Evaluation of a Pseudotyped Virus Neutralisation Test for the Measurement of Equine Influenza Virus-Neutralising Antibody Responses Induced by Vaccination and Infection.

Equine influenza is a major respiratory disease of horses that is largely controlled by vaccination in some equine populations. Virus-neutralising antibodies, the mainstay of the protective immune response, are problematic in assaying for equine influenza virus, as most strains do not replicate efficiently in cell culture. Surrogate measures of protective antibody responses include the haemagglutination inhibition (HI) test and single radial haemolysis (SRH) assay. For this study, a pseudotyped virus, bearing an envelope containing the haemagglutinin (HA) from the Florida clade 2 equine influenza virus strain A/equine/Richmond/1/07 (H3N8), was generated to measure HA-specific neutralising antibodies in serum samples (n = 134) from vaccinated or experimentally-infected ponies using a pseudotyped virus neutralization test (PVNT). Overall, the results of PVNT were in good agreement with results from the SRH assay (100% sensitivity, 68.53% specificity) and HI test (99.2% sensitivity, 49.03% specificity). The PVNT was apparently more sensitive than either the SRH assay or the HI test, which could be advantageous for studying the antibody kinetics, particularly when antibody levels are low. Nevertheless, further studies are required to determine whether a protective antibody level can be defined for the SRH assay and to ascertain the inter-laboratory reproducibility. In conclusion, the PVNT efficiently measures neutralising antibodies after immunization and/or experimental infection in the natural host, and may complement existing antibody assays.

Refinement of the equine influenza model in the natural host: A meta-analysis to determine the benefits of individual nebulisation for experimental infection and vaccine evaluation in the face of decreased strain pathogenicity.

Equine Influenza (EI) is an important respiratory disease of horses caused by H3N8 equine influenza viruses (EIV). Vaccination is a key strategy to prevent or control this disease. However, EIV undergoes continuous antigenic drift and whilst numerous EI vaccines are commercially available worldwide, an accurate evaluation of their efficacy is frequently required through clinical trials conducted in the natural host. Room nebulisation is one of the chosen methods to challenge horses during EI vaccine studies. A potential decreased pathogenicity observed with recent Florida Clade 2 (FC2) EIV isolates have increased the heterogeneity of the clinical response and virus shedding measured after infection by room nebulisation, which reduced the statistical power of studies. Our objectives were to compare clinical and virological parameters following experimental infection with several different EIV strains and to confirm that individual nebulisation is a model refinement that prevents an increase of the number of animals per group. This study is a retrospective comparison and meta-analysis of clinical and virological results collected from 9 independent EIV infection studies in the natural host. Naïve Welsh mountain ponies were experimentally infected by room or individual nebulisation with FC2 EIV strains, including A/equine/Richmond/1/07 (R/07), A/equine/East Renfrewshire/11 (ER/11), A/equine/Cambremer/1/2012 (C/12) and A/equine/Northamptonshire/1/13 (N/1/13). The retrospective meta-analysis confirmed a decreased pathogenicity of the EIV ER/11 and C/12 strains when compared with R/07. Experimental infection by individual nebulisation improved the clinical and virological parameters induced by recent FC2 strains, when compared with conventional room nebulisation. In conclusion, individual nebulisation offers a better control of the challenge dose administered and a greater homogeneity of the response measured in control animals. This in turn, helps maintain the number of animals per group to the minimum necessary required to obtain meaningful results in vaccine efficacy studies, which adheres to the 3Rs (Replacement, Reduction and Refinement) principles.

In vitro antimicrobial activity of miconazole and polymyxin B against canine meticillin-resistant Staphylococcus aureus and meticillin-resistant Staphylococcus pseudintermedius isolates.

