Occurrence and identification of yeasts in dogs external ear canal with and without otitis

ABSTRACT Objective. To analyze the presence of yeast in the external ear canal of 116 dogs with and without a diagnosis of otitis from veterinary clinic in the Chapecó city, Santa Catarina, Brazil, and to examine the secretion of the proteinase in isolates. Materials and methods. Were collected cerumen of conduct hearing of dogs of 16 different races 71% with pendular ear type, 5% of semi-pendular and 24% of the erect type. All dogs were previously evaluated by otoscopy and grouped in dogs with and without otitis. Results. Yeasts were isolated in 44 samples (approximately 36%), where Malassezia pachydermatis was identified in 95% of samples where were observed growth of yeasts. On 20 samples the proteinase enzyme showed strong activity in 31% isolates, were 21% of the dogs with otitis tested showed high proteolytic activity. Conclusions. We observed a variation of strains of M. pachydermatis-producing enzymes. The variation in production of these enzymes is probably more associated with different response to the action of the immune system of the animal in the tissue injury

Quantification of Malassezia pachydermatis by real-time PCR in swabs from the external ear canal of dogs

Financial support for this project came from Servei Veterinari de Bacteriologia i Micologia from the Universitat Autònoma de Barcelona.Malassezia pachydermatis is part of the normal microbiota of canine skin and external ear canal, and is also associated with otitis externa in dogs. Laboratory detection of Malassezia otitis relies on the presence of elevated numbers of the yeast on cytologic examination of otic exudate. Although cytology has high specificity, it has low sensitivity, resulting in false-negatives and posing a challenge for clinicians to accurately diagnose Malassezia otitis. We developed a quantitative PCR (qPCR) to detect and quantify M. pachydermatis yeasts and validate the method with swabs from external ear canals of dogs. Our qPCR uses the β-tubulin gene, a single-copy gene, as a target. The limit of quantification was established as 0.18 ng/reaction, equivalent to 2.0 × 10 genome equivalents (gEq). Swabs from healthy dogs yielded quantification values of ≤2.7 × 10 gEq in the qPCR, whereas swabs from dogs with otitis yielded quantification values of ≥2.5 × 10 gEq. Our qPCR assay provides accurate quantification of M. pachydermatis yeasts from swab samples from dogs, is more sensitive than cytology, and could be used to monitor response to treatment. Our assay could also be valuable in a research setting to better understand the pathogenesis of M. pachydermatis

Is Malassezia nana the main species in horses' ear canal microbiome?

The objective of this study was to characterize genotypically Malassezia spp. isolated from the external ear canal of healthy horses. Fifty-five horses, 39 (70.9%) males and 16 (29.1%) females, from different breeds and adults were studied. External ear canals were cleaned and a sterile cotton swab was introduced to collect cerumen. A total of 110 samples were cultured into Dixon medium and were incubated at 32 degrees C for up to 15 days. Macro- and micromorphology and phenotypic identification were performed. DNA was extracted, strains were submitted to polymerase chain reaction technique, and the products obtained were submitted to Restriction Fragment Length Polymorphism using the restriction enzymes BstCI and HhaI. Strains were sent off to genetic sequencing of the regions 26S rDNA D1/D2 and ITS1-5.85-ITS2 rDNA. Malassezia spp. were isolated from 33/55 (60%) animals and 52/110 (47%) ear canals. No growth on Sabouraud dextrose agar was observed, confirming the lipid dependence of all strains. Polymerase chain reaction-Restriction fragment length polymorphism permitted the molecular identification of Malassezia nana-42/52 (81%) and Malassezia slooffiae- 10/52 (19%). Sequencing confirmed RFLP identification. It was surprising that M. nana represented over 80% of the strains and no Malassezia equina was isolated in this study, differing from what was expected. (C) 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. This is an open access article under the CC BY-NC-ND licenseCapes-PROSUP fellowship, Federal Government, BrazilUniv Paulista, Mol & Cellular Biol Lab, Sao Paulo, BrazilUniv Fed Sao Paulo, Dept Microbiol Immunol & Parasitol, Sao Paulo, BrazilUniv Fed Sao Paulo, Dept Microbiol Immunol & Parasitol, Sao Paulo, BrazilWeb of Scienc

