Item Published version Open AccessOsteoblast differentiation of equine induced pluripotent stem cells.(The Company of Biologists, 2018-05-10) Baird, Arabella; Lindsay, Timothy; Everett, Alice; Iyemere, Valentine; Paterson, Yasmin Z; McClellan, Alyce; Henson, Frances MD; Guest, Deborah J; Baird, Arabella [0000-0003-2689-1481]; Everett, Alice [0000-0002-1679-3936]; Guest, Deborah J [0000-0002-0034-3332]Bone fractures occur in horses following traumatic and non-traumatic (bone overloading) events. They can be difficult to treat due to the need for the horse to bear weight on all legs during the healing period. Regenerative medicine to improve fracture union and recovery could significantly improve horse welfare. Equine induced pluripotent stem cells (iPSCs) have previously been derived. Here we show that equine iPSCs cultured for 21 days in osteogenic induction media on an OsteoAssay surface upregulate the expression of osteoblast associated genes and proteins, including COL1A1, SPARC, SPP1, IBSP, RUNX2 and BGALP We also demonstrate that iPSC-osteoblasts are able to produce a mineralised matrix with both calcium and hydroxyapatite deposition. Alkaline phosphatase activity is also significantly increased during osteoblast differentiation. Although the genetic background of the iPSC donor animal affects the level of differentiation observed after 21 days of differentiation, less variation between lines of iPSCs derived from the same horse was observed. The successful, direct, differentiation of equine iPSCs into osteoblasts may provide a source of cells for future regenerative medicine strategies to improve fracture repair in horses undergoing surgery. iPSC-derived osteoblasts will also provide a potential tool to study equine bone development and disease. Item Accepted version Open AccessModeling infectious disease dynamics in the complex landscape of global health.(American Association for the Advancement of Science (AAAS), 2015-03-13) Heesterbeek, Hans; Anderson, Roy M; Andreasen, Viggo; Bansal, Shweta; De Angelis, Daniela; Dye, Chris; Eames, Ken TD; Edmunds, W John; Frost, Simon DW; Funk, Sebastian; Hollingsworth, T Deirdre; House, Thomas; Isham, Valerie; Klepac, Petra; Lessler, Justin; Lloyd-Smith, James O; Metcalf, C Jessica E; Mollison, Denis; Pellis, Lorenzo; Pulliam, Juliet RC; Roberts, Mick G; Viboud, Cecile; Isaac Newton Institute IDD Collaboration; De Angelis, Daniela [0000-0001-6619-6112]; Frost, Simon [0000-0002-5207-9879]Despite some notable successes in the control of infectious diseases, transmissible pathogens still pose an enormous threat to human and animal health. The ecological and evolutionary dynamics of infections play out on a wide range of interconnected temporal, organizational, and spatial scales, which span hours to months, cells to ecosystems, and local to global spread. Moreover, some pathogens are directly transmitted between individuals of a single species, whereas others circulate among multiple hosts, need arthropod vectors, or can survive in environmental reservoirs. Many factors, including increasing antimicrobial resistance, increased human connectivity and changeable human behavior, elevate prevention and control from matters of national policy to international challenge. In the face of this complexity, mathematical models offer valuable tools for synthesizing information to understand epidemiological patterns, and for developing quantitative evidence for decision-making in global health. Item Accepted version Open AccessMicrobiological quality and antimicrobial resistance characterization of Salmonella spp. in fresh milk value chains in Ghana.(Elsevier BV, 2018-07-20) Parry-Hanson Kunadu, Angela; Holmes, Mark; Miller, Eric L; Grant, Andrew J; Holmes, Mark [0000-0002-5454-1625]; Miller, Eric [0000-0002-7157-6213]; Grant, Andrew [0000-0001-9746-2989]Consumer perception of poor hygiene of fresh milk products is a major barrier to promotion of milk consumption as an intervention to alleviate the burden of malnutrition in Ghana. Fresh milk is retailed raw, boiled, or processed into unfermented cheese and spontaneously fermented products in unlicensed outlets. In this study, we have determined microbiological quality of informally retailed fresh milk products and characterized the genomic diversity and antimicrobial resistance (AMR) patterns of non-typhoidal Salmonella (NTS) in implicated products. A total of 159 common dairy products were purchased from five traditional milk markets in Accra. Samples were analysed for concentrations of aerobic bacteria, total and fecal coliforms, Escherichia coli, staphylococci, lactic acid bacteria and yeast and moulds. The presence of Salmonella, E. coli O157:H7, Listeria monocytogenes and Staphylococcus aureus were determined. AMR of Salmonella against 18 antibiotics was