Genetic Determinants of Thiazide-Induced Hyperuricemia, Hyperglycemia, and Urinary Electrolyte Disturbances - A Genome-Wide Evaluation of the UK Biobank.

Thiazide diuretics, widely used in hypertension, cause a variety of adverse reactions, including hyperglycemia, hyperuricemia, and electrolyte abnormalities. In this study, we aimed to identify genetic variants that interact with thiazide-use to increase the risk of these adverse reactions. Using UK Biobank data, we first performed genomewide variance quantitative trait locus (vQTL) analysis of ~ 6.2 million SNPs on 95,493 unrelated hypertensive White British participants (24,313 on self-reported bendroflumethiazide treatment at recruitment) for 2 blood (glucose and urate) and 2 urine (potassium and sodium) biomarkers. Second, we conducted direct gene-environment interaction (GEI) tests on the significant (P < 2.5 × 10-9) vQTLs, included a second UK Biobank cohort comprising 13,647 unrelated hypertensive White British participants (3,478 on thiazides other than bendroflumethiazide) and set significance at P = 0.05 divided by the number of vQTL SNPs tested for GEIs. The vQTL analysis identified eight statistically significant SNPs for blood glucose (5 SNPs) and serum urate (3 SNPs), with none being identified for the urinary biomarkers. Two of the SNPs (1 glucose SNP: CDKAL1 intron rs35612982, GEI P = 6.24 × 10-3; and 1 serum urate SNP: SLC2A9 intron rs938564, GEI P = 4.51 × 10-4) demonstrated significant GEI effects in the first, but not the second, cohort. Both genes are biologically plausible candidates, with the SLC2A9-mediated interaction having been previously reported. In conclusion, we used a two-stage approach to detect two biologically plausible genetic loci that can interact with thiazides to increase the risk of thiazide-associated biochemical abnormalities. Understanding how environmental exposures (including medications such as thiazides) and genetics interact, is an important step toward precision medicine and improved patient outcomes.

A "Bundle of Care" to Improve Anticoagulation Control in Patients Receiving Warfarin in Uganda and South Africa: Protocol for an Implementation Study.

BACKGROUND: The quality of warfarin anticoagulation among Sub-Saharan African patients is suboptimal. This is due to several factors, including a lack of standardized dosing algorithms, difficulty in providing timely international normalized ratio (INR) results, a lack of patient feedback on their experiences with treatment, a lack of education on adherence, and inadequate knowledge and training of health care workers. Low quality of warfarin anticoagulation, expressed as time in therapeutic range (TTR), is associated with higher adverse event rates, including bleeding and thrombosis, and ultimately, increased morbidity and mortality. Processes and interventions that improve this situation are urgently needed. OBJECTIVE: This study aims to evaluate the implementation of the "warfarin bundle," a package of interventions to improve the quality of anticoagulation and thereby clinical outcomes. The primary outcome for this study is TTR over the initial 3 months of warfarin therapy. METHODS: Patients aged 18 years or older who are newly initiated on warfarin for venous thromboembolism, atrial fibrillation, or valvular heart disease will be enrolled and followed up for 3 months at clinics in Cape Town, South Africa, and Kampala, Uganda, where the warfarin bundle is implemented. A retrospective review of the clinical records of patients on warfarin treatment before implementation (controls) will be used for comparison. This study uses a mixed methods approach of the implementation of patient- and process-centered activities to improve the quality of anticoagulation. Patient-centered activities include the use of clinical dosing algorithms, adherence support, and root cause analysis, whereas process-centered activities include point-of-care INR testing, staff training, and patient education and training. We will assess the impact of these interventions by comparing the TTR and safety outcomes across the 2 groups, as well as the cost-effectiveness and acceptability of the package. RESULTS: We started recruitment in June 2021 and stopped in August 2022, having recruited 167 participants. We obtained ethics approval from the University of Cape Town Faculty of Health Sciences Human Research Ethics Committee, the Provincial Health Research Committees in South Africa, the Joint Clinical Research Centre Institutional Review Board, Kampala, and the University of Liverpool Research Ethics Committee. As of February 2023, data cleaning and formal analysis are underway. We expect to publish the full results by December 2023. CONCLUSIONS: We anticipate that the "bundle of care," which includes a clinical algorithm to guide individualized dosing of warfarin, will improve INR control and TTR of patients in Uganda and South Africa. We will use these findings to design a larger, multisite clinical trial across several Sub-Saharan African countries. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/46710.

