A quantitative release assessment for the noncommercial movement of companion animals: risk of rabies reintroduction to the United kingdom.

In 2004, the European Union (EU) implemented a pet movement policy (referred to here as the EUPMP) under EU regulation 998/2003. The United Kingdom (UK) was granted a temporary derogation from the policy until December 2011 and instead has in place its own Pet Movement Policy (Pet Travel Scheme (PETS)). A quantitative risk assessment (QRA) was developed to estimate the risk of rabies introduction to the UK under both schemes to quantify any change in the risk of rabies introduction should the UK harmonize with the EU policy. Assuming 100 % compliance with the regulations, moving to the EUPMP was predicted to increase the annual risk of rabies introduction to the UK by approximately 60-fold, from 7.79 × 10(-5) (5.90 × 10(-5), 1.06 × 10(-4)) under the current scheme to 4.79 × 10(-3) (4.05 × 10(-3), 5.65 × 10(-3)) under the EUPMP. This corresponds to a decrease from 13,272 (9,408, 16,940) to 211 (177, 247) years between rabies introductions. The risks associated with both the schemes were predicted to increase when less than 100 % compliance was assumed, with the current scheme of PETS and quarantine being shown to be particularly sensitive to noncompliance. The results of this risk assessment, along with other evidence, formed a scientific evidence base to inform policy decision with respect to companion animal movement.

Real-time and multiplex real-time polymerase chain reactions for the detection of Bartonella henselae within cat flea, Ctenocephalides felis, samples.

Bartonella henselae (Rhizobiales: Bartonellacae), the agent of cat-scratch disease, is an emerging bacterial pathogen which can be transmitted via infective faecal material of Ctenocephalides felis Bouché (Siphonaptera: Pulicidae). Worldwide, B. henselae has been identified in 1-53% of felines and 2.9-17.4% of fleas. Although culture is the routine method for detection, the procedure is time-consuming and is rarely used for isolation directly from flea vectors. The current study reports the development of a quantitative real-time polymerase chain reaction (qPCR) to detect and quantify B. henselae organisms from vector samples. The qPCR is specific and detects as few as 2.5 genome copies. To enable direct quantification of Bartonella organisms in different vector samples, we developed a qPCR to detect C. felis DNA that also acts as an extraction control. Combining both PCRs into a multiplex format validates B. henselae results when sampling flea populations, although there is a reduction in sensitivity. This reduction might be counteracted by a different combination of probe fluorophores.

The development of a qPCR assay to detect tick (Ixodida) DNA and its implementation for the study of tick-borne pathogen transmission.

Polymerase chain reaction (PCR) analysis is regularly used to detect pathogens within arthropod vectors, but has also been applied to investigate vector DNA. This study details a novel highly sensitive quantitative PCR (qPCR) which detects and quantifies DNA from Ixodes ricinus, the European vector of Anaplasma phagocytophilum. By pairing this with a qPCR to detect A. phagocytophilum, valid comparisons of pathogen load can be made between different sized tick-tissue samples. These qPCRs were validated in I. ricinus that were fed A. phagocytophilum-infected blood using an artificial membrane feeder. Pathogens were detected in the tick haemolymph within 36h, indicating that successful infection had taken place. This study illustrates the application of vector-targeted qPCRs to confirm and validate pathogen load in samples as part of investigations of vector-pathogen interactions.

Angiostrongylus vasorum from South America and Europe represent distinct lineages.

Angiostrongylus vasorum is a nematode parasite of sylvan and domestic species of the family Canidae. It has a broad but patchy distribution worldwide, and there is evidence for geographical spread and increasing incidence of infection in recent years. While historically Angiostrongylus-like nematodes identified in dogs and foxes have been described as A. vasorum in Europe and Angiocaulus raillieti in South America, more recent taxonomic revision has amalgamated these into a single species, A. vasorum. Here we report, for the first time, the molecular characterization of isolates of A. vasorum from Germany, Portugal, Denmark and the United Kingdom on the basis of the mitochondrial COI gene and the second ribosomal internal transcribed spacer. When compared with isolates from Brazil, sequence analysis revealed 2 distinct genotypes. Estimated rates of evolution based on COI sequences for both nematode and host are consistent with the hypothesis that the presence of A. vasorum in South America is a result of an ancient evolutionary event. Angiostrongylus vasorum in South America potentially represents a separate species to that observed in Europe.

Improving care at scale: process evaluation of a multi-component quality improvement intervention to reduce mortality after emergency abdominal surgery (EPOCH trial).

