Cytological evaluation, culture and genomics to evaluate the microbiome in healthy rabbit external ear canals.

BACKGROUND: Lop-eared rabbits may be predisposed to otitis externa (OE) as a consequence of their ear conformation. Although otoscopy, otic cytological evaluation and culture are valuable tools in dogs and cats, published data on rabbits remain lacking. HYPOTHESIS/OBJECTIVES: This study aimed to assess the utility of otoscopy and cytological results in evaluating healthy rabbit external ear canals (EECs) and to characterise ear cytological and microbiological findings through culture techniques and metagenomic sequencing. ANIMALS: Sixty-three otitis-free client-owned rabbits. MATERIALS AND METHODS: All rabbits underwent otoscopy and ear cytological evaluation. In a subset of 12 rabbits, further bacterial and fungal culture, fungal DNA assessment and metagenomic sequencing were performed. RESULTS: Otic cytological results revealed yeast in 73%, cocci in 42.9% and rods in 28.6% of healthy rabbit EECs. Compared to upright-eared rabbits, lop-eared rabbits had more discharge and more bacteria per oil immersion field. Culture isolated eight different species yet metagenomic sequencing identified 36, belonging to the Bacillota (Firmicutes), Pseudomonadota and Actinomycetota phyla. Staphylococcus were the most commonly observed species with both methods. Ten of 12 rabbits were yeast-positive on cytological evaluation with only three yielding fungal growth identified as Yarrowia (Candida) lipolytica, Eurotium echinulatum and Cystofilobasidium infirmominiatum. CONCLUSIONS AND CLINICAL RELEVANCE: Healthy rabbit EECs lack inflammatory cells yet can host yeast and bacteria, emphasising the need to evaluate cytological results alongside the clinical signs. Lop-ear anatomy may predispose to bacterial overgrowth and OE. Notably, yeasts may be present despite a negative culture.

Latent gammaherpesvirus infection enhances type I IFN response and reduces virus spread in an influenza A virus co-infection model.

Infections with persistent or latent viruses alter host immune homeostasis and have potential to affect the outcome of concomitant acute viral infections such as influenza A virus (IAV). Gammaherpesviruses establish life-long infections and require an on-going immune response to control reactivation. We have used a murine model of co-infection to investigate the response to IAV infection in mice latently infected with the gammaherpesvirus MHV-68. Over the course of infection, latently infected BALB/c mice showed less weight loss, clinical signs, pulmonary cellular infiltration and expression of inflammatory mediators than naïve mice infected with IAV and had significantly more activated CD8+ T cells in the lungs. Four days after IAV infection, virus spread in the lungs of latently infected animals was significantly lower than in naïve animals. By 7 days after IAV infection latently infected lungs express elevated levels of cytokines and chemokines indicating they are primed to respond to the secondary infection. Investigation at an early time point showed that 24 h after IAV infection co-infected animals had higher expression of IFNß and Ddx58 (RIG-I) and a range of ISGs than mice infected with IAV alone suggesting that the type I IFN response plays a role in the protective effect. This effect was mouse strain dependent and did not occur in 129/Sv/Ev mice. These results offer insight into innate immune mechanisms that could be utilized to protect against IAV infection and highlight on-going and persistent viral infections as a significant factor impacting the severity of acute respiratory infections.

Animal welfare requirements in publishing guidelines.

Descriptions of measures taken to optimize animal welfare are often absent from scientific reports of animal experiments. One reason may be that journal guidelines inadequately compel authors to provide such information. In this study, online English language versions of the 'Guidelines to authors' (GTAs) from 54 national biomedical journals were examined for neutral (unrelated to welfare) and non-neutral keywords referring to: animal welfare; the '3Rs'; the ARRIVE (2010) guidelines, and regulations pertaining to animal experimentation. Journals were selected from nine countries (UK, US, China, Canada, India, Brazil, Germany, Japan and Australia) and seven biomedical specialties (oncology, rheumatology, surgery, pharmacology, medicine, anaesthesia and veterinary medicine). Total GTA word counts varied from 1137 to 31,609. The keyword count identified per category were expressed per myriad (10,000) of total word count. One-way analyses of variance followed by post hoc Tukey pairwise comparisons revealed greater non-neutral per myriad word counts for (a) veterinary GTAs compared with medicine, oncology, rheumatology or surgery; (b) British, compared with Australian, Canadian, German and Japanese GTAs; and (c) no differences between non-neutral categories. The English language versions of GTAs of British and veterinary medical journals contain more words associated with animal welfare, the 3Rs and the ARRIVE guidelines than those from eight other countries and six other medical specialities. The exclusion of 'national' language versions from analysis precludes attempts to identify national differences in attitudes to laboratory animal welfare.