Drug-Drug-Gene Interactions in Cardiovascular Medicine.

Cardiovascular disease remains a leading cause of both morbidity and mortality worldwide. It is widely accepted that both concomitant medications (drug-drug interactions, DDIs) and genomic factors (drug-gene interactions, DGIs) can influence cardiovascular drug-related efficacy and safety outcomes. Although thousands of DDI and DGI (aka pharmacogenomic) studies have been published to date, the literature on drug-drug-gene interactions (DDGIs, cumulative effects of DDIs and DGIs) remains scarce. Moreover, multimorbidity is common in cardiovascular disease patients and is often associated with polypharmacy, which increases the likelihood of clinically relevant drug-related interactions. These, in turn, can lead to reduced drug efficacy, medication-related harm (adverse drug reactions, longer hospitalizations, mortality) and increased healthcare costs. To examine the extent to which DDGIs and other interactions influence efficacy and safety outcomes in the field of cardiovascular medicine, we review current evidence in the field. We describe the different categories of DDIs and DGIs before illustrating how these two interact to produce DDGIs and other complex interactions. We provide examples of studies that have reported the prevalence of clinically relevant interactions and the most implicated cardiovascular medicines before outlining the challenges associated with dealing with these interactions in clinical practice. Finally, we provide recommendations on how to manage the challenges including but not limited to expanding the scope of drug information compendia, interaction databases and clinical implementation guidelines (to include clinically relevant DDGIs and other complex interactions) and work towards their harmonization; better use of electronic decision support tools; using big data and novel computational techniques; using clinically relevant endpoints, preemptive genotyping; ensuring ethnic diversity; and upskilling of clinicians in pharmacogenomics and personalized medicine.

A genome-wide association study of plasma concentrations of warfarin enantiomers and metabolites in sub-Saharan black-African patients.

Diversity in pharmacogenomic studies is poor, especially in relation to the inclusion of black African patients. Lack of funding and difficulties in recruitment, together with the requirement for large sample sizes because of the extensive genetic diversity in Africa, are amongst the factors which have hampered pharmacogenomic studies in Africa. Warfarin is widely used in sub-Saharan Africa, but as in other populations, dosing is highly variable due to genetic and non-genetic factors. In order to identify genetic factors determining warfarin response variability, we have conducted a genome-wide association study (GWAS) of plasma concentrations of warfarin enantiomers/metabolites in sub-Saharan black-Africans. This overcomes the issue of non-adherence and may have greater sensitivity at genome-wide level, to identify pharmacokinetic gene variants than focusing on mean weekly dose, the usual end-point used in previous studies. Participants recruited at 12 outpatient sites in Uganda and South Africa on stable warfarin dose were genotyped using the Illumina Infinium H3Africa Consortium Array v2. Imputation was conducted using the 1,000 Genomes Project phase III reference panel. Warfarin/metabolite plasma concentrations were determined by high-performance liquid chromatography with tandem mass spectrometry. Multivariable linear regression was undertaken, with adjustment made for five non-genetic covariates and ten principal components of genetic ancestry. After quality control procedures, 548 participants and 17,268,054 SNPs were retained. CYP2C9*8, CYP2C9*9, CYP2C9*11, and the CYP2C cluster SNP rs12777823 passed the Bonferroni-adjusted replication significance threshold (p < 3.21E-04) for warfarin/metabolite ratios. In an exploratory GWAS analysis, 373 unique SNPs in 13 genes, including CYP2C9*8, passed the Bonferroni-adjusted genome-wide significance threshold (p < 3.846E-9), with 325 (87%, all located on chromosome 10) SNPs being associated with the S-warfarin/R-warfarin outcome (top SNP rs11188082, CYP2C19 intron variant, p = 1.55E-17). Approximately 69% of these SNPs were in linkage disequilibrium (r 2 > 0.8) with CYP2C9*8 (n = 216) and rs12777823 (n = 8). Using a pharmacokinetic approach, we have shown that variants other than CYP2C9*2 and CYP2C9*3 are more important in sub-Saharan black-Africans, mainly due to the allele frequencies. In exploratory work, we conducted the first warfarin pharmacokinetics-related GWAS in sub-Saharan Africans and identified novel SNPs that will require external replication and functional characterization before they can be considered for inclusion in warfarin dosing algorithms.