experimentally determined. Genome sequencing of 19 Salmonella isolates allowed determination of serovars, antigenic profiles, prediction of AMR genes in silico and inference of phylogenetic relatedness between strains. Raw and heat-treated milk did not differ significantly in overall bacterial quality (P = 0.851). E. coli O157:H7 and Staphylococcus aureus were present in 34.3% and 12.9% of dairy products respectively. Multidrug resistant (MDR) Salmonella enterica serovars Muenster and Legon were identified in 11.8% and 5.9% of unfermented cheese samples respectively. Pan genome analysis revealed a total of 3712 core genes. All Salmonella strains were resistant to Trimethoprim/Sulfamethoxazole, Cefoxitin, Cefuroxime Axetil and Cefuroxime. Resistance to Chloramphenicol (18%) and Ciprofloxacin (100%), which are first line antibiotics used in treatment of NTS bacteremia in Ghana, was evident. AMR was attributed to presence and/or mutations in the following genes: golS, sdiA for cephalosporins, aac(6')-Iy, ant(9) for aminoglycosides, mdtK, gyrA, gyrB, parC, parE for quinolones and cat1, cat4 for phenicols. Phylogenetic analysis based on accessory genes clustered S. Legon strains separately from the S. Muenster strains. These strains were from different markets suggesting local circulation of related strains. Our study justifies consumer resistance to consumption of unripened soft cheese without further lethal heat treatment, and provides evidence that supports the Ghana Health Service recommendation for use of 3rd generation cephalosporins for the treatment of MDR NTS infections. Item Accepted version Open AccessHereditary selective cobalamin malabsorption and concurrent pancreatitis in a young Border collie(BMJ, 2018-03-01) McCallum, KE; Watson, PJ; McCallum, Katie [0000-0001-6153-0687]; Watson, Penelope [0000-0002-7241-9412]A one-year-old neutered male Border collie presented with failure to gain weight, lethargy, intermittent leucopenia, borderline anaemia and intermittent gastrointestinal symptoms. He was diagnosed with pancreatitis based on blood results and abdominal ultrasonography and hereditary selective cobalamin malabsorption based on hypocobalaminaemia, methylmalonic aciduria and genetic testing for cubilin mutation. The dog responded to oral cobalamin supplementation with resolution of clinical signs and normalisation of serum cobalamin. There was no recurrence of signs after 27 months of follow-up. An association between organic acidaemias and pancreatitis has been reported in humans but to the authors' knowledge, this is the first report of hereditary selective cobalamin malabsorption and concurrent pancreatitis in a dog. Furthermore, this is the first report of inherited canine cobalamin deficiency responding to oral cobalamin supplementation. Item Published version Open AccessCharacterizing Nanoparticles in Biological Matrices: Tipping Points in Agglomeration State and Cellular Delivery In Vitro.(American Chemical Society (ACS), 2017-12-26) Wills, John W; Summers, Huw D; Hondow, Nicole; Sooresh, Aishwarya; Meissner, Kenith E; White, Paul A; Rees, Paul; Brown, Andy; Doak, Shareen H; Wills, John W [0000-0002-4347-5394]Understanding the delivered cellular dose of nanoparticles is imperative in nanomedicine and nanosafety, yet is known to be extremely complex because of multiple interactions between nanoparticles, their environment, and the cells. Here, we use 3-D reconstruction of agglomerates preserved by cryogenic snapshot sampling and imaged by electron microscopy to quantify the "bioavailable dose" that is presented at the cell surface and formed by the process of individual nanoparticle sequestration into agglomerates in the exposure media. Critically, using 20 and 40 nm carboxylated polystyrene-latex and 16 and 85 nm silicon dioxide nanoparticles, we show that abrupt, dose-dependent "tipping points" in agglomeration state can arise, subsequently affecting cellular delivery and increasing toxicity. These changes are triggered by shifts in the ratio of the total nanoparticle surface area to biomolecule abundance, with the switch to a highly agglomerated state effectively changing the test article midassay, challenging the dose-response paradigm for nanosafety experiments. By characterizing nanoparticle numbers per agglomerate, we show these tipping points can lead to the formation of extreme agglomeration states whereby 90% of an administered dose is contained and delivered to the cells by just the top 2% of the largest agglomerates. We thus demonstrate precise definition, description, and comparison of the nanoparticle dose formed in different experimental environments and show that this description is critical to understanding cellular delivery and toxicity. We further empirically "stress-test" the commonly used dynamic light scattering approach, establishing its limitations to present an analysis strategy that significantly improves the usefulness of this popular nanoparticle characterization technique. Item Published version Open AccessOutbreakTools: a new platform for disease outbreak analysis using the R software.