Detection of Escherichia coli strains producing cytotoxic necrotizing factor type two (CNF2) by enzyme-linked immunosorbent assay

Sheep and rabbit antisera were produced against lysates of E. coli strain 711 (pVir). This K-12 strain carries the Vir plasmid which codes for Cytotoxic Necrotizing Factor type 2 (CNF2). Immunoglobulin G (IgG) fractions of both immune sera were subsequently purified by a two-step precipitation method. To increase the specificity for CNF2, the sheep IgG preparation was extensively adsorbed against both a sonicated extract of isogenic K-12 strain 711 and intact phenol-treated cells of vaccine strain 711 (pVir). An enzyme-linked immunosorbent assay (ELISA) was developed to detect clinical isolates of E. coli producing CNF2, using the final preparations of rabbit and sheep IgG in a double sandwich technique. The results obtained with this CNF2-ELISA were compared to those obtained with the conventional HeLa cell cytotoxicity assay. The testing of 133 E. coli strains (49 CNF2 positive strains and 84 negative strains) resulted in no false-negative and no false-positive. Therefore, the CNF2-ELISA offers a good alternative to the HeLa cell culture assay for the detection of CNF2-producing strains where facilities for and experience with cell cultures is lacking.     

Modelling the interactions between phenology and insecticide resistance genes in the codling moth Cydia pomonella

In the codling moth Cydia pomonella (L), insecticide resistance genes have been associated with pleiotropic effects affecting phenology. In this paper, we investigated whether an increase in the frequency of insecticide resistance in field populations of C pomonella was likely to entail significant divergences in the temporal occurrence of both susceptible and insecticide-resistant individuals. For this purpose, we built a phenological model that provided suitable predictions of the distinct and diverging seasonal evolutions of populations of a susceptible and two insecticide-resistant (at two and three loci) homozygous genotypes of C pomonella. Model simulations for each genotype were further compared with pheromone trap catches recorded in a field insecticide-treated population over an 8-year period (from 1992 to 2000), which reflected the progressive annual increase in the frequency of resistance in southeastern France. We found a significant delay in field adult emergence relative to those predicted by the homozygous susceptible model, and the magnitude of such a delay was positively correlated with increasing frequencies of insecticide resistance in the sampled field population of C pomonella. Adult emergence predicted in the theoretical population that was homozygous for resistance at two loci converged with those recorded in the field during the investigated 8-year period. This suggested that the pleiotropic effects of resistance were likely to result in a significant phenological segregation of insecticide-resistant alleles in the field. The results of this study emphasized the potential for pest populations exposed to chemical selection to evolve qualitatively with respect to phenology. This may raise critical questions regarding the use of phenological modelling as a forecasting tool for appropriate resistance management strategies that would take into account the diverging seasonal evolutions of both insecticide resistance and susceptibility.     

Diabetes mellitus in a domesticated Spanish mustang

An 18-year-old Spanish Mustang mare was referred for evaluation of progressive weight loss and persistent hyperglycemia. Clinicopathologic abnormalities included marked hyperglycemia and glycosuria. Serum cortisol concentration was appropriately decreased following administration of dexamethasone, indicating that the horse did not have pituitary pars intermedia dysfunction. Serum insulin and plasma C-peptide concentrations were low, suggesting that hyperglycemia was a result of decreased secretion of insulin by pancreatic beta cells. In addition, glucose concentration did not return to the baseline concentration until 5 hours after i.v. administration of a glucose bolus, suggesting that insulin secretion, insulin effect, or both were reduced. However, i.v. administration of insulin caused only a slight decrease in the plasma glucose concentration, giving the impression that the action of insulin was impaired. Within 5 hours after administration of a combination of glyburide and metformin, which is used to treat diabetes mellitus in humans, the glucose concentration was within reference limits. The horse was euthanized, and a postmortem examination was done. Immunohistochemical staining of sections of the pancreas revealed attenuation of the pancreatic islet beta-cell population, with beta cells that remained generally limited to the periphery of the islets. These findings indicate that, albeit rare, pancreatic beta-cell failure may contribute to the development of diabetes mellitus in horses.     

Molecular epidemiology of white pine blister rust: recombination and spatial distribution

ABSTRACT Multilocus haplotypes (MLHs) were derived for the spermogonial (monokaryotic haploid) stage of Cronartium ribicola, the causal agent of white pine blister rust. Six random amplified polymorphic DNA loci and three single-strand conformational polymorphism markers were analyzed for 246 rust samples collected from two heavily infected white pine plantations. All cankers sampled were spatially located within the plantations. The hypothesis that spores are not locally disseminated was supported by the absence of any spatial clustering in the distribution of the MLHs. A large number of MLHs was found at both sites and the haplotypic diversity was close to the maximum (one) in both populations. All measures of recombination were not different from expectations under a scenario of sexual recombination. Genetic differentiation between the two sites was very low (theta = 0.023), yet it was significantly different from zero (P < 0.01). This analysis is in agreement with a scenario of extensive sexual recombination followed by some long-distance dispersal.     

Iridium-192 interstitial brachytherapy as adjunctive treatment for canine cutaneous mast cell tumors

Eleven dogs with cutaneous mast cell tumors (MCTs) were treated with surgery and iridium-192 ((192)Ir) interstitial brachytherapy. Minimum tumor doses ranged from 47.2 to 63.3 Gy. Treated tumors were classified as grade II (n=7) or III (n=4). Five dogs had recurrences with a median progression-free interval of 1391 days, and six dogs had no recurrence at a median follow-up time of 942 days. Acute adverse effects were well tolerated, and late effects were mild. One dog developed a second tumor of a different cell type in the radiation treatment field.     