Ethnic Diversity and Warfarin Pharmacogenomics.

Warfarin has remained the most commonly prescribed vitamin K oral anticoagulant worldwide since its approval in 1954. Dosing challenges including having a narrow therapeutic window and a wide interpatient variability in dosing requirements have contributed to making it the most studied drug in terms of genotype-phenotype relationships. However, most of these studies have been conducted in Whites or Asians which means the current pharmacogenomics evidence-base does not reflect ethnic diversity. Due to differences in minor allele frequencies of key genetic variants, studies conducted in Whites/Asians may not be applicable to underrepresented populations such as Blacks, Hispanics/Latinos, American Indians/Alaska Natives and Native Hawaiians/other Pacific Islanders. This may exacerbate health inequalities when Whites/Asians have better anticoagulation profiles due to the existence of validated pharmacogenomic dosing algorithms which fail to perform similarly in the underrepresented populations. To examine the extent to which individual races/ethnicities are represented in the existing body of pharmacogenomic evidence, we review evidence pertaining to published pharmacogenomic dosing algorithms, including clinical utility studies, cost-effectiveness studies and clinical implementation guidelines that have been published in the warfarin field.

Cardiovascular drugs and COVID-19 clinical outcomes: a systematic review and meta-analysis of randomized controlled trials.

AIMS: To update our previously reported systematic review and meta-analysis of observational studies on cardiovascular drug exposure and COVID-19 clinical outcomes by focusing on newly published randomized controlled trials (RCTs). METHODS: More than 500 databases were searched between 1 November 2020 and 2 October 2021 to identify RCTs that were published after our baseline review. One reviewer extracted data with other reviewers verifying the extracted data for accuracy and completeness. RESULTS: After screening 22 414 records, we included 24 and 21 RCTs in the qualitative and quantitative syntheses, respectively. The most investigated drug classes were angiotensin-converting enzyme inhibitors (ACEIs)/angiotensin receptor blocker (ARBs) and anticoagulants, investigated by 10 and 11 studies respectively. In meta-analyses, ACEI/ARBs did not affect hospitalization length (mean difference -0.42, 95% confidence interval [CI] -1.83; 0.98 d, n = 1183), COVID-19 severity (risk ratio/RR 0.90, 95% CI 0.71; 1.15, n = 1661) or mortality (risk ratio [RR] 0.92, 95% CI 0.58; 1.47, n = 1646). Therapeutic anticoagulation also had no effect (hospitalization length mean difference -0.29, 95% CI -1.13 to 0.56 d, n = 1449; severity RR 0.86, 95% CI 0.70; 1.04, n = 2696; and, mortality RR 0.93, 95% CI 0.77; 1.13, n = 5689). Other investigated drug classes were antiplatelets (aspirin, 2 trials), antithrombotics (sulodexide, 1 trial), calcium channel blockers (amlodipine, 1 trial) and lipid-modifying drugs (atorvastatin, 1 trial). CONCLUSION: Moderate- to high-certainty RCT evidence suggests that cardiovascular drugs such as ACEIs/ARBs are not associated with poor COVID-19 outcomes, and should therefore not be discontinued. These cardiovascular drugs should also not be initiated to treat or prevent COVID-19 unless they are needed for an underlying currently approved therapeutic indication.

Stable warfarin dose prediction in sub-Saharan African patients: A machine-learning approach and external validation of a clinical dose-initiation algorithm.

