Detection of DNA from undeclared animal species in commercial elimination diets for dogs using PCR.

BACKGROUND: Elimination diets are the gold standard for the diagnosis of adverse food reactions (AFR). A broad variety of commercial diets are available containing either hydrolysed protein or novel ingredients which claim to be suitable for elimination diets. Contamination may be one factor accounting for the failure of commercial elimination diet trials. HYPOTHESIS/OBJECTIVES: To test commercial diets labelled as suitable for elimination diets for dogs, for DNA of animal origin other than that declared on the label. METHODS: Twelve commercial dry and tinned dog food products were investigated for DNA of animal origin (chicken, turkey, beef, mutton and pork) using PCR testing. RESULTS: In nine of 10 over-the-counter diets, DNA of one or more animal species other than declared on the label was identified. The DNA most frequently detected was derived from beef (n = 8) and pork (n = 6). Two hydrolysed diets only contained DNA of the declared animal source. CONCLUSIONS AND CLINICAL IMPORTANCE: Over-the-counter "single protein diets" or canned meat products cannot be recommended for the diagnosis of dogs with AFR because contamination may cause the elimination diet to fail.

Phylogenetic relationships and new genetic tools for the detection and discrimination of the three feline Demodex mites.

Two feline Demodex mite species have been described as causative agents of feline demodicosis, until recently a third species was detected. We provide an updated analysis on the phylogenetic relationship of Demodex mites. In addition, we present the first qPCR assay for the detection and differentiation of all three feline mite species in a single reaction. Specimen of Demodex cati, Demodex gatoi, and the recently discovered third species were collected from skin scrapings and fecal flotation for DNA extraction, conventional PCR, sequencing, and alignment. A total of 24 sequences of the partial 16S rRNA gene were used to estimate the evolutionary divergence in a p-distance model and a maximum likelihood phylogenetic tree. For the qPCR assay, new primers and fluorescent probes for the simultaneous detection of all three feline Demodex mites were designed. A consensus fragment of 351 bp was phylogenetically analyzed. The third species sequence of our study shares 98.6 % similarity to the available sequence in GenBank®. It is most similar to D. gatoi (82.41 %) and most distant to the canine Demodex injai (78.28 %). In contrast, D. gatoi is most similar to human Demodex brevis (87.01 %). The multiplex qPCR detected and discriminated the three different mite species in one reaction. The detection limit is ≤1.4 ng of mite DNA. The three feline Demodex species have distinct genotypes and did not cluster in one genetic clade. The species differentiation and assessment of evolutionary relationships will ultimately support correct diagnostics and treatment approaches.

The first case of Demodex gatoi in Austria, detected with fecal flotation.

Feline demodicosis is a rare parasitic condition caused by three different species of mites (Demodex cati, Demodex gatoi, and an unnamed species). D. gatoi inhabits the superficial skin layer (stratum corneum) and is easily transmitted between individual cats. A 2-year-old female spayed Cornish Rex was presented with alopecia and pruritus. The dermatological examination revealed bilateral alopecia and excoriations on trunk, limbs, and belly. The second cat in the household, a 3-year-old female spayed Thai, showed no clinical signs. Superficial and deep skin scrapings were performed and cellophane tapes applied, and living D. gatoi mites could be detected in both cats. Oral ivermectin (0.25 mg/kg every other day) was subscribed. Feces were collected from both cats and fecal flotation with sugar and zinc solutions performed. When compared to skin scrapings and cellophane tapes, D. gatoi was detected more frequently and in higher numbers in fecal samples. Our findings suggest that D. gatoi can be efficiently diagnosed with coproscopy, particularly in asymptomatic carrier animals. DNA was extracted from the flotation liquid, and a PCR protocol for the species verification was designed. A fragment targeting a 325-bp DNA fragment of the D. gatoi mitochondrial 16S rDNA gene was amplified with a 100% similarity to the D. gatoi entry in GenBank® (GI 421920216). We report the first finding of D. gatoi in Austria and propose fecal flotation as a valuable tool for mite detection. Fecal flotation liquid is suitable for DNA extraction and PCR-based species verification of D. gatoi.

Tepoxalin reduces pruritus and modified CADESI-01 scores in dogs with atopic dermatitis: a prospective, randomized, double-blinded, placebo-controlled, cross-over study.

Thirty dogs with atopic dermatitis were given tepoxalin (Zubrin, Intervet/Schering-Plough Animal Health, Boxmeer, the Netherlands) or placebo once daily for 4 weeks, followed by a wash-out period of 1 week before reversing the treatments. Pruritus was scored by the owners using the Edinburgh Pruritus Scale and one investigator employed a modification of the Canine Atopic Dermatitis Extent and Severity Index-01 (mCADESI-01) to score the physical lesions. After administration of tepoxalin there was a > or = 50% reduction in pruritus and mCADESI-01 scores in 36% and 25% of the dogs, respectively, whereas following administration of the placebo there was a > or = 50% reduction in pruritus and mCADESI-01 scores in only 25% and 16% of the dogs, respectively. Analysis of pooled data indicated that tepoxalin resulted in a significant reduction in pruritus (P = 0.012) and mCADESI-01 (P = 0.002) scores but there was no significant change after placebo. The median pruritus scores before and after tepoxalin were 2 (range 1-5) and 1 (range 0-5), respectively, and before and after placebo were 2 (range 0-4) and 2 (range 0-4), respectively. The median mCADESI scores before and after tepoxalin were 23 (range 0-68) and 16 (range 0-72), respectively, and before and after placebo were 18 (range 3-79) and 24 (range 0-65), respectively. At the dose used in this study (10.0-19.1 mg kg(-1)), tepoxalin was well-tolerated and no adverse effects were noted.