(Elsevier BV, 2014-06) Jombart, Thibaut; Aanensen, David M; Baguelin, Marc; Birrell, Paul; Cauchemez, Simon; Camacho, Anton; Colijn, Caroline; Collins, Caitlin; Cori, Anne; Didelot, Xavier; Fraser, Christophe; Frost, Simon; Hens, Niel; Hugues, Joseph; Höhle, Michael; Opatowski, Lulla; Rambaut, Andrew; Ratmann, Oliver; Soubeyrand, Samuel; Suchard, Marc A; Wallinga, Jacco; Ypma, Rolf; Ferguson, Neil; Birrell, Paul [0000-0001-8131-4893]; Frost, Simon [0000-0002-5207-9879]The investigation of infectious disease outbreaks relies on the analysis of increasingly complex and diverse data, which offer new prospects for gaining insights into disease transmission processes and informing public health policies. However, the potential of such data can only be harnessed using a number of different, complementary approaches and tools, and a unified platform for the analysis of disease outbreaks is still lacking. In this paper, we present the new R package OutbreakTools, which aims to provide a basis for outbreak data management and analysis in R. OutbreakTools is developed by a community of epidemiologists, statisticians, modellers and bioinformaticians, and implements classes and methods for storing, handling and visualizing outbreak data. It includes real and simulated outbreak datasets. Together with a number of tools for infectious disease epidemiology recently made available in R, OutbreakTools contributes to the emergence of a new, free and open-source platform for the analysis of disease outbreaks. Item Published version Open AccessPathogenesis of bat rabies in a natural reservoir: Comparative susceptibility of the straw-colored fruit bat (Eidolon helvum) to three strains of Lagos bat virus.(Public Library of Science (PLoS), 2018-03) Suu-Ire, Richard; Begeman, Lineke; Banyard, Ashley C; Breed, Andrew C; Drosten, Christian; Eggerbauer, Elisa; Freuling, Conrad M; Gibson, Louise; Goharriz, Hooman; Horton, Daniel L; Jennings, Daisy; Kuzmin, Ivan V; Marston, Denise; Ntiamoa-Baidu, Yaa; Riesle Sbarbaro, Silke; Selden, David; Wise, Emma L; Kuiken, Thijs; Fooks, Anthony R; Müller, Thomas; Wood, James LN; Cunningham, Andrew A; Begeman, Lineke [0000-0001-8856-9135]Rabies is a fatal neurologic disease caused by lyssavirus infection. People are infected through contact with infected animals. The relative increase of human rabies acquired from bats calls for a better understanding of lyssavirus infections in their natural hosts. So far, there is no experimental model that mimics natural lyssavirus infection in the reservoir bat species. Lagos bat virus is a lyssavirus that is endemic in straw-colored fruit bats (Eidolon helvum) in Africa. Here we compared the susceptibility of these bats to three strains of Lagos bat virus (from Senegal, Nigeria, and Ghana) by intracranial inoculation. To allow comparison between strains, we ensured the same titer of virus was inoculated in the same location of the brain of each bat. All bats (n = 3 per strain) were infected, and developed neurological signs, and fatal meningoencephalitis with lyssavirus antigen expression in neurons. There were three main differences among the groups. First, time to death was substantially shorter in the Senegal and Ghana groups (4 to 6 days) than in the Nigeria group (8 days). Second, each virus strain produced a distinct clinical syndrome. Third, the spread of virus to peripheral tissues, tested by hemi-nested reverse transcriptase PCR, was frequent (3 of 3 bats) and widespread (8 to 10 tissues positive of 11 tissues examined) in the Ghana group, was frequent and less widespread in the Senegal group (3/3 bats, 3 to 6 tissues positive), and was rare and restricted in the Nigeria group (1/3 bats, 2 tissues positive). Centrifugal spread of virus from brain to tissue of excretion in the oral cavity is required to enable lyssavirus transmission. Therefore, the Senegal and Ghana strains seem most suitable for further pathogenesis, and for transmission, studies in the straw-colored fruit bat. Item Published version Open AccessThe IUPHAR/BPS Guide to PHARMACOLOGY in 2018: updates and expansion to encompass the new guide to IMMUNOPHARMACOLOGY.