MRI of the optic nerve sheath and globe in cats with and without presumed intracranial hypertension.

OBJECTIVES: The study aimed to: (1) test MRI repeatability of measurements of optic nerve sheath diameter (ONSD), optic nerve diameter (OND) and eye globe transverse diameter (ETD); (2) investigate the associations between the OND, ONSD and ETD; (3) assess whether these measurements are affected by age or body weight; and (4) test the association between ONSD, OND, ETD and ONSD:ETD ratio with presumed intracranial pressure (ICP) status. METHODS: This was a retrospective and blinded study where patients were allocated to presumed normal or intracranial hypertension groups based on MRI findings. The ONSD and ETD were measured and recorded. Interclass correlation coefficient (ICC) was calculated to investigate interobserver agreement. Data were analysed using the Pearson correlation coefficient, two-sample t-test and general linear model ANOVA. RESULTS: Seventy-seven cats were included, 62 with presumed normal ICP and 15 with presumed intracranial hypertension. The ICC showed moderate-to-good reliability for all measurements. Positive correlations were identified for: (1) ETD and weight; (2) ONSD and age; (3) OND and age; (4) ONSD and ETD; (5) ONSD:ETD ratio and presumed ICP status; and (6) ONSD and presumed ICP status. No difference was detected between the presumed normal and intracranial hypertension groups and ONSD, as well as ONSD:ETD ratio and presumed ICP status when patient age was considered. CONCLUSIONS AND RELEVANCE: The measurement of the ONSD and the ONSD:ETD ratio on T2-weighted MRI might not be reliable as non-invasive tests for diagnosing intracranial hypertension in cats.

Lack of IFNγ signaling attenuates spread of influenza A virus in vivo and leads to reduced pathogenesis.

IFNγ is a key regulator of inflammatory responses but its role in influenza A virus (IAV) pathogenesis is unclear. Our studies show that infection of mice lacking the IFNγ receptor (IFNγR-/-) at a dose which caused severe disease in wild type 129 Sv/Ev (WT) mice resulted in milder clinical symptoms and significantly lower lung virus titers by 6 days post-infection (dpi). Viral spread was reduced in IFNγR-/- lungs at 2 and 4 dpi. Levels of inflammatory cytokines and chemokines were lower in IFNγR-/- mice at 2 dpi and there was less infiltration of monocyte/macrophage lineage cells than in WT mice. There was no difference in CD4+ and CD8+ T cells and alveolar macrophages in the bronchoalveolar lavage fluid (BALF) at 2 and 4 dpi but by 4 dpi IFNγR-/- mice had significantly higher percentages of neutrophils. Our data strongly suggest that IAV can use the inflammatory response to promote viral spread.

Thermography as an early predictive measurement for evaluating epidural and femoral-sciatic block success in dogs.

