The effect of Psoroptes ovis infestation on ovine epidermal barrier function.

Sheep scab is an intensively pruritic, exudative and allergic dermatitis of sheep caused by the ectoparasitic mite Psoroptes ovis. The purpose of the present study was to investigate the effect of P. ovis infestation on different components of the ovine epidermal barrier within the first 24 hours post-infestation (hpi). To achieve this, the expression of epidermal differentiation complex (EDC) genes and epidermal barrier proteins, the nature and severity of epidermal pathology and transepidermal water loss (TEWL) were evaluated.By 1 hpi a significant dermal polymorphonuclear infiltrate and a significant increase in TEWL with maximal mean TEWL (598.67 g/m2h) were observed. Epidermal pathology involving intra-epidermal pustulation, loss of epidermal architecture and damage to the basement membrane was seen by 3 hpi. Filaggrin and loricrin protein levels in the stratum corneum declined significantly in the first 24 hpi and qPCR validation confirmed the decrease in expression of the key EDC genes involucrin, filaggrin and loricrin observed by microarray analysis, with 5.8-fold, 4.5-fold and 80-fold decreases, respectively by 24 hpi.The present study has demonstrated that early P. ovis infestation disrupts the ovine epidermal barrier causing significant alterations in the expression of critical barrier components, epidermal pathology, and TEWL. Many of these features have also been documented in human and canine atopic dermatitis suggesting that sheep scab may provide a model for the elucidation of events occurring in the early phases of atopic sensitisation.

Prednisolone therapy for atopic dermatitis is less effective in dogs with lower pretreatment serum 25-hydroxyvitamin D concentrations.

Serum 25-hydroxyvitamin D [25(OH)D] concentrations were measured in 20 dogs with atopic dermatitis prior to treatment with a standard therapeutic dosage of prednisolone (0.93-1.06 mg/kg) every other day for 5 weeks after 7 days of treatment with the same dosage once daily. The severity of their physical signs was scored before and 6 weeks after prednisolone treatment by the canine atopic dermatitis extent and severity index version 3 (CADESI-03) and the Edinburgh Pruritus Scale (EPS). The 20 dogs with atopic dermatitis that were treated with prednisolone did not have significantly lower serum concentrations of 25(OH)D than a group of 36 healthy dogs, and the physical severity of the atopic dermatitis was not correlated to pretreatment serum 25(OH)D concentrations. However, dogs which had a marked improvement of their physical signs, defined by a post-treatment EPS score of 0 and/or an 85% reduction in CADESI-03 score, had significantly higher pretreatment serum 25(OH)D concentrations than dogs with a suboptimal response (P = 0.003 and P = 0.03, respectively). Serum 25(OH)D concentrations were also measured in a previously published cohort of atopic dogs that were treated with ciclosporin. This cohort of dogs was recruited in a similar time frame to the prednisolone-treated dogs, and all samples were handled in the same way. In contrast to the prednisolone-treated dogs, there was no significant difference in 25(OH)D concentrations in dogs that responded optimally to ciclosporin compared with suboptimal responders. Additional studies are required to establish whether vitamin D has a synergistic therapeutic effect with prednisolone in dogs with atopic dermatitis.

Staphylococccus pseudintermedius surface proteins SpsD and SpsO mediate adherence to ex vivo canine corneocytes.

The Gram-positive bacterium Staphylococcus pseudintermedius is regarded as the major cause of canine bacterial pyoderma. Despite its clinical importance, there is only very limited knowledge about the pathogenesis of S. pseudintermedius infection and the specific bacterial virulence factors involved in causing disease. Using a whole-genome approach, we have previously identified 18 predicted cell-wall-anchored surface proteins representing possible virulence factors in a clinical isolate of S. pseudintermedius (strain ED99). They were designated S. pseudintermedius surface proteins A-R (SpsA-SpsR). The present study tested three of the putative Sps proteins (SpsD, SpsL and SpsO) for their ability to mediate adherence of bacteria to canine corneocytes. The three proteins were expressed on the surface of the nonpathogenic surrogate host Lactococcus lactis, a Gram-positive bacterium that does not adhere to canine corneocytes. Adherence assays were performed using corneocytes from different healthy canine donors (n = 5), and bacterial cells were quantified using computerized image analysis. Two of the proteins, SpsD and SpsO, mediated adherence of L. lactis to canine corneocytes, suggesting that they contribute to S. pseudintermedius pathogenesis and may represent novel therapeutic targets to combat infection.