(Oxford University Press (OUP), 2018-01-04) Harding, Simon D; Sharman, Joanna L; Faccenda, Elena; Southan, Chris; Pawson, Adam J; Ireland, Sam; Gray, Alasdair JG; Bruce, Liam; Alexander, Stephen PH; Anderton, Stephen; Bryant, Clare; Davenport, Anthony P; Doerig, Christian; Fabbro, Doriano; Levi-Schaffer, Francesca; Spedding, Michael; Davies, Jamie A; NC-IUPHAR; Bryant, Clare [0000-0002-2924-0038]; Davenport, Anthony [0000-0002-2096-3117]The IUPHAR/BPS Guide to PHARMACOLOGY (GtoPdb, www.guidetopharmacology.org) and its precursor IUPHAR-DB, have captured expert-curated interactions between targets and ligands from selected papers in pharmacology and drug discovery since 2003. This resource continues to be developed in conjunction with the International Union of Basic and Clinical Pharmacology (IUPHAR) and the British Pharmacological Society (BPS). As previously described, our unique model of content selection and quality control is based on 96 target-class subcommittees comprising 512 scientists collaborating with in-house curators. This update describes content expansion, new features and interoperability improvements introduced in the 10 releases since August 2015. Our relationship matrix now describes ∼9000 ligands, ∼15 000 binding constants, ∼6000 papers and ∼1700 human proteins. As an important addition, we also introduce our newly funded project for the Guide to IMMUNOPHARMACOLOGY (GtoImmuPdb, www.guidetoimmunopharmacology.org). This has been 'forked' from the well-established GtoPdb data model and expanded into new types of data related to the immune system and inflammatory processes. This includes new ligands, targets, pathways, cell types and diseases for which we are recruiting new IUPHAR expert committees. Designed as an immunopharmacological gateway, it also has an emphasis on potential therapeutic interventions. Item Published version Open AccessPrevalence, duration and risk factors for appendicular osteoarthritis in a UK dog population under primary veterinary care.(Springer Science and Business Media LLC, 2018-04-04) Anderson, Katharine L; O'Neill, Dan G; Brodbelt, David C; Church, David B; Meeson, Richard L; Sargan, David; Summers, Jennifer F; Zulch, Helen; Collins, Lisa M; Collins, Lisa M [0000-0002-8596-5498]Osteoarthritis is the most common joint disease diagnosed in veterinary medicine and poses considerable challenges to canine welfare. This study aimed to investigate prevalence, duration and risk factors of appendicular osteoarthritis in dogs under primary veterinary care in the UK. The VetCompassTM programme collects clinical data on dogs attending UK primary-care veterinary practices. The study included all VetCompassTM dogs under veterinary care during 2013. Candidate osteoarthritis cases were identified using multiple search strategies. A random subset was manually evaluated against a case definition. Of 455,557 study dogs, 16,437 candidate osteoarthritis cases were identified; 6104 (37%) were manually checked and 4196 (69% of sample) were confirmed as cases. Additional data on demography, clinical signs, duration and management were extracted for confirmed cases. Estimated annual period prevalence (accounting for subsampling) of appendicular osteoarthritis was 2.5% (CI95: 2.4-2.5%) equating to around 200,000 UK affected dogs annually. Risk factors associated with osteoarthritis diagnosis included breed (e.g. Labrador, Golden Retriever), being insured, being neutered, of higher bodyweight and being older than eight years. Duration calculation trials suggest osteoarthritis affects 11.4% of affected individuals' lifespan, providing further evidence for substantial impact of osteoarthritis on canine welfare at the individual and population level. Item Published version Open AccessMitochondrial damage contributes to Pseudomonas aeruginosa activation of the inflammasome and is downregulated by autophagy.(Informa UK Limited, 2015) Jabir, Majid Sakhi; Hopkins, Lee; Ritchie, Neil D; Ullah, Ihsan; Bayes, Hannah K; Li, Dong; Tourlomousis, Panagiotis; Lupton, Alison; Puleston, Daniel; Simon, Anna Katharina; Bryant, Clare; Evans, Thomas J; Tourlomousis, Panagiotis [0000-0002-6152-8066]; Bryant, Clare [0000-0002-2924-0038]The nucleotide-binding domain, leucine-rich repeat containing family caspase recruitment domain containing 4 (NLRC4) inflammasome can be activated by pathogenic bacteria via products translocated through the microbial type III secretion apparatus (T3SS). Recent work has shown that activation of the NLRP3 inflammasome is downregulated by autophagy, but the influence of autophagy on NLRC4 activation is unclear. We set out to determine how autophagy might influence this process, using the bacterium Pseudomonas aeruginosa, which activates the NLRC4 inflammasome via its T3SS. Infection resulted in T3SS-dependent mitochondrial damage with increased production of reactive oxygen intermediates and release of mitochondrial DNA. Inhibiting mitochondrial reactive oxygen release or degrading intracellular mitochondrial DNA abrogated NLRC4 inflammasome activation. Moreover, macrophages lacking mitochondria failed to activate NLRC4 following infection. Removal of damaged