OBJECTIVE: To evaluate skin temperature increase as an early predictive measure for evaluating epidural and femoral-sciatic block success in dogs. STUDY DESIGN: Prospective clinical trial. ANIMALS: A total of 29 dogs undergoing orthopaedic surgery on one hindlimb. METHODS: Dogs were anaesthetized and placed into lateral recumbency with the affected limb uppermost and the coat was clipped. Baseline infrared thermographic images (T0) of the affected limb, of the paw pad of the affected leg and of the ipsilateral paw pad were taken. Subsequently, dogs were administered either an epidural (EPI; n=11) or a femoral-sciatic block (FS; n=18) using bupivacaine 1 mg kg-1. Then, 2 minutes after placement of the block, thermographic images were obtained every 3 minutes for a total of four measurements (T1-T4) and surgery was commenced. Rescue analgesia consisting of fentanyl 1 µg kg-1 was administered if needed. A regional block was considered successful if the dose of fentanyl administered was less than the lower 95% confidence interval of the geometric mean of the total fentanyl used in each group. A ≥ 1 °C increase of skin temperature was considered as the minimum increase required for detection of a successful block. RESULTS: A total of 12 out of 18 blocks in the FS and eight of 11 in the EPI group were considered successful based on fentanyl consumption. Out of these, only four of 12 in the FS and one of eight in the EPI group developed an increase in temperature of ≥ 1 °C. Contrarily, four of six of the nonsuccessful cases in the FS and three of three in the EPI group developed an increase in temperature of ≥ 1 °C. CONCLUSIONS AND CLINICAL RELEVANCE: Contrary to reports in humans, thermography did not indicate regional block success prior to surgery in dogs. However further studies under more controlled conditions are needed to determine whether thermography can be used to indicate failure of regional blockade.

Pathological and phylogenetic characterization of Amphibiothecum sp. infection in an isolated amphibian (Lissotriton helveticus) population on the island of Rum (Scotland).

Outbreaks of cutaneous infectious disease in amphibians are increasingly being attributed to an overlooked group of fungal-like pathogens, the Dermocystids. During the last 10 years on the Isle of Rum, Scotland, palmate newts (Lissotriton helveticus) have been reportedly afflicted by unusual skin lesions. Here we present pathological and molecular findings confirming that the pathogen associated with these lesions is a novel organism of the order Dermocystida, and represents the first formally reported, and potentially lethal, case of amphibian Dermocystid infection in the UK. Whilst the gross pathology and the parasite cyst morphology were synonymous to those described in a study from infected L. helveticus in France, we observed a more extreme clinical outcome on Rum involving severe subcutaneous oedema. Phylogenetic topologies supported synonymy between Dermocystid sequences from Rum and France and as well as their distinction from Amphibiocystidium spp. Phylogenetic analysis also suggested that the amphibian-infecting Dermocystids are not monophyletic. We conclude that the L. helveticus-infecting pathogen represents a single, novel species; Amphibiothecum meredithae.

Susceptibility of bone marrow-derived macrophages to influenza virus infection is dependent on macrophage phenotype.

The role of the macrophage in influenza virus infection is complex. Macrophages are critical for resolution of influenza virus infections but implicated in morbidity and mortality in severe infections. They can be infected with influenza virus and consequently macrophage infection is likely to have an impact on the host immune response. Macrophages display a range of functional phenotypes, from the prototypical pro-inflammatory classically activated cell to alternatively activated anti-inflammatory macrophages involved in immune regulation and wound healing. We were interested in how macrophages of different phenotype respond to influenza virus infection and therefore studied the infection of bone marrow-derived macrophages (BMDMs) of classical and alternative phenotype in vitro. Our results show that alternatively activated macrophages are more readily infected and killed by the virus than classically activated. Classically activated BMDMs express the pro-inflammatory markers inducible nitric oxide synthase (iNOS) and TNF-α, and TNF-α expression was further upregulated following infection. Alternatively activated macrophages express Arginase-1 and CD206; however, following infection, expression of these markers was downregulated whilst expression of iNOS and TNF-α was upregulated. Thus, infection can override the anti-inflammatory state of alternatively activated macrophages. Importantly, however, this results in lower levels of pro-inflammatory markers than those produced by classically activated cells. Our results showed that macrophage phenotype affects the inflammatory macrophage response following infection, and indicated that modulating the macrophage phenotype may provide a route to develop novel strategies to prevent and treat influenza virus infection.

Sheep lung segmental delivery strategy demonstrates adenovirus priming of local lung responses to bacterial LPS and the role of elafin as a response modulator.