Ciclosporin therapy is associated with minimal changes in calcium metabolism in dogs with atopic dermatitis.

BACKGROUND: Ciclosporin is widely used in the management of canine atopic dermatitis. In humans, ciclosporin therapy has been linked to disturbances in calcium metabolism and resultant skeletal disorders. OBJECTIVES: The objective of this study was to assess calcium homeostasis in dogs before and after a 6 week course of once daily oral ciclosporin at the licensed dose (5 mg/kg). ANIMALS: Sixteen client-owned dogs with spontaneous atopic dermatitis. METHODS: Serum concentrations of calcium, phosphate, creatinine, 25-hydroxyvitamin D, 1,25-dihyroxyvitamin D and plasma concentrations of ionized calcium and parathyroid hormone (PTH) were measured, together with the urinary fractional excretion of calcium and phosphate. The extent of skin lesions was scored using the Canine Atopic Dermatitis Extent and Severity Index (CADESI)-03 and the degree of pruritus by the Edinburgh Pruritus Scale prior to and at the end of the study. RESULTS: The CADESI-03 and the Edinburgh Pruritus Scale scores decreased satisfactorily in all dogs by the end of the study. Plasma PTH concentrations were significantly increased (P = 0.02) following ciclosporin treatment, whereas all other biochemical parameters were not significantly different from their starting values. The increase in PTH was mild in most cases and the proportion of dogs that had a PTH concentration above the reference range was not significantly different following treatment. CONCLUSIONS AND CLINICAL IMPORTANCE: This study indicates that ciclosporin has minimal impact on calcium metabolism in dogs with atopic dermatitis when used at the licensed and clinically effective dosage for 6 weeks.

Complete genome sequence of the canine pathogen Staphylococcus pseudintermedius.

We report the first whole-genome sequence for a clinical isolate of Staphylococcus pseudintermedius (ED99), the major pathogen responsible for canine bacterial pyoderma. S. pseudintermedius contains numerous mobile genetic elements and encodes an array of putative virulence factors, including superantigenic, cytolytic, and exfoliative toxins and cell wall-associated surface proteins.

Genomic and surface proteomic analysis of the canine pathogen Staphylococcus pseudintermedius reveals proteins that mediate adherence to the extracellular matrix.

Cell wall-associated (CWA) proteins made by Gram-positive pathogens play a fundamental role in pathogenesis. Staphylococcus pseudintermedius is a major animal pathogen responsible for the canine skin disease bacterial pyoderma. Here, we describe the bioinformatic analysis of the family of 18 predicted CWA proteins encoded in the genome of S. pseudintermedius strain ED99 and determine their distribution among a phylogenetically diverse panel of S. pseudintermedius clinical isolates and closely related species of the Staphylococcus intermedius group. In parallel, we employed a proteomic approach to identify proteins presented on the surface of strain ED99 in vitro, revealing a total of 60 surface-localized proteins in one or more phases of growth, including 6 of the 18 genome-predicted CWA proteins. Based on these analyses, we selected two CWA proteins (SpsD and SpsL) encoded by all strains examined and investigated their capacity to mediate adherence to extracellular matrix proteins. We discovered that SpsD and SpsL mediated binding of a heterologous host, Lactococcus lactis, to fibrinogen and fibronectin and that SpsD mediated binding to cytokeratin 10, a major constituent of mammalian skin. Of note, the interaction with fibrinogen was host-species dependent, suggestive of a role for SpsD and SpsL in the host tropism of S. pseudintermedius. Finally, we identified IgG specific for SpsD and SpsL in sera from dogs with bacterial pyoderma, implying that both proteins are expressed during infection. The combined genomic and proteomic approach employed in the current study has revealed novel host-pathogen interactions which represent candidate therapeutic targets for the control of bacterial pyoderma.