mitochondria by autophagy significantly attenuated NLRC4 inflammasome activation. Mitochondrial DNA bound specifically to NLRC4 immunoprecipitates and transfection of mitochondrial DNA directly activated the NLRC4 inflammasome; oxidation of the DNA enhanced this effect. Manipulation of autophagy altered the degree of inflammasome activation and inflammation in an in vivo model of P. aeruginosa infection. Our results reveal a novel mechanism contributing to NLRC4 activation by P. aeruginosa via mitochondrial damage and release of mitochondrial DNA triggered by the bacterial T3SS that is downregulated by autophagy. Item Accepted version Open AccessUnusual clinical presentation of cryptococcus in an immunocompetent cat(BMJ Journals, 2017-11) Sanchini, Lucia; Reading, Mark; Williams, TL; Archer, Frances; Williams, Timothy [0000-0002-9775-8285]This case report describes an atypical presentation of cryptococcal infection in a cat initially presented with multiple persistent pruritic exudative skin lesions, which did not subside following administration of antibiotics and corticosteroids. Both fungal culture and feline immunodeficiency virus (FIV)/feline leukaemia virus (FeLV) ELISA test yielded negative results. Cytological examination of the skin scrapings was consistent with infection by Cryptococcus, which was confirmed by both postmortem inspection and histopathological examination of the lesions. The observed multifocal skin lesions are the result of haematogenous dissemination of the yeast, which is generally seen in immunocompromised cats. Clinical signs of systemic infection by Cryptococcus include apathy and cachexia and may or may not follow classical nasal disease. Surprisingly, the cat described in this report was immunocompetent, presented in good general condition and with no nasal discharge. Item Accepted version Open AccessUnusual presentation of bilateral stifle osteochondritis dissecans in two labrador retrievers(Wiley, 2017-11) Earley, NF; Piola, V; Radke, H; Salgüero, R; Radke, Heidi [0000-0003-0092-4628]There are few published reports of CT findings for the diagnosis of stifle osteochondritis dissecans (OCD) in the dog. This report describes the radiographic and multidetector CT (MDCT) findings of two cases with bilateral stifle osteochondrosis and OCD lesions. In both cases, radiographs did not provide a complete picture of the pathology and CT was needed for a definitive diagnosis. In case 1, CT showed bilateral defects on the caudomedial part of the lateral femoral condyles with joint mice within the cranial right stifle joint. In case 2, CT showed large bilateral joint mice within the intercondylar fossa originating from the medial aspect of the lateral femoral condyles as well as a defect on the right medial femoral condyle. MDCT provided an accurate diagnosis in both these cases and facilitated surgical planning. Item Published version Open AccessLigand-Doped Copper Oxo-hydroxide Nanoparticles are Effective Antimicrobials.(Springer, 2018-04-19) Passos Bastos, Carlos; Rodrigues Faria, Nuno; Ivask, Angela; Bondarenko, Olesja M; Kahru, Anne; Powell, Jonathan; Bastos, Carlos [0000-0003-3528-2178]; Ivask, Angela [0000-0003-3451-7211]; Powell, Jonathan [0000-0003-2738-1715]Bacterial resistance to antimicrobial therapies is an increasing clinical problem. This is as true for topical applications as it is for systemic therapy. Topically, copper ions may be effective and cheap antimicrobials that act through multiple pathways thereby limiting opportunities to bacteria for resistance. However, the chemistry of copper does not lend itself to facile formulations that will readily release copper ions at biologically compatible pHs. Here we have developed nanoparticulate copper hydroxide adipate tartrate (CHAT) as a cheap, safe and readily synthesised material that should enable antimicrobial copper ion release in an infected wound environment. First, we synthesised CHAT and showed that this had disperse aquated particle sizes of 2-5 nm, and mean zeta potential of -40 mV. Next, when diluted into bacterial medium, CHAT demonstrated similar efficacy to copper chloride against Escherichia coli and Staphylococcus aureus, with dose-dependent activity occurring mostly around 12.5-50 mg/L of copper. Indeed, at these levels, CHAT very rapidly dissolved and, as confirmed by a bacterial copper biosensor, showed identical intracellular loading to copper ions derived from copper chloride. However, when formulated at 250 mg/L in a topically-applied matrix, namely hydroxyethyl cellulose, the benefit of CHAT over copper chloride was apparent. The former yielded rapid sustained release of copper within the bactericidal range but the copper chloride, which formed insoluble precipitates at such concentration and pH, achieved a maximum release of 10 ± 7 