Viral lung infections increase susceptibility to subsequent bacterial infection. We questioned whether local lung administration of recombinant adenoviral vectors in the sheep would alter the susceptibility of the lung to subsequent challenge with bacterial lipopolysaccharide (LPS). We further questioned whether local lung expression of elafin, a locally produced alarm anti-LPS/anti-bacterial molecule, would modulate the challenge response. We established that adenoviral vector treatment primed the lung for an enhanced response to bacterial LPS. Whereas this local effect appeared to be independent of the transgene used (Ad-o-elafin or Ad-GFP), Ad-o-elafin treated sheep demonstrated a more profound lymphopenia in response to local lung administration of LPS. The local influence of elafin in modulating the response to LPS was restricted to maintaining neutrophil myeloperoxidase activity, and levels of alveolar macrophage and neutrophil phagocytosis at higher levels post-LPS. Adenoviral vector-bacterial synergism exists in the ovine lung and elafin expression modulates such synergism both locally and systemically.

Ovine herpesvirus-2-encoded microRNAs target virus genes involved in virus latency.

Herpesviruses encode microRNAs (miRNAs) that target both virus and host genes; however, their role in herpesvirus biology is understood poorly. We identified previously eight miRNAs encoded by ovine herpesvirus-2 (OvHV-2), the causative agent of malignant catarrhal fever (MCF), and have now investigated the role of these miRNAs in regulating expression of OvHV-2 genes that play important roles in virus biology. ORF20 (cell cycle inhibition), ORF50 (reactivation) and ORF73 (latency maintenance) each contain predicted targets for several OvHV-2 miRNAs. Co-transfection of miRNA mimics with luciferase reporter constructs containing the predicted targets showed the 5' UTRs of ORF20 and ORF73 contain functional targets for ovhv-miR-2 and ovhv2-miR-8, respectively, and the 3' UTR of ORF50 contains a functional target for ovhv2-miR-5. Transfection of BJ1035 cells (an OvHV-2-infected bovine T-cell line) with the relevant miRNA mimic resulted in a significant decrease in ORF50 and a smaller but non-significant decrease in ORF20. However, we were unable to demonstrate a decrease in ORF73. MCF is a disease of dysregulated lymphocyte proliferation; miRNA inhibition of ORF20 expression may play a role in this aberrant lymphocyte proliferation. The proteins encoded by ORF50 and ORF73 play opposing roles in latency. It has been hypothesized that miRNA-induced inhibition of virus genes acts to ensure that fluctuations in virus mRNA levels do not result in reactivation under conditions that are unfavourable for viral replication and our data supported this hypothesis.

The safety profile of a cationic lipid-mediated cystic fibrosis gene transfer agent following repeated monthly aerosol administration to sheep.

Clinically effective gene therapy for Cystic Fibrosis has been a goal for over 20 years. A plasmid vector (pGM169) that generates persistent expression and reduced host inflammatory responses in mice has raised prospects for translation to the clinic. The UK CF Gene Therapy Consortium is currently evaluating long-term repeated delivery of pGM169 complexed with the cationic lipid GL67A in a large Multidose Trial. This regulatory-compliant evaluation of aerosol administration of nine doses of pGM169/GL67A at monthly intervals, to the sheep lung, was performed in preparation for the Multidose Trial. All sheep tolerated treatment well with no adverse effects on haematology, serum chemistry, lung function or histopathology. Acute responses were observed in relation to bronchoalveolar cellularity comprising increased neutrophils and macrophage numbers 1 day post-delivery but these increases were transient and returned to baseline. Importantly there was no cumulative inflammatory effect or lung remodelling with successive doses. Molecular analysis confirmed delivery of pGM169 DNA to the airways and pGM169-specific mRNA was detected in bronchial brushing samples at day 1 following doses 1, 5 and 9. In conclusion, nine doses of pGM169/GL67A were well tolerated with no significant evidence of toxicity that would preclude adoption of a similar strategy in CF patients.

Salmonella transforms follicle-associated epithelial cells into M cells to promote intestinal invasion.

Salmonella Typhimurium specifically targets antigen-sampling microfold (M) cells to translocate across the gut epithelium. Although M cells represent a small proportion of the specialized follicular-associated epithelium (FAE) overlying mucosa-associated lymphoid tissues, their density increases during Salmonella infection, but the underlying molecular mechanism remains unclear. Using in vitro and in vivo infection models, we demonstrate that the S. Typhimurium type III effector protein SopB induces an epithelial-mesenchymal transition (EMT) of FAE enterocytes into M cells. This cellular transdifferentiation is a result of SopB-dependent activation of Wnt/ß-catenin signaling leading to induction of both receptor activator of NF-κB ligand (RANKL) and its receptor RANK. The autocrine activation of RelB-expressing FAE enterocytes by RANKL/RANK induces the EMT-regulating transcription factor Slug that marks epithelial transdifferentiation into M cells. Thus, via the activity of a single secreted effector, S. Typhimurium transforms primed epithelial cells into M cells to promote host colonization and invasion.