Warfarin remains the most widely prescribed oral anticoagulant in sub-Saharan Africa. However, because of its narrow therapeutic index, dosing can be challenging. We have therefore (a) evaluated and compared the performance of 21 machine-learning techniques in predicting stable warfarin dose in sub-Saharan Black-African patients and (b) externally validated a previously developed Warfarin Anticoagulation in Patients in Sub-Saharan Africa (War-PATH) clinical dose-initiation algorithm. The development cohort included 364 patients recruited from eight outpatient clinics and hospital departments in Uganda and South Africa (June 2018-July 2019). Validation was conducted using an external validation cohort (270 patients recruited from August 2019 to March 2020 in 12 outpatient clinics and hospital departments). Based on the mean absolute error (MAE; mean of absolute differences between the actual and predicted doses), random forest regression (12.07 mg/week; 95% confidence interval [CI], 10.39-13.76) was the best performing machine-learning technique in the external validation cohort, whereas the worst performing technique was model trees (17.59 mg/week; 95% CI, 15.75-19.43). By comparison, the simple, commonly used regression technique (ordinary least squares) performed similarly to more complex supervised machine-learning techniques and achieved an MAE of 13.01 mg/week (95% CI, 11.45-14.58). In summary, we have demonstrated that simpler regression techniques perform similarly to more complex supervised machine-learning techniques. We have also externally validated our previously developed clinical dose-initiation algorithm, which is being prospectively tested for clinical utility.

Cardiovascular drugs and COVID-19 clinical outcomes: A living systematic review and meta-analysis.

AIMS: The aim of this study was to continually evaluate the association between cardiovascular drug exposure and COVID-19 clinical outcomes (susceptibility to infection, disease severity, hospitalization, hospitalization length, and all-cause mortality) in patients at risk of/with confirmed COVID-19. METHODS: Eligible publications were identified from more than 500 databases on 1 November 2020. One reviewer extracted data with 20% of the records independently extracted/evaluated by a second reviewer. RESULTS: Of 52 735 screened records, 429 and 390 studies were included in the qualitative and quantitative syntheses, respectively. The most-reported drugs were angiotensin-converting enzyme inhibitors (ACEIs)/angiotensin receptor blockers (ARBs) with ACEI/ARB exposure having borderline association with confirmed COVID-19 infection (OR 1.14, 95% CI 1.00-1.31). Among COVID-19 patients, unadjusted estimates showed that ACEI/ARB exposure was associated with hospitalization (OR 1.76, 95% CI 1.34-2.32), disease severity (OR 1.40, 95% CI 1.26-1.55) and all-cause mortality (OR 1.22, 95% CI 1.12-1.33) but not hospitalization length (mean difference -0.27, 95% CI -1.36-0.82 days). After adjustment, ACEI/ARB exposure was not associated with confirmed COVID-19 infection (OR 0.92, 95% CI 0.71-1.19), hospitalization (OR 0.93, 95% CI 0.70-1.24), disease severity (OR 1.05, 95% CI 0.81-1.38) or all-cause mortality (OR 0.84, 95% CI 0.70-1.00). Similarly, subgroup analyses involving only hypertensive patients revealed that ACEI/ARB exposure was not associated with confirmed COVID-19 infection (OR 0.93, 95% CI 0.79-1.09), hospitalization (OR 0.84, 95% CI 0.58-1.22), hospitalization length (mean difference -0.14, 95% CI -1.65-1.36 days), disease severity (OR 0.92, 95% CI 0.76-1.11) while it decreased the odds of dying (OR 0.76, 95% CI 0.65-0.88). A similar trend was observed for other cardiovascular drugs. However, the validity of these findings is limited by a high level of heterogeneity and serious risk of bias. CONCLUSION: Cardiovascular drugs are not associated with poor COVID-19 outcomes in adjusted analyses. Patients should continue taking these drugs as prescribed.

Developing and Validating a Clinical Warfarin Dose-Initiation Model for Black-African Patients in South Africa and Uganda.