mg/L copper by 24 hours. We provide a practical formulation for topical copper - based antimicrobial therapy. Further studies, especially in vivo, are merited. Item Published version Open AccessClinical, histological, immunohistochemical and genetic factors associated with measurable response of high‑risk canine mast cell tumours to tyrosine kinase inhibitors(Spandidos Publications, 2018-01) Dobson, JM; Dobson, Jane [0000-0002-7121-014X]Abstract. The aim of the present prospective‑retrospective study was to evaluate the response of high‑risk canine mast cell tumours (MCTs) to tyrosine kinase inhibitors (TKIs) and to correlate this with prognostic factors. A total of 24 dogs presented with macroscopic cutaneous MCTs at disease stage II or III, and therefore, at high‑risk of associated mortality, were included in the study and treated with masitinib (n=20) or toceranib (n=4). A total of 12/24 dogs achieved an objective response and the overall survival (OS) for all subjects was 113 days. Dogs responding to treatment had a significant increase in OS compared to non‑responders (146.5 days vs. 47 days, P=0.02). Internal tandem duplications in exon 11 of the c‑kit gene were identified in 6/24 cases. Ki67, KIT immunolabelling and c‑kit mutation did not provide information regarding prognosis or prediction of response to TKIs in this population. Initial response to TKIs appears to be the most reliable prognostic factor for survival duration. Item Published version Open AccessThe role of venues in structuring HIV, sexually transmitted infections, and risk networks among men who have sex with men.(BioMed Central, 2018-02-07) Drumright, Lydia; Weir, Sharon S; Frost, Simon; Drumright, Lydia [0000-0002-3361-8080]; Frost, Simon [0000-0002-5207-9879]Background Venues form part of the sampling frame for time-location sampling, an approach often used for HIV surveillance. While sampling location is often regarded as a nuisance factor, venues may play a central role in structuring risk networks. We investigated individual reports of risk behaviors and infections among men who have sex with men (MSM) attending different venues to examine structuring of HIV risk behaviors. However, teasing apart ‘risky people’ from ‘risky places’ is difficult, as individuals cannot be randomized to attend different venues. However, we can emulate this statistically using marginal structural models, which inversely weight individuals according to their estimated probability of attending the venue. Methods We conducted a cross-sectional survey of 609 MSM patrons of 14 bars in San Diego, California, recruited using the Priorities for Local AIDS Control Efforts (PLACE) methodology, which consists of a multi-level identification and assessment of venues for HIV risk through population surveys. Results and Discussion Venues differed by many factors, including participants’ reported age, ethnicity, number of lifetime male partners, past sexually transmitted infection (STI), and HIV status. In multivariable marginal structural models, venues demonstrated structuring of HIV+ status, past STI, and methamphetamine use, independently of individual-level characteristics. Conclusions Studies using time-location sampling should consider venue as an important covariate, and the use of marginal structural models may help to identify risky venues. This may assist in widespread, economically feasible and sustainable targeted surveillance and prevention. A more mechanistic understanding of how 'risky venues' emerge and structure risk is needed. Item Published version Open AccessMeningococcal carriage within households in the African meningitis belt: A longitudinal pilot study.(Elsevier BV, 2018-02) Basta, Nicole E; Berthe, Abdoulaye; Keita, Mahamadou; Onwuchekwa, Uma; Tamboura, Boubou; Traore, Awa; Hassan-King, Musa; Manigart, Olivier; Nascimento, Maria; Stuart, James M; Trotter, Caroline; Blake, Jayne; Carr, Anthony D; Gray, Stephen J; Newbold, Lynne S; Deng, Yangqing; Wolfson, Julian; Halloran, M Elizabeth; Greenwood, Brian; Borrow, Ray; Sow, Samba O; Trotter, Caroline [0000-0003-4000-2708]OBJECTIVES: Carriers of Neisseria meningitidis are a key source of transmission. In the African meningitis belt, where risk of meningococcal disease is highest, a greater understanding of meningococcal carriage dynamics is needed. METHODS: We randomly selected an age-stratified sample of 400 residents from 116 households in Bamako, Mali, and collected pharyngeal swabs in May 2010. A month later, we enrolled all 202 residents of 20 of these households (6 with known carriers) and collected swabs monthly for 6 months prior to MenAfriVac vaccine introduction and returned 10 months later to collect swabs monthly for 3 months. We used standard bacteriological methods to identify N. meningitidis carriers and fit