Mycobacterial disease in cats in Great Britain: I. Culture results, geographical distribution and clinical presentation of 339 cases.

This study investigated 339 cases of feline mycobacterial disease from cats with cutaneous lesions or masses found at exploratory laparotomy. Tissue samples were submitted to the Veterinary Laboratories Agency for mycobacterial culture over a 4-year period to December 2008. The study assessed which species of culturable mycobacteria were involved, where the cats lived, and their clinical presentation (physical findings, serum biochemistry, radiography, feline leukaemia virus and feline immunodeficiency virus status). Mycobacterium microti was cultured from 19%, Mycobacterium bovis 15%, Mycobacterium avium 7%, non-M avium non-tuberculous mycobacteria 6%, with no growth in 53% of samples. M microti, M bovis and M avium were found in almost mutually exclusive clusters within Great Britain (GB) (ie, M bovis in South-West England/Wales/Welsh Border, M avium in eastern England and M microti south of London and in South-West Scotland). While differences were seen in the clinical presentation and distribution of lesions caused by the different infections, these were not sufficiently different to be diagnostic. Cats commonly presented with single or multiple cutaneous lesions (74%), which were sometimes ulcerated or discharging, located most frequently on the head (54%). Lymph nodes were usually involved (47%); typically the submandibular nodes. Systemic or pulmonary signs were rarely seen (10-16%). When a cat is suspected of having mycobacteriosis, accurate identification of the species involved helps to determine appropriate action. Our findings show that knowing the cat's geographic location can be helpful, while the nature of the clinical presentation is less useful. Most cases of feline mycobacterial disease in GB are cutaneous.

Mycobacterial disease in a population of 339 cats in Great Britain: II. Histopathology of 225 cases, and treatment and outcome of 184 cases.

This study investigated 339 cases of feline mycobacterial infection, with histopathology findings from 225 cases, and treatment and outcome information from 184 cases. Tissue samples from cats with cutaneous lesions or suspicious masses at exploratory laparotomy were submitted to the Veterinary Laboratories Agency for mycobacterial culture over a 4-year period to December 2008. The study reviewed the files for information about histopathology, treatment and outcome, and blindly reviewed histopathological changes (including staining for acid-fast bacteria [AFB]) in a sub-set of 45 cases. When a cat is suspected of having a mycobacterial infection, accurate identification of the species involved helps to determine possible treatment options and prognosis. The study confirmed that histopathology and the presence of AFB are useful tools in the recognition of mycobacterial infection. Unfortunately, they did little to help determine the species of mycobacteria involved. The study identified a group of cats that were negative for AFB at the primary laboratory, but from which mycobacteria could be cultured; commonly Mycobacterium bovis or Mycobacterium microti. The study also identified a group of cats which where culture negative, despite typical signs of mycobacterial infection and positive AFB staining. Many cases responded favourably to treatment (56% of the cases where information was available), and many cats gained complete remission (42%). However, relapses were common (64%) and often followed by pulmonary and/or systemic spread that may have resulted from treatment with short courses of single drugs. This study shows that the diagnosis and treatment of feline mycobacteriosis is complex and challenging.

Comparative analysis of EspF variants in inhibition of Escherichia coli phagocytosis by macrophages and inhibition of E. coli translocation through human- and bovine-derived M cells.