Warfarin remains the oral anticoagulant of choice in sub-Saharan Africa. However, dosing is challenging due to a highly variable clinical response for a given dose. This study aimed to develop and validate a clinical warfarin dose-initiation model in sub-Saharan Black-African patients. For the development cohort, we used data from 364 patients who were recruited from 8 outpatient clinics and hospital departments in Uganda and South Africa (June 2018-July 2019). Validation was undertaken using the International Warfarin Pharmacogenetics Consortium (IWPC) dataset (690 black patients). Four predictors (age, weight, target International Normalized Ratio range, and HIV status) were included in the final model, which achieved mean absolute errors (MAEs; mean of absolute differences between true dose and dose predicted by the model) of 11.6 (95% confidence interval (CI) 10.4-12.8) and 12.5 (95% CI 11.6-13.4) mg/week in the development and validation cohorts, respectively. Two other clinical models, IWPC and Gage, respectively, obtained MAEs of 12.5 (95% CI 11.3-13.7) and 12.7 (95% CI 11.5-13.8) mg/week in the development cohort, and 12.1 (95% CI 11.2-13.0) and 12.2 (95% CI 11.4-13.1) mg/week in the validation cohort. Compared with fixed dose-initiation, our model decreased the percentage of patients at high risk of suboptimal anticoagulation by 7.5% (1.5-13.7%) and 11.9% (7.1-16.8%) in the development and validation cohorts, respectively. The clinical utility of this model will be tested in a prospective study. Study Highlights WHAT IS THE CURRENT KNOWLEDGE ON THE TOPIC? ☑ Warfarin dosing remains challenging due to a highly variable clinical response for a given dose. WHAT QUESTION DID THIS STUDY ADDRESS? ☑ Can a clinical dose-initiation model be developed and validated for sub-Saharan Black-African patients? WHAT DOES THIS STUDY ADD TO OUR KNOWLEDGE? ☑ We have developed the first warfarin dose-initiation clinical model for Black-African patients in Uganda and South Africa. HOW MIGHT THIS CHANGE CLINICAL PHARMACOLOGY OR TRANSLATIONAL SCIENCE? ☑ We will be implementing and validating this model in a prospective cohort to inform future large-scale implementation. More optimized dosing should improve the quality of warfarin anticoagulation in these two developing countries.

Warfarin dosing algorithms: A systematic review.

AIMS: Numerous algorithms have been developed to guide warfarin dosing and improve clinical outcomes. We reviewed the algorithms available for various populations and the covariates, performances and risk of bias of these algorithms. METHODS: We systematically searched MEDLINE up to 20 May 2020 and selected studies describing the development, external validation or clinical utility of a multivariable warfarin dosing algorithm. Two investigators conducted data extraction and quality assessment. RESULTS: Of 10 035 screened records, 266 articles were included in the review, describing the development of 433 dosing algorithms, 481 external validations and 52 clinical utility assessments. Most developed algorithms were for dose initiation (86%), developed by multiple linear regression (65%) and mostly applicable to Asians (49%) or Whites (43%). The most common demographic/clinical/environmental covariates were age (included in 401 algorithms), concomitant medications (270 algorithms) and weight (229 algorithms) while CYP2C9 (329 algorithms), VKORC1 (319 algorithms) and CYP4F2 (92 algorithms) variants were the most common genetic covariates. Only 26% and 7% algorithms were externally validated and evaluated for clinical utility, respectively, with <2% of algorithm developments and external validations being rated as having a low risk of bias. CONCLUSION: Most warfarin dosing algorithms have been developed in Asians and Whites and may not be applicable to under-served populations. Few algorithms have been externally validated, assessed for clinical utility, and/or have a low risk of bias which makes them unreliable for clinical use. Algorithm development and assessment should follow current methodological recommendations to improve reliability and applicability, and under-represented populations should be prioritized.

Genetic Factors Influencing Warfarin Dose in Black-African Patients: A Systematic Review and Meta-Analysis.

Warfarin is the most commonly used oral anticoagulant in sub-Saharan Africa. Dosing is challenging due to a narrow therapeutic index and high interindividual variability in dose requirements. To evaluate the genetic factors affecting warfarin dosing in black-Africans, we performed a meta-analysis of 48 studies (2,336 patients). Significant predictors for CYP2C9 and stable dose included rs1799853 (CYP2C9*2), rs1057910 (CYP2C9*3), rs28371686 (CYP2C9*5), rs9332131 (CYP2C9*6), and rs28371685 (CYP2C9*11) reducing dose by 6.8, 12.5, 13.4, 8.1, and 5.3 mg/week, respectively. VKORC1 variants rs9923231 (-1639G>A), rs9934438 (1173C>T), rs2359612 (2255C>T), rs8050894 (1542G>C), and rs2884737 (497T>G) decreased dose by 18.1, 21.6, 17.3, 11.7, and 19.6 mg/week, respectively, whereas rs7294 (3730G>A) increased dose by 6.9 mg/week. Finally, rs12777823 (CYP2C gene cluster) was associated with a dose reduction of 12.7 mg/week. Few studies were conducted in Africa, and patient numbers were small, highlighting the need for further work in black-Africans to evaluate genetic factors determining warfarin response.