Parasitic otitis and dermatitis in dogs in Tuscany Otiti e dermatiti parassitarie nei cani della Toscana

Otitis externa (OE) and dermatitis are very common diseases in dogs. In a population of affected dogs we focused on parasitic dermatitis and OE with the aim to evaluate their frequency and the parasitic species more frequently involved. The main symptoms associated with each isolated parasite and the frequency of some predisposing factors (sex, age, breed and living situation), were also evaluated. In the period between March 2011 and September 2012, skin and wax samples were collected from 87 dogs including owned animals living in Pisa municipality or animals hosted in kennels located in Pisa and Florence districts. For sampling collection, skin scraping, scotch test, trichogram and ear swab methods were used. In order to find parasitic arthropods, skin scrapings were digested by NaOH 10% or just mixed with few drops of paraffin oil and then checked under the microscope (100 X- 400 X ), ear wax samples were mixed with paraffin oil or directly observed , while the scotch test and trichogram samples were directly observed under the microscope. To find Malassezia, smears of ear wax samples were stained with Diff-Quick and microscopically observed at 400 X and 1000X magnifications. All data was statistically analyzed. Overall, 54% of the dogs had dermatitis, 43% had otitis and 3% had both. Out of the cases of dermatitis 44% were diagnosed with Demodicosis and 38% with Sarcoptic mange. Out of the cases of OE 67.5% were diagnosed with Malassezia, 7.5% with Malassezia and Otodectes cynotis, 2.5% with Malassezia and Trombicula autumnalis, 2.5% with O. cynotis and 20% were negative for parasites. From statistical analysis, parasitic OE was more frequent in kenneled and crossbreed dogs (P0.05). Sex was correlated neither with parasitic dermatitis nor with parasitic OE. Pruritus (100%) and erythema (89%) resulted the main clinical signs of sarcoptic mange, while pruritus (82%), alopecia (68%) and erythema (68%) were the main clinical signs of demodicosis. The main symptoms of parasitic OE (Malassezia and O. cynotis) were abundant (77%, 100%) and dark (68%, 100%) wax

Studies of the microenvironment and microflora of the canine external ear canal

The aims of this study were to investigate aspects of the aural microenvironment in dogs. The aural microflora, histological features of the aural integument, and biochemical components of cerumen from healthy canine ears and those with otitis externa were studied. Interactions between canine cerumen and one member of the resident aural flora, Malassezia pachydermatis were also investigated. Gram-positive, coagulase-negative cocci, and M.pachydermatis were the most common resident flora found in 52 healthy canine external ears. Microscopic examination of cytological smears from cerumen and microbiological culture indicated that these organisms were present in low numbers. Gram-positive coagulase positive staphylococci, Gram-negative rods, and M.pachydermatis were isolated most frequently from 27 canine ears affected by otitis externa. Eighty percent of these inflamed ears were associated with microbial overgrowth. The numbers of microorganisms found in cerumen cytological smears appear to be correlated to the growth density of microbial colonies on culture plates. In an anatomical survey of the external ear canal, 40 canine ears were examined. The average length of the cartilaginous part of these canals was 5.3 cm. The diameter at the most proximal end of the annular cartilage averaged 0.5 cm; at the proximal end of the auricular cartilage the mean diameter was 0.7 cm; at the distal extremity of the ear canal, the average diameter was 4.8 cm. Morphometric stereology was used to evaluate histological features of 28 clinically normal and 15 otitic canine ears at four anatomical levels. Marked variation was found in the distribution of sebaceous and apocrine glands in the aural integument in healthy ears and those with otitis

Antibacterial and antifungal activity of essential oils against pathogens responsible for otitis externa in dogs and cats