BACKGROUND: Improving the quality and safety of perioperative care is a global priority. The Enhanced Peri-Operative Care for High-risk patients (EPOCH) trial was a stepped-wedge cluster randomised trial of a quality improvement (QI) programme to improve 90-day survival for patients undergoing emergency abdominal surgery in 93 hospitals in the UK National Health Service. METHODS: The aim of this process evaluation is to describe how the EPOCH intervention was planned, delivered and received, at both cluster and local hospital levels. The QI programme comprised of two interventions: a care pathway and a QI intervention to aid pathway implementation, focussed on stakeholder engagement, QI teamwork, data analysis and feedback and applying the model for improvement. Face-to-face training and online resources were provided to support senior clinicians in each hospital (QI leads) to lead improvement. For this evaluation, we collated programme activity data, administered an exit questionnaire to QI leads and collected ethnographic data in six hospitals. Qualitative data were analysed with thematic or comparative analysis; quantitative data were analysed using descriptive statistics. RESULTS: The EPOCH trial did not demonstrate any improvement in survival or length of hospital stay. Whilst the QI programme was delivered as planned at the cluster level, self-assessed intervention fidelity at the hospital level was variable. Seventy-seven of 93 hospitals responded to the exit questionnaire (60 from a single QI lead response on behalf of the team); 33 respondents described following the QI intervention closely (35%) and there were only 11 of 37 care pathway processes that > 50% of respondents reported attempting to improve. Analysis of qualitative data suggests QI leads were often attempting to deliver the intervention in challenging contexts: the social aspects of change such as engaging colleagues were identified as important but often difficult and clinicians frequently attempted to lead change with limited time or organisational resources. CONCLUSIONS: Significant organisational challenges faced by QI leads shaped their choice of pathway components to focus on and implementation approaches taken. Adaptation causing loss of intervention fidelity was therefore due to rational choices made by those implementing change within constrained contexts. Future large-scale QI programmes will need to focus on dedicating local time and resources to improvement as well as on training to develop QI capabilities. EPOCH TRIAL REGISTRATION: ISRCTN80682973 https://doi.org/10.1186/ISRCTN80682973 Registered 27 February 2014 and Lancet protocol 13PRT/7655.

Correction to: Improving care at scale: process evaluation of a multi-component quality improvement intervention to reduce mortality after emergency abdominal surgery (EPOCH trial).

Following the publication of this article [1], the authors reported a number of errors which are given below.

Developmental specificity of the interaction between the locus control region and embryonic or fetal globin genes in transgenic mice with an HS3 core deletion.

The human beta-globin locus control region (LCR) consists of five erythroid-lineage-specific DNase I-hypersensitive sites (HSs) and is required for activation of the beta-globin locus chromatin domain and globin gene expression. Each DNase I-HS of the LCR consists of a highly conserved core element and flanking sequences. To analyze the functional role of the core elements of the HSs, we deleted a 234-bp fragment encompassing the core of HS3 (HS3c) from a beta-globin locus residing on a 248-kb beta-locus yeast artificial chromosome and analyzed its function in F2 progeny of transgenic mice. Human epsilon-globin gene expression was absent at day 10 and severely reduced in the day 12 embryonic erythropoiesis of mice lacking HS3c. In contrast, gamma-globin gene expression was normal in embryonic erythropoiesis but it was absent in definitive erythropoiesis in the fetal liver. These results indicate that the core element of HS3 is necessary for epsilon-globin gene transcription in embryonic cells and for gamma-globin gene transcription in definitive cells. Normal gamma-globin gene expression in embryonic cells and the absence of gamma-globin gene expression in definitive cells show that different HSs interact with gamma-globin gene promoters in these two stages of development. Such results provide direct evidence for developmental stage specificity of the interactions between the core elements of HSs and the promoters of the globin genes.

Molecular evidence of tick-transmitted infections in dogs and cats in the United Kingdom.

PCR analysis was used to determine the prevalence of tick-transmitted infections in 120 systemically ill dogs and 60 cats recruited over a period of three months from 52 veterinary practices in the UK. The animals had not travelled outside the UK and had one or more of the following clinical criteria: acute or recurrent pyrexia, anaemia and/or thrombocytopenia, polyarthritis/muscle pain, splenomegaly/lymphadenopathy, and intraocular inflammation with systemic signs. Blood samples from the animals were tested for the presence of DNA from Borrelia burgdorferi sensu lato and Anaplasma phagocytophilum by using simple PCR targeting. B. burgdorferi sensu lato was detected in five dogs and two cats, and A. phagocytophilum was detected in one dog and one cat. These results provide the first molecular evidence of naturally occurring B. burgdorferi sensu lato infection in cats in the UK and confirm that A. phagocytophilum infection is present in cats. There were no statistically significant associations between the infections and the clinical signs shown by the dogs and cats.