hidden Markov models to assess acquisition and clearance overall and by sex and age. RESULTS: During the cross-sectional study 5.0% of individuals (20/400) were carriers. During the longitudinal study, 73 carriage events were identified from 1422 swabs analyzed, and 16.3% of individuals (33/202) were identified as carriers at least once. The majority of isolates were non-groupable; no serogroup A carriers were identified. CONCLUSIONS: Our results suggest that the duration of carriage with any N. meningitidis averages 2.9 months and that males and children acquire and lose carriage more frequently in an urban setting in Mali. Our study informed the design of a larger study implemented in seven countries of the African meningitis belt. Item Accepted version Open AccessEvolutionary routes and KRAS dosage define pancreatic cancer phenotypes.(Springer Nature, 2018-02-01) Mueller, Sebastian; Engleitner, Thomas; Maresch, Roman; Zukowska, Magdalena; Lange, Sebastian; Kaltenbacher, Thorsten; Konukiewitz, Björn; Öllinger, Rupert; Zwiebel, Maximilian; Strong, Alex; Yen, Hsi-Yu; Banerjee, Ruby; Louzada, Sandra; Fu, Beiyuan; Seidler, Barbara; Götzfried, Juliana; Schuck, Kathleen; Hassan, Zonera; Arbeiter, Andreas; Schönhuber, Nina; Klein, Sabine; Veltkamp, Christian; Friedrich, Mathias; Rad, Lena; Barenboim, Maxim; Ziegenhain, Christoph; Hess, Julia; Dovey, Oliver M; Eser, Stefan; Parekh, Swati; Constantino-Casas, Fernando; de la Rosa, Jorge; Sierra, Marta I; Fraga, Mario; Mayerle, Julia; Klöppel, Günter; Cadiñanos, Juan; Liu, Pentao; Vassiliou, George; Weichert, Wilko; Steiger, Katja; Enard, Wolfgang; Schmid, Roland M; Yang, Fengtang; Unger, Kristian; Schneider, Günter; Varela, Ignacio; Bradley, Allan; Saur, Dieter; Rad, Roland; Vassiliou, George [0000-0003-4337-8022]; Bradley, Allan [0000-0002-2349-8839]The poor correlation of mutational landscapes with phenotypes limits our understanding of the pathogenesis and metastasis of pancreatic ductal adenocarcinoma (PDAC). Here we show that oncogenic dosage-variation has a critical role in PDAC biology and phenotypic diversification. We find an increase in gene dosage of mutant KRAS in human PDAC precursors, which drives both early tumorigenesis and metastasis and thus rationalizes early PDAC dissemination. To overcome the limitations posed to gene dosage studies by the stromal richness of PDAC, we have developed large cell culture resources of metastatic mouse PDAC. Integration of cell culture genomes, transcriptomes and tumour phenotypes with functional studies and human data reveals additional widespread effects of oncogenic dosage variation on cell morphology and plasticity, histopathology and clinical outcome, with the highest KrasMUTlevels underlying aggressive undifferentiated phenotypes. We also identify alternative oncogenic gains (Myc, Yap1 or Nfkb2), which collaborate with heterozygous KrasMUTin driving tumorigenesis, but have lower metastatic potential. Mechanistically, different oncogenic gains and dosages evolve along distinct evolutionary routes, licensed by defined allelic states and/or combinations of hallmark tumour suppressor alterations (Cdkn2a, Trp53, Tgfβ-pathway). Thus, evolutionary constraints and contingencies direct oncogenic dosage gain and variation along defined routes to drive the early progression of PDAC and shape its downstream biology. Our study uncovers universal principles of Ras-driven oncogenesis that have potential relevance beyond pancreatic cancer. Item Published version Open AccessIdentification of LukPQ, a novel, equid-adapted leukocidin of Staphylococcus aureus.(Springer Science and Business Media LLC, 2017-01-20) Koop, Gerrit; Vrieling, Manouk; Storisteanu, Daniel ML; Lok, Laurence SC; Monie, Tom; van Wigcheren, Glenn; Raisen, Claire; Ba, Xiaoliang; Gleadall, Nicholas; Hadjirin, Nazreen; Timmerman, Arjen J; Wagenaar, Jaap A; Klunder, Heleen M; Fitzgerald, J Ross; Zadoks, Ruth; Paterson, Gavin K; Torres, Carmen; Waller, Andrew S; Loeffler, Anette; Loncaric, Igor; Hoet, Armando E; Bergström, Karin; De Martino, Luisa; Pomba, Constança; de Lencastre, Hermínia; Ben Slama, Karim; Gharsa, Haythem; Richardson, Emily J; Chilvers, Edwin R; de Haas, Carla; van Kessel, Kok; van Strijp, Jos AG; Harrison, Ewan M; Holmes, Mark A; Lok, Laurence [0000-0002-9364-4213]; Monie, Tom [0000-0003-4097-1680]; Ba, Xiaoliang [0000-0002-3882-3585]; Chilvers, Edwin [0000-0002-4230-9677]; Harrison, Ewan [0000-0003-2720-0507]; Holmes, Mark [0000-0002-5454-1625]Bicomponent pore-forming leukocidins are a family of potent toxins secreted by Staphylococcus aureus, which target white blood cells preferentially and consist of an S- and an F-component. The S-component recognizes a receptor on the host cell, enabling high-affinity binding to the cell surface, after which the toxins form a pore that