Exploring the longitudinal dynamics of herd BVD antibody test results using model-based clustering.

Determining the Bovine Viral Diarrhoea (BVD) infection status of cattle herds is a challenge for control and eradication schemes. Given the changing dynamics of BVD  virus (BVDV) antibody responses in cattle, classifying herds based on longitudinal changes in the results of BVDV antibody tests could offer a novel, complementary approach to categorising herds that is less likely than the present system to result in a herd's status changing from year to year, as it is more likely to capture the true exposure dynamics of the farms. This paper describes the dynamics of BVDV antibody test values (measured as percentage positivity (PP)) obtained from 15,500 bovines between 2007 and 2010 from thirty nine cattle herds located in Scotland and Northern England. It explores approaches of classifying herds based on trend, magnitude and shape of their antibody PP trajectories and investigates the epidemiological similarities between farms within the same cluster. Gaussian mixture models were used for the magnitude and shape clustering. Epidemiologically meaningful clusters were obtained. Farm cluster membership depends on clustering approach used. Moderate concordance was found between the shape and magnitude clusters. These methods hold potential for application to enhance control efforts for BVD and other infectious livestock diseases.

An empirical comparison of isolate-based and sample-based definitions of antimicrobial resistance and their effect on estimates of prevalence.

Antimicrobial resistance is primarily a problem in human medicine but there are unquantified links of transmission in both directions between animal and human populations. Quantitative assessment of the costs and benefits of reduced antimicrobial usage in livestock requires robust quantification of transmission of resistance between animals, the environment and the human population. This in turn requires appropriate measurement of resistance. To tackle this we selected two different methods for determining whether a sample is resistant - one based on screening a sample, the other on testing individual isolates. Our overall objective was to explore the differences arising from choice of measurement. A literature search demonstrated the widespread use of testing of individual isolates. The first aim of this study was to compare, quantitatively, sample level and isolate level screening. Cattle or sheep faecal samples (n=41) submitted for routine parasitology were tested for antimicrobial resistance in two ways: (1) "streak" direct culture onto plates containing the antimicrobial of interest; (2) determination of minimum inhibitory concentration (MIC) of 8-10 isolates per sample compared to published MIC thresholds. Two antibiotics (ampicillin and nalidixic acid) were tested. With ampicillin, direct culture resulted in more than double the number of resistant samples than the MIC method based on eight individual isolates. The second aim of this study was to demonstrate the utility of the observed relationship between these two measures of antimicrobial resistance to re-estimate the prevalence of antimicrobial resistance from a previous study, in which we had used "streak" cultures. Boot-strap methods were used to estimate the proportion of samples that would have tested resistant in the historic study, had we used the isolate-based MIC method instead. Our boot-strap results indicate that our estimates of prevalence of antimicrobial resistance would have been considerably lower in the historic study had the MIC method been used. Finally we conclude that there is no single way of defining a sample as resistant to an antimicrobial agent. The method used greatly affects the estimated prevalence of antimicrobial resistance in a sampled population of animals, thus potentially resulting in misleading results. Comparing methods on the same samples allows us to re-estimate the prevalence from other studies, had other methods for determining resistance been used. The results of this study highlight the importance of establishing what the most appropriate measure of antimicrobial resistance is, for the proposed purpose of the results.

Clinical Research Abstracts of the British Equine Veterinary Association Congress 2015.

