Domenico Santoro

Canine atopic dermatitis: detailed guidelines for diagnosis and allergen identification.

BackgroundCanine atopic dermatitis (AD) is a common, genetically predisposed, inflammatory and pruritic skin disease. The variation in clinical presentations, due to genetic factors, extent of the lesions, stage of the disease, secondary infections, as well as resemblance to other non-atopic related skin diseases, can complicate a diagnosis of canine AD. A sub-group of the International Committee for Allergic Diseases in Animals (ICADA) was tasked with the development of a set of practical guidelines that can be used to assist practitioners and researchers in the diagnosis of canine AD. Online citation databases and abstracts from international meetings were searched for publications related to the topic, and combined with expert opinion where necessary. The final set of guidelines was approved by the entire ICADA committee.ResultsA total of 81 publications relevant for this review were identified. The guidelines generated focus on three aspects of the diagnostic approach:1.Ruling out of other skin conditions with clinical signs resembling, or overlapping with canine AD.2.Detailed interpretation of the historical and clinical features of patients affected by canine AD.3.Allergy testing by intradermal versus allergen-specific IgE serum testing.ConclusionsThe diagnosis of canine AD is based on meeting clinical criteria and ruling out other possible causes with similar clinical signs. Flea combing, skin scraping and cytology should be performed, where necessary, as part of a thorough work-up. Elimination diet trials are required for patients with perennial pruritus and/or concurrent gastrointestinal signs. Once a clinical diagnosis of canine AD is made, allergy testing can be performed to identify potential causative allergens for allergen-specific immunotherapy.

Review: Pathogenesis of canine atopic dermatitis: skin barrier and host–micro-organism interaction

BACKGROUND Canine atopic dermatitis (AD) is a common, genetically predisposed, inflammatory and pruritic skin disease. The pathogenesis of canine AD is incompletely understood. OBJECTIVES The aim of this review is to provide an in-depth update on the involvement of skin barrier and host-microbiome interaction in the pathogenesis of canine AD. METHODS Online citation databases and abstracts from international meetings were searched for publications related to skin barrier and host-microbiome interaction (e.g. bacteria, yeast, antimicrobial peptides). RESULTS A total of 126 publications were identified. This review article focuses on epidermal barrier dysfunction and the interaction between cutaneous microbes (bacteria and yeasts) and the host (antimicrobial peptides). Epidemiological updates on the presence of pathogenic organisms and canine AD are also provided. CONCLUSIONS AND CLINICAL IMPORTANCE Major advances have been made in the investigation of skin barrier dysfunction in canine AD, although many questions still remain. Skin barrier dysfunction and host-microbiome interactions are emerging as primary alterations in canine AD. Based on this review, it is clear that future studies focused on the development of drugs able to restore the skin barrier and increase the natural defences against pathogenic organisms are needed.

Early exposure to probiotics in a canine model of atopic dermatitis has long-term clinical and immunological effects

Probiotics modulate the immune response and may have protective effects against atopic dermatitis (AD). Clinical trials using dogs with spontaneous disease are limited by confounding factors such as different diets, environments and sensitizations while a more controlled evaluation is possible using experimental models. A validated model of canine AD showed that early exposure to Lactobacillus rhamnosus GG (LGG) significantly decreases allergen-specific IgE and partially prevents AD in the first 6 months of life. This study is a follow-up three years after discontinuation of LGG. Clinical signs were evaluated after allergen challenge with ragweed, timothy, Dermatophagoides farinae. Allergen-specific IgE, IL-10 and TGF-β were measured on the 1st day of challenge, before allergen exposure. Normal dogs were included as controls. Analyses included seven dogs in the non-probiotic and nine in the probiotic litter. For clinical scores, a 2-Group × 9-Time Analysis of Variance showed significant effects of group (p=0.0003, probiotic<controls), time (p<0.0001) and group × time interaction (p<0.0001). IL-10 for all allergens was significantly higher in the control group than probiotics-exposed dogs. Allergen-specific IgE and TGF-β did not differ between litters. Early exposure to probiotics has long-term clinical and immunological effects in this model and larger studies using dogs with spontaneous disease are needed.