Epidemiological survey of Angiostrongylus vasorum in dogs and slugs around a new endemic focus in Scotland.

The nematode parasite Angiostrongylus vasorum is an increasingly important cause of respiratory and other diseases in dogs. Geographical spread from previously limited endemic foci has occurred rapidly. This paper investigates parasite epidemiology around the location of the first reported case in Scotland in 2009: by detection of A vasorum-specific DNA in gastropod intermediate hosts, and in dogs circulating DNA and specific antibodies, and first stage larvae in faeces. Overall prevalence in gastropods was 6.7 per cent (16/240), with parasite DNA found in slugs in the Arion ater and Arion hortensis species aggregates and the snail Helix aspersa (syn. Cornu aspersum). Of 60 dogs presenting with clinical signs compatible with angiostrongylosis, none tested positive using PCR on peripheral blood or Baermann test on faeces, and none of 35 tested for circulating anti-A vasorum antibodies were positive. PCR prevalence in gastropods was highest (11 per cent) in the park frequented by the canine angiostrongylosis index case. Molecular survey for infection in gastropods is a potentially informative and efficient method for characterising the distribution of A vasorum and therefore local risk of canine infection. However, there appears to be a complex relationship between prevalence in gastropods and emergence of canine clinical disease, which requires further work to advance understanding of parasite transmission and geographical disease spread.

Tick-borne infectious diseases of dogs.

Tick-transmitted infections are an emerging problem in dogs. In addition to causing serious disease in traditional tropical and semi-tropical regions, they are now increasingly recognized as a cause of disease in dogs in temperate climates and urban environments. Furthermore, subclinically infected companion animals could provide a reservoir for human tick-transmitted infectious agents, such as Ehrlichia chaffeensis, Ehrlichia ewingll, the Ehrlichia phagocytophila group and Rickettsia conorii. Here, we discuss the emergence of new canine tick-transmitted diseases, which results from several factors, including the expansion of the tick range into urban and semi-urban areas worldwide, the movement of infected dogs into previously non-endemic areas, and the advent of novel molecular techniques for diagnosis and pathogen identification.

Pathogen carriage by the cat flea Ctenocephalides felis (Bouché) in the United Kingdom.

The carriage of Bartonella, Rickettsia felis and haemoplasma species was investigated in cat fleas (Ctenocephalides felis) collected from 121 cats and dogs in the United Kingdom. DNA extracted from fleas was analysed using genus and species-specific PCR and amplicons were characterised using DNA sequencing. Fifty percent of flea samples were PCR positive for at least one pathogen. Twenty one percent were positive for R. felis, 17% for Bartonella henselae, 40% for haemoplasma species and 20% were infected with more than one of the pathogen species studied. It is clear from the results in this study that companion cats and dogs are commonly infested with Ct. felis carrying bacterial pathogens of significance to human and animal health. These findings raise the possibility that Ct. felis found on dogs and cats are a potential source of infection with such pathogens for humans.

Immunologic study of systemic aspergillosis in German shepherd dogs.

Data are presented from a series of eight cases of disseminated canine aspergillosis (A. terreus) in German Shepherd dogs referred to Murdoch University Veterinary Hospital. Immunoglobulin determination revealed depression of serum IgA (cases 1 and 5) and IgM (case 2) levels and elevated levels of IgG in all cases. Total complement activity (CH50) and complement components tests, (C3, C4) were present in normal amounts in all cases. Using agar gel diffusion, serum antibody to A. terreus was found in only one case and aspergillus antigenaemia in two of the remainder. Lectin transformation of lymphocytes in two dogs was found to be depressed relative to normal controls in case 1 and initially in case 2. Two dogs failed to respond to the intradermal injection of A. terreus antigen.

Comparison of intradermal and serum testing for allergen-specific IgE using a Fcepsilon RIalpha-based assay in atopic dogs in the UK.