Ciclosporin A therapy is associated with disturbances in glucose metabolism in dogs with atopic dermatitis.

The effect of ciclosporin A (CsA) on glucose homeostasis was investigated in 16 dogs with atopic dermatitis by determining plasma glucose, serum fructosamine and insulin concentrations, and serial insulin and glucose concentrations following a glucagon stimulation test, before and 6 weeks after CsA therapy at 5 mg/kg once daily. All dogs completed the study. Following CsA treatment, the median serum fructosamine concentrations were significantly higher (pretreatment 227.5 µmol/L; post-treatment 246.5 µmol/L; P = 0.001; reference range 162-310 µmol/L). Based on analyses of the areas under concentration-time curves (AUC) pre- and post-CsA treatment, plasma glucose concentrations were significantly higher (AUC without baseline correction 31.0 mmol/L/min greater; P = 0.021) and serum insulin concentrations were significantly lower (AUC without baseline correction 217.1 µIU/mL/min lower; P = 0.044) following CsA treatment. Peak glucose concentrations after glucagon stimulation test were significantly higher following CsA treatment (10.75 versus 12.05 mmol/L; P = 0.021), but there was no significant difference in peak serum insulin (52.0 versus 35.0 µIU/mL; P = 0.052). There was a negative correlation between baseline uncorrected insulin AUC and trough serum log CsA concentrations (r = -0.70, P = 0.005). The administration of CsA to dogs with atopic dermatitis leads to disturbances in glucose homeostasis. The clinical significance of this is unclear, but it should be taken into account when considering CsA treatment in dogs that already have such impairments.

Idiopathic generalized sebaceous gland hyperplasia of the Border terrier: a morphometric study.

Skin biopsies from five Border terriers with histologically confirmed idiopathic, generalized sebaceous gland hyperplasia (Group A) were compared morphometrically to those from four unaffected Border terriers (Group B) and the unaffected dogs to biopsies from four other terrier breeds (Group C). Dogs in Group A had significantly higher numbers of sebaceous gland lobules per hair follicle than those in Group B (P=0.020) but there was no significant difference between Groups B and C. The total sebaceous gland lobular area per hair follicle was significantly higher in dogs in Group A than Group B (P=0.020) but there were no differences between Groups B and C. There were no significant differences in the size of the individual sebaceous gland lobules. There were no significant differences in the total number of basal or mature sebocytes between Groups A and B but significantly lower numbers of both cell types were seen in Group C compared to Group B. There were significantly more basal sebaceous cells undergoing mitosis in dogs in Group A than Group B (P=0.017) but no significant difference between Groups B and C. These results indicate that the physical signs of sebaceous gland hyperplasia are caused by an increase in the number of sebaceous gland lobules and total lobular area. Border terriers may be genetically predisposed to the development of idiopathic generalized sebaceous gland hyperplasia compared with other dog breeds but also to sebaceous gland hyperplasia secondary to other dermatopathies.

Cutaneous Alternaria infectoria infection in a dog in association with therapeutic immunosuppression for the management of immune-mediated haemolytic anaemia.