Expression and distribution of canine antimicrobial peptides in the skin of healthy and atopic beagles.

Antimicrobial peptides (AMPs) are small immuno-modulatory proteins important in defense against pathogenic organisms. Defensins and cathelicidin are the most frequently studied human AMPs. An increase in AMPs in atopic humans has been reported recently. Our goals were to determine the distribution of AMPs and evaluate their mRNA and protein expression in non-lesional (Day 0), acute lesional skin (Day 3) and post-challenged skin after resolution of skin lesions (Day 10) using a canine model of atopic dermatitis (AD). All dogs were environmentally challenged for three consecutive days with house dust mite. Clinical evaluation of atopic beagles was performed using a CADESI score at each time point before and after environmental challenge. Skin biopsies were taken from six healthy and seven atopic beagles before and after allergen challenge (Day 0, Day 3 and Day 10). The transcription of canine cathelicidin (cCath) and beta-defensins (cBD)-1, -2 and -3 mRNA was quantified using quantitative-RT-PCR while the protein distribution of cBD2, cBD3 and cCath was detected by indirect immunofluorescence. A significant effect, over-time, was seen in CADESI score in AD beagles with an increase score after challenge (Day 3). Quantitative analysis showed a significant difference in mRNA transcript levels between groups (with atopic dogs having more than controls) for all AMPs but cBD2. No effect over time was evident for either group. No significant differences were seen for the AMP protein patterns of distribution (homogenous distribution). Although, these results showed no differences in AMPs localization after allergen exposure in each group; atopic dogs had a higher mRNA expression of AMPs when compared with healthy dogs, a similar finding to humans.

Review: Role of genetics and the environment in the pathogenesis of canine atopic dermatitis

BACKGROUND Multiple levels of evidence support the role of genetics and the environment in the pathogenesis of canine atopic dermatitis (AD). OBJECTIVES This review summarizes the current evidence in genetics and the effect of environmental factors on the development and perpetuation of canine AD. METHODS Citation databases, abstracts and proceedings from international meetings published between 2001 and 2013 were reviewed in this update. Where necessary, older articles were included for background information. RESULTS Canine AD is a heritable disease, in which interaction with environmental factors influences disease risk and phenotype. A study of British guide dogs indicated that nearly 50% of the risk of developing AD was determined by an individuals genotype. Genomic studies performed so far in canine AD have uncovered numerous gene candidates likely to be involved in pathogenesis through their role in immunity, skin barrier formation, apoptosis and inflammation. In addition to genetics, there is evidence to suggest that exposure to certain environmental factors influences the prevalence and course of canine AD. For example, living in rural areas or feeding noncommercial diets was negatively associated with the development of AD in dogs, while exposure to high levels of smoke was associated with increased prevalence of allergic skin disease. CONCLUSIONS It is becoming clear that canine AD is genotypically complex and influenced by a variety of environmental factors. Well-designed studies with sufficient statistical power will be critical to identify the complex genetic and environmental factors involved in disease development and progression. Recognition of such factors may help to identify new targets for therapy and enable better disease prevention and management.