The EspF protein is secreted by the type III secretion system of enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC, respectively). EspF sequences differ between EHEC O157:H7, EHEC O26:H11, and EPEC O127:H6 in terms of the number of SH3-binding polyproline-rich repeats and specific residues in these regions, as well as residues in the amino domain involved in cellular localization. EspF(O127) is important for the inhibition of phagocytosis by EPEC and also limits EPEC translocation through antigen-sampling cells (M cells). EspF(O127) has been shown to have effects on cellular organelle function and interacts with several host proteins, including N-WASP and sorting nexin 9 (SNX9). In this study, we compared the capacities of different espF alleles to inhibit (i) bacterial phagocytosis by macrophages, (ii) translocation through an M-cell coculture system, and (iii) uptake by and translocation through cultured bovine epithelial cells. The espF gene from E. coli serotype O157 (espF(O157)) allele was significantly less effective at inhibiting phagocytosis and also had reduced capacity to inhibit E. coli translocation through a human-derived in vitro M-cell coculture system in comparison to espF(O127) and espF(O26). In contrast, espF(O157) was the most effective allele at restricting bacterial uptake into and translocation through primary epithelial cells cultured from the bovine terminal rectum, the predominant colonization site of EHEC O157 in cattle and a site containing M-like cells. Although LUMIER binding assays demonstrated differences in the interactions of the EspF variants with SNX9 and N-WASP, we propose that other, as-yet-uncharacterized interactions contribute to the host-based variation in EspF activity demonstrated here.

Evaluation of formalin-fixed ileum as the optimum method to diagnose equine dysautonomia (grass sickness) in simulated intestinal biopsies.

Equine dysautonomia, or grass sickness, is a frequently fatal disease of unknown etiology, manifested as poor gastrointestinal motility and colic as a result of degenerative changes in the autonomic nervous system. Examination of ileal biopsies collected at laparotomy is currently the best antemortem diagnostic method to distinguish equine dysautonomia from colic cases, which can present with similar signs, but their value has not been previously critically evaluated. Using simulated biopsies collected postmortem from 23 cases of equine dysautonomia and 11 of colic, the sensitivity and specificity of 1-cm long, formalin-fixed ileal biopsies was 100% for the diagnosis of equine dysautonomia. There was therefore no advantage to using larger biopsies or examining jejunum either in addition to or instead of ileal biopsies. Furthermore, although cryostat sections of ileum, 1-cm long, had a sensitivity of 100%, the specificity was only 73%, meaning that 27% of cases would have been misclassified, resulting in unnecessary euthanasia. Increasing the size of the cryostat or examining jejunum in addition to ileum cryostat sections did not significantly improve the specificity. Results of the current study indicate that in diagnostic practice, 1-cm long, formalin-fixed biopsies are likely to be the most suitable for accurate diagnosis, despite the slower turnaround time compared with cryostat sections.

Pulsed-wave Doppler tissue imaging velocities in normal geriatric cats and geriatric cats with primary or systemic diseases linked to specific cardiomyopathies in humans, and the influence of age and heart rate upon these velocities.

Pulsed-wave Doppler tissue imaging (pw-DTI) techniques allow the non-invasive assessment of myocardial dynamics. pw-DTI has demonstrated regional and global diastolic impairment in various forms of human and feline cardiomyopathy. We hypothesise that in geriatric cats with systemic diseases that have been linked to specific cardiomyopathies in human beings, the myocardial velocity profile will be altered when compared to either normal or hypertrophic cardiomyopathy (HCM) cats; and that both age and heart rate have a significant affect upon pw-DTI velocities. The aims of this study were to determine whether the feline M-mode or myocardial velocity profile is altered in geriatric cats with disease states that have been linked to specific cardiomyopathies in humans when compared to normal geriatric cats or geriatric cats with HCM and to determine whether age or heart rate has a significant effect upon pw-DTI velocities within these groups of cats. Sixty-six cats aged 8 years or above were included in the study, and were divided as follows: Unaffected (n=8), basilar septal bulge (BSB) (17), HCM (14), hyperthyroid (HiT(4)) (12) and chronic renal failure (CRF) (15). Systolic blood pressure was normal in all the cats. pw-DTI systolic (S'), early (E') and late diastolic (A') velocities were assessed from standardised sites within the myocardium, and the relationships between these and disease group, age and heart rate were then assessed. In cats with HCM, the E' velocity was decreased at various sites. Conversely, the HiT(4) cats demonstrated increased S' velocities. The only site at which the age of the cat was significantly related to myocardial velocities was the S' velocity from the apical mid-septum. There were also significant positive relationships between heart rate and the magnitude of myocardial S', E' and A' velocities of radial motion and S' and A' velocities of longitudinal motion. pw-DTI detected diastolic dysfunction in untreated cats with HCM and increased systolic function in HiT(4) cats. The age of the cat was of little significance, whereas heart rate significantly influenced myocardial velocity profiles.