REASONS FOR PERFORMING STUDY: Although fasciolosis is an important livestock disease worldwide, the public health importance of human fasciolosis has increased in recent years and it is recognised as an important re-emerging zoonotic disease, its epidemiology and pathogenicity in donkeys, and the epidemiological role they may play have not been determined. OBJECTIVES: To investigate the epidemiology and pathogenicity of fasciolosis in donkeys. STUDY DESIGN: Cross-sectional coprological and retrospective post-mortem study. METHODS: Faecal samples collected from 803 randomly selected working donkeys from the central region of Ethiopia were analysed by a sedimentation-centrifugation-flotation technique. Further data on liver-flukes and associated pathologies were obtained by routine post mortem examinations of 112 donkeys, subjected to euthanasia on welfare grounds or died. Data were analysed using a generalised linear model and multivariate binary logistic regression in R statistical package with significance level of statistical tests set at P<0.05. RESULTS: Infection prevalences of 44.4% and 41.9% were obtained in coprologically and post mortem examined donkeys, respectively, irrespective of their age. Both Fasciola hepatica and Fasciola gigantica were identified with the mean infection intensity of 30 flukes. Older donkeys (≥8 years) were found harbouring a significantly higher worm burden (P<0.0001). Gross and histopathologies of hyperplasia and thickening of the bile ducts, fibrosis of large portal areas and irregular bile duct proliferation and hypertrophy were noted. CONCLUSIONS: The high infection prevalence of fasciolosis and the associated hepatic pathologies in working donkeys shows not only the susceptibility of donkeys and the impact it has on their health, but also indicates the important role they can play in the epidemiology of both livestock and human fasciolosis. These further demonstrate the need for these animals to be considered in the overall epidemiological studies and for sound control strategies and prevention of fasciolosis. Ethical animal research: The research underwent ethical review and the use of animals was approved by the Directors of The Donkey Sanctuary. Consent of the owners was obtained to use their animals. SOURCE OF FUNDING: The Donkey Sanctuary. Competing interests: None declared.

Identifying the seasonal origins of human campylobacteriosis.

Human campylobacteriosis exhibits a distinctive seasonality in temperate regions. This paper aims to identify the origins of this seasonality. Clinical isolates [typed by multi-locus sequence typing (MLST)] and epidemiological data were collected from Scotland. Young rural children were found to have an increased burden of disease in the late spring due to strains of non-chicken origin (e.g. ruminant and wild bird strains from environmental sources). In contrast the adult population had an extended summer peak associated with chicken strains. Travel abroad and UK mainland travel were associated with up to 17% and 18% of cases, respectively. International strains were associated with chicken, had a higher diversity than indigenous strains and a different spectrum of MLST types representative of these countries. Integrating empirical epidemiology and molecular subtyping can successfully elucidate the seasonal components of human campylobacteriosis. The findings will enable public health officials to focus strategies to reduce the disease burden.

Field efficacy of praziquantel oral paste against naturally acquired equine cestodes in Ethiopia.

The efficacy of an oral formulation of praziquantel (Equitape, Horse paste, Fort Dodge) in the reduction of cestode egg counts and serum antibody level against Anoplocephala perfoliata was assessed in 44 donkeys under field conditions. The donkeys were confirmed both by faecal examination and serum antibody assessed by an enzyme-linked immunosorbent assay to have natural infection with tapeworms. The donkeys were randomly allocated into treatment (n = 22) and control (n = 22) groups. The treatment group was treated with both praziquantel and ivermectin (Ivomec, Merial) at a dose rate of 1 mg/kg and 200 µg/kg, respectively while the control group was treated only with ivermectin. Faecal samples were collected before treatment (day-0) and 2, 6, 8, 12, and 16 weeks post-treatment while blood samples were collected before treatment and 8 and 16 weeks after treatment and analysed. The results of the study demonstrated that praziquantel paste was highly effective in reducing cestode eggs in donkeys and had an efficacy of more than 99 % until week 16 (day 112). No cestode egg reappearance by 16 weeks post-treatment in any animal in the treatment group was observed while donkeys in the control group continued shedding cestode eggs. The immunological assay also showed a significant reduction in serum antibody level against A. perfoliata in treated donkeys compared to the control group (p = 0.0001). This marked decrease in serum antibody level indicates reduced risk of cestode-associated colic and other gastrointestinal disorders and clinical diseases. No adverse reactions or clinical effects were encountered in any animal within either group throughout the trial period.

Quantifying the sources of variability in equine faecal egg counts: implications for improving the utility of the method.