Essential oils (EOs) are recommended by some veterinarians to treat otitis externa in pets, but data about their efficacy in scientific literature are very scant. Methods: Nine commercial EOs, from roman chamomile (Anthemis nobilis L.), star anise (Illicium verum), lavender (Lavandula hybrida), litsea (Litsea cubeba (Lour.) Pers.), basil (Ocimum basilicum L.), oregano (Origanum vulgare L. subsp. hirticum), rosemary (Rosmarinus officinalis L.), clary sage (Salvia sclarea L.), and thyme (Thymus vulgaris L.) were tested against bacterial and fungal pathogens previously isolated from dogs and cats with otitis externa. In particular, the analyses were carried out against Pseudomonas aeruginosa, Staphylococcus aureus, Staphylococcus pseudointermedius, Aspergillus niger, Aspergillus fumigatus, Aspergillus terreus, Candida albicans, Candida tropicalis, Trichosporon sp., and Rhodotorula sp. Results: O. vulgare and S. sclarea showed superior antibacterial activity, even if not against all the strains. Trichosporon sp., C. albicans, and A. terreus were insensitive to most Eos, while other yeasts and molds showed different degrees of sensitivity. In particular, most fungi were inhibited by O. vulgare and R. officinalis. Conclusions: The obtained results suggest that some EOs could be included in treatment as an alternative therapeutic option in bacterial otitis complicated by fungi, in association with conventional drugs

Study of the variation of the Malassezia load in the interdigital fold of dogs with pododermatitis

Acord transformatiu CRUE-CSICThe yeast Malassezia pachydermatis is a common inhabitant of the skin and mucosae of dogs. However, under certain circumstances this yeast can overgrow and act as an opportunistic pathogen causing otitis and dermatitis in dogs. Canine pododermatitis is a common disorder in dogs in which M. pachydermatis acts as an opportunistic pathogen. In the present study, the presence of Malassezia yeasts was assessed and quantified in samples collected from the interdigital space of dogs with pododermatitis before and after treatment, and from healthy dogs. The samples were subjected to two different cytological examinations, culture on Sabouraud glucose agar and modified Dixon's agar and a quantitative PCR targeting the internal transcribed spacer (ITS) genomic region. A selection of samples was analyzed by next generation sequencing (NGS) using the D1D2 domain of the large subunit of the ribosomal DNA as target. The pododermatitis samples before treatment showed higher cell counts, colony-forming units and ITS copies than the rest of samples. The NGS analysis revealed that Ascomycota was the main phylum in the healthy and post-treatment samples. However, Basidiomycota and M. pachydermatis was more abundant in the pododermatitis samples before treatment. These results support M. pachydermatis as an opportunistic agent in canine pododermatitis by a variety of methods, and demonstrate the correlation between cytologic and molecular methods for quantification