Altered mRNA and protein expression of filaggrin in the skin of a canine animal model for atopic dermatitis

BACKGROUND Filaggrin is a structural protein that has attracted increasing interest over the past decade for its role in the pathogenesis of human atopic dermatitis (AD). Null mutations in its sequence are considered risk factors in the development of AD. HYPOTHESIS/OBJECTIVES To investigate canine filaggrin mRNA and protein expression in the skin of atopic beagles with experimentally induced AD compared with breed-matched healthy control dogs. METHODS All dogs were environmentally challenged for 3 days consecutively with allergens to which the atopic dogs had been sensitized. Skin biopsy specimens were taken from six healthy and seven atopic beagles before and after allergen challenge. Canine filaggrin mRNA was measured using quantitative real-time PCR. Indirect immunofluorescence was used to localize the filaggrin protein in canine skin. Analysis of variance with Tukeys multiple comparison test (over-time effect) and unpaired Students t-test (treatment effect) were used. Values of P ≤ 0.05 were considered significant. RESULTS Analysis of variance showed a significantly higher expression of filaggrin mRNA in atopic dogs compared with healthy control dogs (P = 0.004 on day 3 and P = 0.01 on day 10) and a decreased mRNA expression on day 3 in healthy control dogs (effect of time, P = 0.006). On blinded evaluation, filaggrin immunofluorescence was distributed homogeneously in the stratum granulosum and the stratum corneum in healthy dogs. Atopic dogs showed a patchy immunofluorescence pattern, which was exacerbated after environmental challenge. CONCLUSIONS AND CLINICAL IMPORTANCE Altered epidermal filaggrin mRNA expression and protein distribution was detected in this experimental model.

Expression and distribution of antimicrobial peptides in the skin of healthy beagles.

Antimicrobial peptides (AMPs) are small proteins involved in defense against pathogenic organisms. Defensins and cathelicidin are the most frequently studied human AMPs. Our goals were to determine the distribution of AMPs and evaluate their mRNA expression in normal canine skin. Skin biopsies were taken from six healthy beagles. The relative transcript level of canine cathelicidin (cCath) and β-defensin (cBD)-1, cBD2 and cBD3 mRNA was quantified using quantitative real-time polymerase chain reaction. Indirect immunofluorescence (IIF), using polyclonal antibodies against cBD2, cBD3 and cCath, was used to evaluate protein localization in the skin of healthy dogs. The Pfaffl method, using experimentally determined primer efficiencies of amplification, was used to determine the expression level of cCath, cBD1 and cDB3 relative to cDB2. The levels of cCath, cBD3 and cBD1 mRNA were 358, 296 and 177 times higher than those of cBD2, respectively. Using IIF, cBD2 and cBD3 protein fluorescence was detected in all layers of the epidermis, whereas cCath was detected predominantly in the stratum granulosum and corneum. In addition, antimicrobial peptide detection was limited to the infundibular portion of the pilosebaceous units. We have validated useful methods to evaluate AMPs in canine skin. Further studies are needed to compare AMP expression in healthy dogs with that of dogs with inflammatory skin conditions.

Evaluation of Canine Antimicrobial Peptides in Infected and Noninfected Chronic Atopic Skin

BACKGROUND Antimicrobial peptides (AMPs) are small immunomodulatory peptides produced by epithelial and immune cells. β-Defensins (BDs) and cathelicidins (Caths) are the most studied AMPs. Recently, increased cutaneous expression of AMPs was reported in atopic humans and in beagles with experimentally induced atopy. HYPOTHESIS/OBJECTIVES Our goal was to analyse mRNA expression and protein levels of canine (c)BD1-like, cBD2-like/122, cBD3-like, cBD103 and cCath in healthy and naturally affected atopic dogs, with and without active skin infection, along with their distribution in the epidermis using indirect immunofluorescence. ANIMALS   Skin biopsies were taken from 14 healthy and 11 atopic privately owned dogs. METHODS The mRNA levels of cBD1-like, cBD2-like/122, cBD3-like, cBD103 and cCath were quantified using quantitative real-time PCR. The protein levels of cBD3-like and cCath were analysed by relative competitive inhibition enzyme-linked immunosorbent assay, while the distributions of cBD2-like/122, cBD3-like and cCath were detected by indirect immunofluorescence. RESULTS Dogs with atopic dermatitis had significantly greater mRNA expression of cBD103 (P = 0.04) than control dogs. Furthermore, atopic skin with active infection had a higher cBD103 mRNA expression (P = 0.01) and a lower cBD1-like mRNA expression (P = 0.04) than atopic skin without infection. No significant differences in protein levels (cBD3-like and cCath) or epidermal distribution of AMPs (cBD2-like/122, cBD3-like and cCath) were seen between healthy and atopic dogs. CONCLUSIONS AND CLINICAL IMPORTANCE Expression of cBD103 mRNA was greater, while expression of cBD1-like mRNA was lower in dogs with atopic dermatitis that had active infections. Work is needed to clarify the biological mechanisms and possible therapeutic options to maintain a healthy canine skin.