The faecal egg count (FEC) is the most widely used means of quantifying the nematode burden of horses, and is frequently used in clinical practice to inform treatment and prevention. The statistical process underlying the FEC is complex, comprising a Poisson counting error process for each sample, compounded with an underlying continuous distribution of means between samples. Being able to quantify the sources of variability contributing to this distribution of means is a necessary step towards providing estimates of statistical power for future FEC and FECRT studies, and may help to improve the usefulness of the FEC technique by identifying and minimising unwanted sources of variability. Obtaining such estimates require a hierarchical statistical model coupled with repeated FEC observations from a single animal over a short period of time. Here, we use this approach to provide the first comparative estimate of multiple sources of within-horse FEC variability. The results demonstrate that a substantial proportion of the observed variation in FEC between horses occurs as a result of variation in FEC within an animal, with the major sources being aggregation of eggs within faeces and variation in egg concentration between faecal piles. The McMaster procedure itself is associated with a comparatively small coefficient of variation, and is therefore highly repeatable when a sufficiently large number of eggs are observed to reduce the error associated with the counting process. We conclude that the variation between samples taken from the same animal is substantial, but can be reduced through the use of larger homogenised faecal samples. Estimates are provided for the coefficient of variation (cv) associated with each within animal source of variability in observed FEC, allowing the usefulness of individual FEC to be quantified, and providing a basis for future FEC and FECRT studies.

Equine cestodosis: a sero-epidemiological study of Anoplocephala perfoliata infection in Ethiopia.

A 12/13 kDa antigen, tapeworm ELISA test, developed for use in horses, was used to detect parasite-specific serum antibody, IgG(T), in the serum of donkeys. In a pilot study the 12/13 kDa antigen was tested and proved to detect the antibody, IgG(T), in donkey sera. Blood samples from 797 donkeys, naturally exposed to cestode infection, from four geographical localities were collected and sera were prepared and analysed. There was substantial serological evidence that donkeys were potentially infected with A. perfoliata. A range of ELISA OD values were obtained from the serological assay. Over 26% and 7.5% of the donkeys were moderately and highly infected, respectively, showing at least a 34% sero-prevalence. The rest, 66.1%, were either with low infection intensity or negative for A. perfoliata infection. The risk of infections, both in sero-prevalence and intensity, as determined by ELISA optical density (OD), were highest in the highland areas of Ethiopia where pastures are low-lying and wet, and permanent pasture management is regularly practised. Sex, age and body condition of the donkeys had no significant effect either on prevalence of the infection or on the serum antibody level. These results indicate a risk of intestinal disorders, particularly, colic, associated with A. perfoliata infection in donkeys.

Assessing the probability of acquisition of meticillin-resistant Staphylococcus aureus (MRSA) in a dog using a nested stochastic simulation model and logistic regression sensitivity analysis.

Meticillin-resistant Staphylococcus aureus (MRSA) is an important nosocomial and community-acquired pathogen with zoonotic potential. The relationship between MRSA in humans and companion animals is poorly understood. This study presents a quantitative exposure assessment, based on expert opinion and published data, in the form of a second order stochastic simulation model with accompanying logistic regression sensitivity analysis that aims to define the most important factors for MRSA acquisition in dogs. The simulation model was parameterised using expert opinion estimates, along with published and unpublished data. The outcome of the model was biologically plausible and found to be dominated by uncertainty over variability. The sensitivity analysis, in the form of four separate logistic regression models, found that both veterinary and non-veterinary routes of acquisition of MRSA are likely to be relevant for dogs. The effects of exposure to, and probability of, transmission of MRSA from the home environment were ranked as the most influential predictors in all sensitivity analyses, although it is unlikely that this environmental source of MRSA is independent of alternative sources of MRSA (human and/or animal). Exposure to and transmission from MRSA positive family members were also found to be influential for acquisition of MRSA in pet dogs, along with veterinary clinic attendance and, while exposure to and transmission from the veterinary clinic environment was also found to be influential, it was difficult to differentiate between the importance of independent sources of MRSA within the veterinary clinic. The implementation of logistic regression analyses directly to the input/output relationship within the simulation model presented in this paper represents the application of a variance based sensitivity analysis technique in the area of veterinary medicine and is a useful means of ranking the relative importance of input variables.