Atopic dermatitis in dogs is a common allergic skin disease that affects substantial numbers of dogs in the UK. The purpose of this study was to compare the results of an intradermal test (IDT) and an in vitro test in a large cohort of dogs. Dogs were intradermal tested with Greer allergens (Greer Labs Inc, Lenoir, NC, USA) using standard techniques. At the same time blood samples were drawn and submitted for evaluation by ELISA using the ALLERCEPT Definitive Allergen Panels for allergen-specific IgE, a commercial assay that uses a biotinylated recombinant extracellular domain of the high affinity Fc-epsilon receptor alpha chain protein (Fcepsilon RIalpha). The allergens used in the two tests included grass, tree and weed pollens, moulds, flea saliva/whole flea extract and house dust mite species. The optical density readings from the ELISA for each allergen were compared with the results of the IDT for 265 dogs. The prevalence of positive reactions in the ELISA was equal to or greater than the results of the IDT in the case of almost all of the allergens, but two notable exceptions were the house dust mites Dermatophagoides farinae and Dermatophagoides pteronyssinus. These two allergens were the most common positive reactions by IDT (prevalence D. farinae 78.9%, D. pteronyssinus 66.4%). The results of the two tests were significantly different (McNemar's test, P<0.05) for 16 of the 22 allergens. The sensitivities of the ELISA compared to the IDT (where there were more than 3 dogs with positive reactions in both tests) varied between 19.3 and 77.1% (D. pteronyssinus 19.3% and D. farinae 67.9%) and the specificities varied between 64.2 and 96.6% (D. pteronyssinus 96.6% and D. farinae 89.3%).

Subclass profile of allergen-specific IgG antibodies in atopic dogs.

Levels of allergen-specific IgE and IgE antibodies were determined in serum samples from 60 atopic and 11 normal dogs by means of commercially available ELISA test kits and a panel of 33 allergens. In the atopic population, IgE antibodies were most commonly identified with a specificity for Dermatophagoides farinae (78.3 per cent of affected dogs), D pteronyssinus (61.6 per cent), mould mix (25 per cent) and house dust (19 per cent), whereas the most frequently detected IgG antibodies had a specificity for D farinae (38.3 per cent), D pteronyssinus (33.3 per cent), mould mix (33.3 per cent), insect mix (16.6 per cent) and meadow fescue (16.6 per cent). The IgG subclass profile of allergen-specific antibodies was determined for five representative allergens from the panel. The IgG response to D farinae and D pteronyssinus was dominated by IgG4 antibodies, although lower levels of IgG2, and IgG3 and IgG1 D pteronyssinus antibodies were also detected. The IgG response to Timothy grass was predominantly within the IgG1 and IgG4 subclasses, IgG subclass selection in the response to mould mix and insect mix was broader, with relatively low level reactions from all four subclasses. The data suggest a degree of IgG subclass restriction in the humoral immune response of canine atopy which may be dependent upon the nature of the allergen.

Studies on the isolation and characterisation of a reaginic antibody in a cat.

A reaginic antibody has been demonstrated, by passive cutaneous anaphylaxis (PCA), in the serum of a cat infected with the microfilariae of Brugia pahangi. Recipient cats and pigs were challenged with an extract of Ascaris suum after either a four-hour or a 72-hour period of sensitisation. When the serum was heat treated at 56 degrees C it lost its PCA activity. Gel filtration of the serum revealed a pattern of positive PCA fractions similar to that observed in other species. Attempts to purify the PCA-positive material by Superose gel filtration and ion exchange chromatography by fast protein liquid chromatography (FPLC) were unsuccessful. Affinity chromatography of PCA-positive material by FPLC on protein A demonstrated two bound peaks, the second of which was PCA-positive and eluted as a single peak by ion exchange chromatography. The PCA-positive material from gel filtration did not bind to protein G. The protein A, PCA-positive peak provides a partially purified reaginic antibody for further study.

Demonstration of thiopurine methyltransferase activity in the erythrocytes of cats.

Azathioprine is a purine analogue used as an immunosuppressive and immunomodulator agent in various mammals, including cats. Several adverse reactions have been reported and have limited the use of the drug in the cat. Adverse reactions to azathioprine in humans have been correlated with reduced activity of thiopurine methyltransferase (TPMT) in erythrocytes. The purpose of this preliminary study was to determine if cats have TPMT activity in their erythrocytes and to compare the values obtained with the normal range for humans and the normal range for dogs in a preliminary report. Activity of the enzyme was measured in blood samples drawn from 41 cats. Blood also was taken from 5 dogs. The mean erythrocyte TPMT activity in the cats was 2.4 +/- 0.4 nmol (range, 1.2-3.9 nmol) per hour per milliliter of red blood cells (U/mL RBC) or 2-8 nmol per hour per gram of hemoglobin (U/g Hb). This range was far lower than the normal human range (8-15 U/mL RBC; 16-33 U/g Hb) and was of monopolar distribution. This observation apparently precludes any diagnostic purpose in assaying erythrocyte TPMT in this species. Erythrocyte TPMT activity in the 5 dogs ranged from 5.5 to 13.1 U/mL RBC (11-27 U/g Hb), which was comparable with normal and carrier ranges for humans, but proof of TPMT genetic polymorphism in either species will require genotyping studies.