A 4-year-old, ovariohysterectomized, English springer spaniel on immunosuppressive therapy was re-examined for the review of its immune-mediated haemolytic anaemia and the recent development of skin lesions. For the 3 months since hospital discharge, the dog had been receiving 1.3 mg/kg prednisolone and 2.6 mg/kg ciclosporin, both administered orally twice daily. Physical examination revealed hepatomegaly and multiple, purulent, crusting, erosive to ulcerative lesions over different body areas. Onychorrhexis had occurred on one digit and the underlying corium had blackened. There were two proliferative and one plaque-like lesions in the mouth. Thick walled fungal hyphae were detected in impression smears from all skin lesions and staining with periodic acid-Schiff's stain confirmed the presence of multiple fungal hyphae and spores in all biopsies examined. Fungal culture isolated a heavy, pure growth of an Alternaria sp. which was identified as A. infectoria by sequencing the internal transcribed spacer 1 region of the rRNA gene. The animal's condition prevented detailed investigation of the oral lesions. Withdrawal of the ciclosporin and reduction of the prednisolone dosage resulted in spontaneous resolution of the skin lesions within 40 days. Further gradual decrements in the prednisolone dosage to zero were carried out without recurrence of the immune-mediated haemolytic anaemia. After 12 months, there has been no recurrence of either the skin lesions or the anaemia. To the authors' knowledge, this is the first reported case of A. infectoria infection in a dog.

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.

Population genetic structure of the Staphylococcus intermedius group: insights into agr diversification and the emergence of methicillin-resistant strains.

The population genetic structure of the animal pathogen Staphylococcus intermedius is poorly understood. We carried out a multilocus sequence phylogenetic analysis of isolates from broad host and geographic origins to investigate inter- and intraspecies diversity. We found that isolates phenotypically identified as S. intermedius are differentiated into three closely related species, S. intermedius, Staphylococcus pseudintermedius, and Staphylococcus delphini. S. pseudintermedius, not S. intermedius, is the common cause of canine pyoderma and occasionally causes zoonotic infections of humans. Over 60 extant STs were identified among the S. pseudintermedius isolates examined, including several that were distributed on different continents. As the agr quorum-sensing system of staphylococci is thought to have evolved along lines of speciation within the genus, we examined the allelic variation of agrD, which encodes the autoinducing peptide (AIP). Four AIP variants were encoded by S. pseudintermedius isolates, and identical AIP variants were shared among the three species, suggesting that a common quorum-sensing capacity has been conserved in spite of species differentiation in largely distinct ecological niches. A lack of clonal association of agr alleles suggests that assortive recombination may have contributed to the distribution of agr diversity. Finally, we discovered that the recent emergence of methicillin-resistant strains was due to multiple acquisitions of the mecA gene by different S. pseudintermedius clones found on different continents. Taken together, these data have resolved the population genetic structure of the S. intermedius group, resulting in new insights into its ancient and recent evolution.

The use of ciclosporin A in veterinary dermatology.

Ciclosporin A (CsA) has potent immunosuppressive and immunomodulatory activity that has been exploited in human medicine to prevent the rejection of transplanted organs and to manage atopic dermatitis and psoriasis. Over the past decade, CsA has been employed more frequently in veterinary dermatology and its value in the management of several canine and feline dermatoses is now well established. CsA inhibits calcineurin phosphatase, suppressing T cell activation and the synthesis of T cell cytokines consequently impairing the activity of B cells, antigen-presenting cells, mast cells, basophils and eosinophils. The pharmacokinetics of CsA are similar in humans, dogs and cats and the drug has a wide safety margin in dogs, cats and rabbits. Adverse effects, principally transient vomiting and soft faeces/diarrhoea, may be seen shortly after instituting treatment but often resolve despite continuing treatment. Gingival hyperplasia and cutaneous effects such as hirsutism may occur after prolonged treatment.

Prednisolone is associated with an increase in serum insulin but not serum fructosamine concentrations in dogs with atopic dermatitis.