penetrates the cell lipid bilayer. Until now, six different leukocidins have been described, some of which are host and cell specific. Here, we identify and characterise a novel S. aureus leukocidin; LukPQ. LukPQ is encoded on a 45 kb prophage (ΦSaeq1) found in six different clonal lineages, almost exclusively in strains cultured from equids. We show that LukPQ is a potent and specific killer of equine neutrophils and identify equine-CXCRA and CXCR2 as its target receptors. Although the S-component (LukP) is highly similar to the S-component of LukED, the species specificity of LukPQ and LukED differs. By forming non-canonical toxin pairs, we identify that the F-component contributes to the observed host tropism of LukPQ, thereby challenging the current paradigm that leukocidin specificity is driven solely by the S-component. Item Published version Open AccessModelling the dynamics of an experimental host-pathogen microcosm within a hierarchical Bayesian framework.(Public Library of Science (PLoS), 2013) Lunn, David; Goudie, Robert JB; Wei, Chen; Kaltz, Oliver; Restif, Olivier; Goudie, Robert [0000-0001-9554-1499]; Restif, Olivier [0000-0001-9158-853X]The advantages of Bayesian statistical approaches, such as flexibility and the ability to acknowledge uncertainty in all parameters, have made them the prevailing method for analysing the spread of infectious diseases in human or animal populations. We introduce a Bayesian approach to experimental host-pathogen systems that shares these attractive features. Since uncertainty in all parameters is acknowledged, existing information can be accounted for through prior distributions, rather than through fixing some parameter values. The non-linear dynamics, multi-factorial design, multiple measurements of responses over time and sampling error that are typical features of experimental host-pathogen systems can also be naturally incorporated. We analyse the dynamics of the free-living protozoan Paramecium caudatum and its specialist bacterial parasite Holospora undulata. Our analysis provides strong evidence for a saturable infection function, and we were able to reproduce the two waves of infection apparent in the data by separating the initial inoculum from the parasites released after the first cycle of infection. In addition, the parameter estimates from the hierarchical model can be combined to infer variations in the parasite's basic reproductive ratio across experimental groups, enabling us to make predictions about the effect of resources and host genotype on the ability of the parasite to spread. Even though the high level of variability between replicates limited the resolution of the results, this Bayesian framework has strong potential to be used more widely in experimental ecology. Item Accepted version Open AccessEffects of a new lumbosacral distraction-stabilization device on lumbosacral kinematics in the dog.Zindl, Claudia; Litsky, Alan; Fitzpatrick, Noel; Crawford, Neil; Allen, MJ; Allen, Matthew [0000-0001-8535-3937]Objective: To determine the biomechanical behaviour of a novel distraction-stabilization system, consisting of an intervertebral distraction bolt, polyaxial screws and connecting rods, in the canine lumbosacral spine. Study design: Biomechanical study. Sample population: Cadaveric canine lumbosacral spines (L4-Cd3) (N=8) Methods: Cadaveric lumbosacral spines were harvested, stripped of musculature, mounted on a 4-point bending jig, and tested in extension, flexion and lateral bending using non-destructive compressive axial loads (0-150N). Angular displacement was recorded from reflective optical trackers rigidly secured to L6, L7 and S1. Data for primary and coupled motion were collected from intact spines; after destabilization at L7-S1, and following surgical stabilisation with the new implant system. Results: As compared with the intact spine, laminectomy resulted in a modest increase in angular displacement at L6-L7 and a marked increase at L7-S1. Instrumentation significantly reduced motion at the operated level (L7-S1) with a concomitant increase at the adjacent level (L6-L7). Conclusion: The combination of a polyaxial pedicle screw-rod system and intervertebral spacer provides a versatile solution of surgical stabilisation of the lumbosacral joint following surgical decompression in the canine lumbosacral spine. The increase in motion at L6-L7 may suggest the potential for adjacent level effects and clinical trials should be designed to address this question. Clinical relevance: These results support the feasibility of using this new implant system for the management of degenerative lumbosacral disease in dogs. The increase in motion at L6-L7 may suggest the potential for adjacent level effects and clinical trials should be designed to address this question.