Molecular investigation of Escherichia coli strains associated with apparently persistent urinary tract infection in dogs

Persistent Escherichia coli urinary tract infection (UTI) in dogs is a frustrating clinical problem. Affected dogs often appear to fail to respond to therapy or to reacquire infection shortly after therapy is completed. Urovirulence factors (UVFs) of the infecting E. coli, antibiotic resistance, and tissue colonization may be contributory but have not been evaluated in dogs with persistent E. coli UTI. In this study, the strain types of E. coli in dogs with persistent UTI were evaluated with pulsed-field gel electrophoresis (PFGE) to determine whether persistence was due to acquisition of new isolates or failure to eradicate existing isolates. UVFs in these isolates, assessed by polymerase chain reaction, and antibiograms were correlated with treatment outcome in these dogs. Results documented a mixed pattern: 9 dogs remained chronically infected with 1 or 2 strains, each with distinct reproducible UVFs, but 1 dog was infected with numerous unrelated E. coli strains over time. Two dogs had a mixed pattern, consisting of 1 or more episodes of persistent E. coli infection attributable to a single strain in addition to episodes caused by unrelated strains. Many isolates had no detectable UVFs, highlighting the likely importance of impaired colonization resistance in the affected dogs. Antibiotic resistance was common, often in response to previous treatments, especially with trimethoprim-sulfamethoxazole. Antibiotic resistance patterns differed significantly within PFGE strain types, suggesting lateral acquisition of resistance plasmids or integrons. These results can be used to help guide testing for and management of persistent E. coli UTI in dogs.     

Pharmacokinetics of enrofloxacin in neonatal kittens

OBJECTIVE: To determine the pharmacokinetics of enrofloxacin in neonatal kittens and compare the pharmacokinetics of enrofloxacin in young and adult cats. ANIMALS: 7 adult cats and 111 kittens (2 to 8 weeks old). PROCEDURE: A single dose of 5 mg of enrofloxacin/kg was administered to adults (i.v.) and kittens (i.v., s.c., or p.o.). Plasma concentrations of enrofloxacin and its active metabolite, ciprofloxacin, were determined. RESULTS: The half-life of enrofloxacin administered i.v. in 2-, 6-, and 8-week-old kittens was significantly shorter and its elimination rate significantly greater than that detected in adults. The apparent volumes of distribution were lower at 2 to 4 weeks and greater at 6 to 8 weeks. This resulted in lower peak plasma concentration (Cmax) at 6 to 8 weeks; however, initial plasma concentration was within the therapeutic range after i.v. administration at all ages. Compared with i.v. administration, s.c. injection of enrofloxacin in 2-week-old kittens resulted in similar Cmax, half-life, clearance, and area under the curve values. Enrofloxacin administered via s.c. injection was well absorbed in 6- and 8-week-old kittens, but greater clearance and apparent volume of distribution resulted in lower plasma concentrations. Oral administration of enrofloxacin resulted in poor bioavailability. CONCLUSIONS AND CLINICAL RELEVANCE: In neonatal kittens, i.v. and s.c. administration of enrofloxacin provided an effective route of administration. Oral administration of enrofloxacin in kittens did not result in therapeutic drug concentrations. Doses may need to be increased to achieve therapeutic drug concentrations in 6- to 8-week-old kittens.     

Evaluation of assays for perinuclear antineutrophilic cytoplasmic antibodies and antibodies to Saccharomyces cerevisiae in dogs with inflammatory bowel disease

OBJECTIVE: To evaluate the use of immunofluorescence asssays for perinuclear antineutrophilic cytoplasmic antibodies (pANCAs) and antibodies to Saccharomyces cerevisiae (ASCAs) in dogs with inflammatory bowel disease (IBD) and assess the clinical value of these serologic markers of the disease. ANIMALS: 39 dogs with IBD, 18 dogs with acute diarrhea, 19 dogs with chronic non-IBD-associated diarrhea, 26 healthy dogs of various breeds and age, and 22 healthy young working dogs. PROCEDURE: Sera obtained from the dogs in each group were added to canine granulocyte- and Saccharomyces cerevisiae-mounted slides for detection of pANCAs and ASCAs via immunofluorescence techniques. Sensitivity and specificity (with 95% confidence intervals [CIs]) were calculated for the group of dogs with IBD versus each of the 2 groups of healthy dogs, the group of dogs with acute diarrhea, and the group of dogs with chronic non-IBD-associated diarrhea. RESULTS: Among the 39 dogs with IBD, 20 yielded positive results via the pANCA assay (sensitivity, 0.51 [95% CI, 0.35 to 0.67]) and 17 yielded positive results via the ASCA assay (sensitivity, 0.44 [95% CI, 0.22 to 0.69]). The specificity of the pANCA assay in the 4 groups of non-IBD-affected dogs ranged from 0.83 (95% CI, 0.85 to 0.96) to 0.95 (95% CI, 0.72 to 1.00). CONCLUSIONS AND CLINICAL RELEVANCE: Immunofluorescence assays for pANCA and ASCA appear to be useful for the detection of IBD in dogs. The pANCA immunofluorescence assay had high specificity for canine IBD, and pANCAs appear to be accurate markers of intestinal inflammation.