Feline diabetes mellitus in the UK: the prevalence within an insured cat population and a questionnaire-based putative risk factor analysis.

Prevalence and risk factors for the development of diabetes mellitus (DM) in cats in the United Kingdom have not previously been reported. The prevalence of DM was evaluated in a large insured population and was found to be 1 in 230 cats. In this insured cat population Burmese cats were 3.7 times more likely to develop DM than non-pedigree cats. A convenience-sampling questionnaire-based study was used in order to identify putative risk factors for the development of DM. The univariate risk factor analysis identified being male, neutered, inactive, weighing >or=5 kg and having a history of corticosteroid treatment as significant risk factors for the development of DM in these cats. In addition, male cats treated with megestrol acetate had a significantly increased risk of developing DM compared to females. In contrast, there was no difference in DM occurrence between male and female Burmese cats. A multivariate classification tree-based model on the questionnaire data looking for interactions between risk factors, identified gender as the most important overall risk factor for the development of DM with low physical activity being the next most important risk factor for female cats and breed the next most important for male cats.

Importance of using guarded techniques for the preparation of endometrial cytology smears in mares.

Material for endometrial cytology can be collected by veterinarians using guarded or unguarded swabs, or digitally with a gloved hand, and is an important diagnostic tool in establishing the endometrial health of mares prior to breeding. The aim of this study was to determine whether the use of unguarded endometrial samples is a reliable indicator of the presence of neutrophils in the uterus. Duplicate endometrial smears were collected from 41 genitally normal, non-pregnant fertile mares by both double-guarded swabs (DGS) and in an unguarded manner by digital scraping (DS) of the endometrium. In 17 of the 41 mares, smears were also collected from the cranial vagina by DS. Cytological samples were collected from a further seven non-pregnant mares at different reproductive stages, and tissues (vestibule, vagina and cervix) from four reproductively normal mares were examined histologically after slaughter to detect the presence of neutrophils. Only 3/41 (7.3%) of the DGS endometrial smears had neutrophils present compared to 36/41 (87.8%) of the DS endometrial smears. The percentage of neutrophils in DGS endometrial smears ranged from 0 to 6% (mean = 0.41%), whereas those in the DS smears ranged from 2 to 90% (mean = 22.02%). Neutrophils were present in all vaginal smears (17/17, range=3-56% (mean = 22.18%)). There was a significant positive correlation (r = 0.84; P < 0.001) between the percentage of neutrophils in the vagina and in the DS endometrial smears. More neutrophils were found in the cervix, vagina and vestibule than in endometrial smears during the cycle (P<0.05). Neutrophils were also observed in tissue collected from the cervix, vagina and vestibule from reproductively normal mares at post-mortem. In conclusion, endometrial smears collected using unguarded techniques are very likely to be contaminated with neutrophils transferred from the vagina potentially leading to incorrect diagnosis of endometritis. When collecting samples for endometrial cytology it is important to use guarded techniques to ensure that only the endometrium is sampled to avoid contamination with cells carried over from other areas of the reproductive tract.

Efficacy of praziquantel against Schistosoma haematobium infection in children.

A study was performed to determine the efficacy of praziquantel (PZQ) against Schistosoma haematobium. Children (n = 592) infected with S. haematobium received either a single treatment with PZQ (40 mg/kg) or two or three treatments with PZQ at three-week intervals after the initial treatment and efficacy was monitored for nine weeks. Cure rates at three-weeks post-treatment were low (< 50%), suggesting either that worms are killed very slowly or, more likely, that eggs continue to be released from tissues after worm death. Interestingly, a single dose of PZQ showed high efficacy (cure rate > 83% and egg reduction rate > 98%) when assessed from six weeks post-treatment onward. There were no significant differences in cure rates or intensity of infection between the three cohorts at any point in the study, despite the different treatment regimens. Since children were in contact with transmission sites during the study period, the results suggest good efficacy of PZQ against all stages of S. haematobium, including the immature worms.