A study of the efficacy of topical and systemic therapy for the treatment of feline Microsporum canis infection.

Microsporum canis infection was induced in 21 healthy SPF-derived cats. Once infection was established (4 weeks after inoculation) the cats were divided into three equal groups housed in separate rooms and monitored for 16 weeks. During this time, group A cats received oral griseofulvin at approximately 50 mg/kg daily and were shampooed twice weekly with a product containing chlorhexidine and miconazole. Group B cats were treated with griseofulvin alone, and group C cats served as untreated controls. The cats were examined on a weekly basis and the severity of lesions was scored semi-quantitatively. In addition, hair samples were collected from each cat on a weekly basis by the MacKenzie brush technique and by the sticky-tape method. A semi-quantitative scoring system was also used for the assessment of fungal (M canis) growth. Generally, significant differences in clinical scores were not seen between the groups although at weeks 3, 4 and 11 there was a significant difference (P< or =0.015) with cats in group A having significantly lower median scores than those in group C. Median times to clinical resolution (return of clinical scores to zero) in groups A, B and C were at treatment weeks 2, 9 and 12, respectively (P>0.05). Median times for mycological resolution (persistently negative culture results) for groups A, B and C were at treatment weeks 2, 9 and 12, respectively, for the MacKenzie brush technique and at weeks 4, 8 and 12 for the sticky-tape technique. For both these results, the groups differed significantly (P< or =0.001) and in both instances group A had significantly more rapid resolution than groups B or C. Median culture scores were significantly different between the three groups using one or both of the sampling techniques at week 2 through to week 12 of treatment with median scores for either group A alone, or groups A and B being significantly lower than group C (P< or =0.026). These results showed a benefit from the addition of twice-weekly chlorhexidine-miconazole shampooing to systemic griseofulvin therapy alone in the treatment of M canis infected cats.

Isolation and functional analysis of normal canine blood monocytes and resident alveolar macrophages.

The percentage of mononuclear phagocytes bearing the Fc receptor for immunoglobulin G, the percentage of cells phagocytic for Candida albicans and latex particles, and the phagocytic index for blood monocytes and alveolar macrophages from healthy dogs are reported. Blood monocytes were concentrated by density-gradient centrifugation, whereas alveolar macrophages were obtained in high yield by bronchoalveolar lavage. Adherent populations of those cells were used for functional assays after repeated washing to remove nonadherent cells. A greater percentage of adherent alveolar macrophages than adherent blood monocytes showed evidence of the Fc receptor for immunoglobulin G. Similarly, adherent alveolar macrophages showed significantly greater phagocytic ability, as measured by percent phagocytic cells and phagocytic index, using C albicans and latex particles, than did adherent canine blood monocytes.

Epidemiological and diagnostic features of canine and feline dermatophytosis in the United Kingdom from 1956 to 1991.

Between 1956 and 1991, 8349 samples from dogs and cats were received for investigation of suspected dermatophytosis, and 1368 (16 per cent) yielded positive cultures. Cats had a significantly higher proportion of positive cultures (26 per cent) than dogs (10 per cent), and of these Microsporum canis accounted for 92 per cent in cats and 65 per cent in dogs. The other isolates were diverse but mainly sylvatic dermatophytes, and M gypseum was isolated on only four occasions. Different breeds of dog and cat had significantly different prevalences of infection, with pedigree and long-haired cats, and Jack Russell and Yorkshire terrier dogs having a particularly high proportion of positive cultures. Animals less than one year old appeared to be predisposed to infection, but there was no apparent sex predisposition and no conclusive evidence of any seasonal variation in the incidence of the disease. In comparison with the results of dermatophyte culture, examination under Wood's lamp had a positive predictive value of 90 per cent and a negative predictive value of 94 per cent in determining M canis infection, and direct microscopy had positive and negative predictive values of 93 per cent in determining the presence of dermatophytosis. However, cultural examination alone was insufficient for the diagnosis of dermatophytosis owing to the occurrence of false positive and false negative results.