BACKGROUND: Meticillin-resistant Staphylococcus aureus (MRSA) and meticillin-resistant Staphylococcus pseudintermedius (MRSP) infections are increasingly reported in dogs, and these bacteria may be isolated from ear infections. HYPOTHESIS/OBJECTIVES: The main aim of the present study was to investigate the in vitro antimicrobial activity of miconazole, polymyxin B and a combination of both against 24 canine MRSA and 50 canine MRSP isolates. The minimal inhibitory concentration (MIC) values of 12 other antimicrobial agents were also determined. METHODS: All MIC values were determined according to a broth microdilution assay. RESULTS: Acquired resistance was found to all tested agents, except for linezolid, miconazole and polymyxin B. The MIC values for miconazole and polymyxin B against MRSA were in the range of 4-8 and 8-64 µg/mL, respectively, while the MIC values for miconazole and polymyxin B against MRSP were in the range of 1-2 and 0.25-4 µg/mL, respectively. Using a combination of miconazole and polymyxin B, there was no evidence for enhanced in vitro activity of the combination (i.e. synergy) of both products. Nevertheless, MIC(90) values of the combination of these antimicrobial agents and of a commercial product containing both agents were at least 1000 times lower than the concentration present in the commercial product. CONCLUSIONS AND CLINICAL IMPORTANCE: These results indicate that the topical use of a combination of miconazole and polymyxin B in a 43.5:1 ratio may have potential for the treatment of MRSA-mediated and MRSP-associated otitis externa in dogs.

Effect of mitratapide on body composition, body measurements and glucose tolerance in obese Beagles.

The objective of this study was to confirm that weight loss after treatment with mitratapide (Yarvitan®) is loss of adipose tissue. Obese dogs were treated with the recommended treatment schedule of mitratapide. Dual-energy X-ray absorptiometry (DEXA) was done before and after the treatment schedule. Body weight, feed consumption and pelvic circumference were recorded and a glucose tolerance test was performed. Dualenergy X-ray absorptiometry measurements showed an impressive loss of fat tissue, corresponding to a mean loss of approximately 41.6% of the body fat mass recorded before treatment. After treatment with mitratapide, the mean body fat percentage had returned within the normal range. At the end of the study, the dogs had lost on average 14.2% of their body weight and 15.2% of their pelvic circumference compared to baseline. The results also suggest that losing weight with mitratapide might help to reverse insulin resistance.

Peptic ulcer disease associated with Helicobacter felis in a dog owner.

The aim of this study was to investigate the identity of the Helicobacter heilmannii-like bacteria found in the stomach of a human patient suffering from stomach ulcers and her asymptomatic pet dog. An elderly woman was referred for gastroscopy because of right hypochondrial pain, nausea, anorexia and vomiting. Gastric ulcers were observed and histology revealed the presence of multiple H. heilmannii-like bacteria. Multiplex polymerase chain reaction (PCR) identified the bacteria as H. felis. Her pet dog was also examined gastroscopically. Only mild gastric lesions were found but PCR showed the presence of H. felis as well as H. bizzozeronii and Candidatus H. heilmannii. This report associates H. felis infection in humans with severe gastric ulceration. Moreover, the suggestion can be made that the patient contracted H. felis from her dog.

Helicobacter felis and Helicobacter bizzozeronii induce gastric parietal cell loss in Mongolian gerbils.

Non-pylori helicobacter infections are associated with gastritis, gastric ulcers and MALT lymphomas in man. Approximately 50% of these are caused by helicobacters commonly found in dogs and cats, including Helicobacter felis, Helicobacter bizzozeronii and H. salomonis. In contrast to Helicobacter pylori, the virulence mechanisms of these species are unknown. In this study the virulence of H. felis, H. bizzozeronii and H. salomonis was investigated in Mongolian gerbils. Female SPF gerbils were inoculated intragastrically with H. felis, H. bizzozeronii or H. salomonis and sacrificed 3 weeks later. Fundus and antrum samples were taken for bacterial detection by PCR. A longitudinal strip covering all stomach regions was taken for histology. Gastric colonization, inflammation, apoptosis, loss of parietal cells and cell proliferation were assessed. Controls and H. salomonis inoculated gerbils were negative in PCR. H. felis and H. bizzozeronii inoculated animals were positive. H. felis inoculated animals showed loss of parietal cells extending from the limiting ridge into the fundus. A high cell proliferation rate was noticed in the mucosal area devoid of parietal cells. A dense band of apoptotic cells and large numbers of Helicobacter bacteria were seen at the transition zone between affected and normal parietal cells. In H. bizzozeronii infected gerbils, this was less pronounced. Focal apoptotic loss of gastric epithelial cells was spatially associated with the presence of bacteria especially in H. felis and to a lesser extent in H. bizzozeronii infected gerbils. This loss of cells may lead to intestinal metaplasia.