Review: Clinical and histological manifestations of canine atopic dermatitis

BACKGROUND Many studies focusing on clinical and histological signs of canine atopic dermatitis (AD) have been published since its early descriptions decades ago. Findings of these studies contributed to our current knowledge about the disease pathogenesis and allowed establishment of diagnostic criteria used by clinicians and researchers. OBJECTIVES This review serves as an update on the clinical and histological features of canine AD published by the American College of Veterinary Dermatology Task Force on Canine Atopic Dermatitis in 2001 and summarizes the recent discoveries in these fields. RESULTS The overall findings of studies focusing on clinical features mirrored those published by the Task Force in 2001. The novelty was the larger number of animals included in these studies, which allowed establishment of a new set of diagnostic criteria that exceeded the sensitivity and specificity of the previous criteria. The same study uncovered some clinical differences between dogs with food-induced and nonfood-induced AD; however, the authors concluded that these two entities cannot be distinguished based on clinical signs only. Another study demonstrated some major breed-specific phenotypes. Several publications addressed the histological features of canine AD skin lesions in experimental models of AD, but none of those addressed naturally occurring lesions. Nevertheless, the histopathological description of the skin reactions was generally similar to that published by the Task Force in 2001. CONCLUSIONS Considerable work has been done in recent years to provide a better definition of the clinical appearance and histopathology of canine AD. New sets of diagnostic criteria have been developed, and additional breed-associated differences in phenotypes have been demonstrated.

Survey of Zoonotic Dermatoses in Client-Owned Exotic Pet Mammals in Southern Italy

Several ‘exotic’ mammalian species (e.g. rabbits, rodents, ferrets and hedgehogs) live in close proximity to humans as companion pets. Skin diseases (SD) are frequent causes of morbidity in exotic pet mammals, and most of those SDs have a zoonotic potential. The purpose of this study was to determine the frequencies and types of zoonotic dermatosis (ZD) in client‐owned, exotic pet mammals in Southern Italy. Six‐hundred and fifty‐five medical records of exotic pet mammals examined between 2011 and 2012, across twenty private practice veterinary clinics around the Naples area (Italy), were retrospectively evaluated and screened for animals diagnosed with SDs (rabbits n = 455, guinea pigs n = 93, ferrets n = 64, hedgehogs n = 19, chinchillas n = 13 and rats n = 11). The records of animals diagnosed with SD, whose causative agents had a zoonotic potential, were selected for analysis. The Mann–Whitney independent test was used for statistical analysis. A P value ≤0.05 was considered statistically significant. Eighty‐two records (12.5%) of animals with ZD were identified. Of those, 56.1% (46/82) were affected by fungal infections and 42.7% (35/82) by parasitic infections. No zoonotic bacterial or viral infections were diagnosed. Dermatophytosis was significantly diagnosed more frequently in younger animals. The results of this survey indicate that exotic pet mammals may serve as active carriers for many highly contagious pathogens with zoonotic potential. Awareness and vigilance by the veterinary practitioner is crucial in the prevention of occurrences of ZDs. Children frequently come in close contact with exotic pets. To prevent the unplanned transmission of pathogen from pet to human, an active routine screening examination and preventative treatments are strongly recommended for every newly purchased pet mammal.