The aim of this study was to assess the effects of a standard therapeutic protocol of prednisolone (Pred) on glucose homeostasis in atopic dogs and compare it with previously published data for ciclosporin A (CsA). The central aim of the study was to assess and compare the effects of standard therapeutic protocols of prednisolone (Pred) and ciclosporin A (CsA) on glucose homeostasis in dogs with atopic dermatitis (CAD). Both treatments significantly reduced the physical signs of CAD, as determined by the canine atopic dermatitis extent and severity index version 3 (CADESI-03) and the Edinburgh Pruritus Scale (EPS). Post-treatment plasma glucose concentrations were not significantly different in the two groups, but serum insulin concentrations were significantly higher following Pred therapy (P<0.05). Serum fructosamine concentrations were not significantly different pre- and post-treatment with Pred, although previous studies had shown that CsA treatment increased fructosamine concentrations (P<0.005). The two treatment groups were recruited in a similar timeframe, were numerically matched and there were no differences in CADESI-03 and EPS scores between the CsA and Pred groups either before or after treatment. Thus, both CsA and Pred treatment were associated with mild disturbances in glucose metabolism, but only CsA therapy resulted in a significant increase in fructosamine concentrations. This information may be relevant to clinicians when considering therapeutic options for dogs with atopic dermatitis which already have impaired glucose homeostasis.

Comparative genomics of the Staphylococcus intermedius group of animal pathogens.

The Staphylococcus intermedius group consists of three closely related coagulase-positive bacterial species including S. intermedius, Staphylococcus pseudintermedius, and Staphylococcus delphini. S. pseudintermedius is a major skin pathogen of dogs, which occasionally causes severe zoonotic infections of humans. S. delphini has been isolated from an array of different animals including horses, mink, and pigeons, whereas S. intermedius has been isolated only from pigeons to date. Here we provide a detailed analysis of the S. pseudintermedius whole genome sequence in comparison to high quality draft S. intermedius and S. delphini genomes, and to other sequenced staphylococcal species. The core genome of the SIG was highly conserved with average nucleotide identity (ANI) between the three species of 93.61%, which is very close to the threshold of species delineation (95% ANI), highlighting the close-relatedness of the SIG species. However, considerable variation was identified in the content of mobile genetic elements, cell wall-associated proteins, and iron and sugar transporters, reflecting the distinct ecological niches inhabited. Of note, S. pseudintermedius ED99 contained a clustered regularly interspaced short palindromic repeat locus of the Nmeni subtype and S. intermedius contained both Nmeni and Mtube subtypes. In contrast to S. intermedius and S. delphini and most other staphylococci examined to date, S. pseudintermedius contained at least nine predicted reverse transcriptase Group II introns. Furthermore, S. pseudintermedius ED99 encoded several transposons which were largely responsible for its multi-resistant phenotype. Overall, the study highlights extensive differences in accessory genome content between closely related staphylococcal species inhabiting distinct host niches, providing new avenues for research into pathogenesis and bacterial host-adaptation.

Generalized alopecic and cystic dermatosis in a cat: a counterpart to the hairless mouse phenotype or a unique congenital dermatosis?

A 2-year-old, male neutered, domestic semi-long-haired cat was presented with a 1.5-year history of progressive, initially nonpruritic alopecia and malodorous greasy exudate affecting the distal extremities, trunk and neck but sparing the head and tail. The extensive alopecia and 'seborrhoea' were associated with severe thickening of the skin and fold formation on the dorsal head and distal extremities as well as the lateral thorax and abdomen. The hair was easily epilated, numerous milia were seen on the ventral abdomen and the caudal and lateral thighs, and mild paronychia was present. Histopathological examination of skin biopsies revealed marked cystic dilation of hair follicles and sebaceous glands with follicular hypoplasia, infundibular hyperkeratosis and variable associated inflammation. Systemic glucocorticoid therapy in combination with topical washes with chlorhexidine and miconazole resulted in a marked improvement and some hair regrowth, but the cat was subsequently lost to follow-up. The dermatosis resembles a number of conditions in other species, but it is not clear whether it is a counterpart to the hairless mutant mouse or is a unique dermatosis.