L-Mesitran® in the management of canine otitis externa

Dissertação de Mestrado Integrado em Medicina VeterináriaAt a time when antimicrobial resistance is rising steadily and the involved microorganisms are demonstrating zoonotic potential, honey and its derived products may prove useful in this ongoing battle. Otitis externa in dogs is considered to be one of the most prominent causes for presentation at veterinary practice. Some of the regularly administered agents in otitis treatments are no longer effective, as resistance has increased, perhaps due to the often long-term periods which are necessary for resolution and the accompanying tendency towards chronicity. In order to address the need for efficient alternative treatments, LMesitran ® Soft, a medical grade honey gel was used to treat 15 dogs with otitis externa of bacterial and/or fungal involvement. Success was based on clinical score decrease, cytology and owner input over time and with basis on culture results. 70% of enrolled dogs achieved clinical cure between days 7 to 14 and over 90% on day 21, the maximum established time limit, with a confidence interval of 95%. Furthermore, by day 7, 20% of dogs had obtained both clinical and cytological cures. This study was successfully able to demonstrate that the use of L-Mesitran® was effective in managing otitis externa in dogs, including cases in which highly resistant pathogens were present, such as methicillinresistant Staphylococcus pseudintermedius (MRSP), thus paving the way to future studies.ABSTRACT - L-MESITRAN® NO MANEIO DE OTITE EXTERNA CANINA – UM ESTUDO PILOTO - Numa altura em que nos deparamos com um aumento de bactérias com resistências aos antibióticos e em que os organismos envolvidos apresentam por vezes inclusivamente potencial zoonótico, o recurso ao mel e seus derivados pode ser uma inestimável ferramenta no decurso desta batalha. A otite externa em cães é um estímulo iatrotrópico frequente e das principais causas de idas ao médico veterinário. Alguns dos tratamentos habitualmente utilizados deixaram de ser eficazes à medida que as resistências surgiram, talvez consequência de terapêuticas prolongadas e recorrentes. Neste estudo avaliou-se uma potencial alternativa à terapêutica, recorrendo-se ao L-Mesitran® Soft, uma pomada contendo mel de grau clínico, para tratar 15 cães com otite externa de envolvimento bacteriano e/ou fúngico. A resposta foi considerada positiva de acordo com a diminuição da pontuação clínica, citologia e opinião dos donos, no decorrer do tempo. Estabelecido o limite de 21 dias, 70% dos cães tratados obteve cura clínica entre os dias 7 e 14 e mais de 90% no dia 21, com um intervalo de confiança de 95%. Ainda, até ao dia 7, 20% dos cães havia obtido cura clínica e citológica. Este estudo demonstrou que o L-Mesitran® foi eficaz no maneio dos casos de otite externa, incluindo aqueles em que estavam presentes bactérias com várias resistências aos antibióticos, como é o caso do Staphylococcus pseudintermedius com resistência à meticilina (SPRM), abrindo assim caminho para futuros estudos

A study of the bacterial and fungal flora of the canine ear: with particular reference to the condition known clinically as 'canker' (otitis externa and media)

The commonest breeds to be affected with otitis were Spaniels and other long- haired dogs with overlapping ears.The most important of the predisposing factors to otitis were skin diseases from which 38 and 59 per cent respectively of the dogs in the unselected and chronic otitis groups were suffering. Only 13 per cent of the dogs were infested with ectoparasites.In most cases the primary lesion of otitis was due to the extension of a skin condition to the external acoustic meatus or, occasionally, to irritation by ear mites and foreign bodies such as grit and grass awns.Infection of the ear lesion was due either to commensal staphylococci or yeasts, or to faecal contamination with Pseudomonas, Proteus or coliform organisms.The incidence of Pseudomonas and Proteus was greatest in dogs with chronic otitis that were also suffering from a clinical skin disease. In a number of cases infection occurred during, or shortly after, a course of treatment with antibiotics.Pseudomonas, Proteus and coliform infections were generally associated with the copious, purulent types of discharge, whereas staphylococci and Pityrosporum predominated in the dryish, dark - coloured exudates.The tissue changes in affected ears were more closely related to the nature of the discharge and the identity of the infectious agent than to the duration of the condition. The numerous, large, cystic diverticula of the tubular portions of the modified ceruminous, or apocrine, glands were a constant feature of the more purulent forms of otitis.All the Pseudomonas strains from infected ears were identified as Ps. aeruginosa, a number of which were sensitive to phages of human strains.Pr. mirabilis accounted for 95 per cent of the Proteus strains, about half of which were antigenically similar. To some extent, species identification was determined by the strain's in vitro sensitivities to antibiotics.The commonest coliform organisms recovered from infected ears were faecal type 1 of E. coli.Unlike Group G. haemolytic streptococci, Group M. strains were rarely isolated from otitic material, although they were frequently present in the tonsils of healthy dogs.Canine staphylococci differed markedly from those from human and certain other animal sources. Most of the dog strains were non -pigmented and coagulated rabbit but not human plasmas. Their coagulase activity was positively correlated with the production of delta but not alpha haemolysin, while a number of coagulase positive strains formed a new haemolysin. Of the few (7 per cent) penicillin resistant dog staphylococci, most were of the alpha -delta haemolytic pattern and produced aureus pigment which which are features of human, rather than canine, strains.Pit rosporum species were isolated from most of the healthy and infected external ears, but from no other site, and differed from other members of the genus by growing well in maltose agar without the addition of oleic acid