natural History of Intracranial Arachnoid Cysts.

BACKGROUND: Intracranial arachnoid cysts are a common incidental finding. Most of these remain asymptomatic with surgical intervention being reserved for those cysts causing symptoms from mass effect or hydrocephalus. It is not properly understood why cysts enlarge nor is it possible to predict in which cysts this will occur. The aim of this study is to review the natural history of incidental arachnoid cysts. METHODS: A retrospective case review was conducted of all patients diagnosed with an arachnoid cyst between 2007 and 2016. Case notes and radiology reports were reviewed for demographics, symptoms, and changes in cyst sizes. RESULTS: Four hundred and eighty-five patients were diagnosed with an arachnoid cyst during this time period; of these, 425 (87.6%) were asymptomatic. Patients with symptomatic cysts were significantly younger and more likely to have associated hydrocephalus. The most common indications for imaging which resulted in the diagnosis of the cyst were headaches (n = 106), stroke (n = 51), and seizures (n = 41). The cyst size remained stable or reduced in 147 patients (99.3%) with follow-up imaging (n = 148), and the enlarging cyst remained asymptomatic. CONCLUSIONS: Asymptomatic arachnoid cysts have a low rate of cyst enlargement on follow-up imaging, which supports not routinely imaging these patients. However, serial imaging and surgery may still be indicated in asymptomatic patients at risk of obstructive hydrocephalus which, in this series, was only seen in the pediatric population. A better understanding of the natural history of intracranial arachnoid cysts allows for more efficient follow-up planning.