Establishment of a gene recombination method for a major human skin commensal fungus, Malassezia restricta, using Agrobacterium tumefaciens-mediated gene transfer system.

Malassezia restricta is the most predominant fungus in the microbiome of human skin. This microorganism can cause or exacerbate Malassezia-associated skin dermatitis, seborrheic dermatitis, atopic dermatitis, and pityriasis versicolor. The virulence factors of M. restricta have not been analyzed because a gene recombination system has not been developed. In this study, we established an Agrobacterium tumefaciens-mediated gene transfer (ATMT) system, optimized for generating gene-deficient mutants of M. restricta. A mutant of FKB1 gene, which encodes the FKBP12 protein that binds to the calcineurin inhibitor tacrolimus, was generated using the ATMT system. Subsequently, the FKB1 gene was reintroduced into the FKB1 gene-deficient mutant for obtaining a gene-complemented strain. The wild-type strain of M. restricta was sensitive to tacrolimus, whereas the FKB1 gene-deficient mutant was resistant to tacrolimus; the phenotypic drug susceptibility in the mutant was restored by reintroducing the FKB1 gene. Contrastingly, the FKB1 gene-deficient mutant was not resistant to cyclosporine A, which also inhibits calcineurin by binding to cyclophilin A. The gene recombination system for M. restricta will facilitate in elucidating the molecular mechanisms causing Malassezia-associated dermatitis.

Leishmaniose visceral em um Pastor Belga: relato de caso / Visceral leishmaniasis in a Belgian Pastor: case report

O presente trabalho tem como objetivo relatar um caso de um Pastor Belga, do município de Ponta Porã, Mato Grosso do Sul, positivo para Leishmaniose Visceral, atendido em 2017 em uma clínica veterinária localizada em Pedro Juan Caballero, Paraguai. O diagnóstico foi confirmado através dos sinais clínicos característicos, e dos exames ELISA e PCR positivos. O animal foi submetido ao tratamento clínico para melhora dos sintomas, cujo tratamento antiparasitário inicial foi realizado com a associação de estibogluconato de sódio 75 mg/kg e alopurinol 100 mg seguido de aloputinol 100mg de uso contínuo e uso da coleira antileishmaniose. Tratamento esse considerado eficiente, com melhora clínica do animal. Após 24 meses o animal foi diagnosticado com tumor de mama e lesão da bolsa escrotal, sendo submetido a tratamento clínico e cirúrgico. Com 30 e 36 meses do diagnóstico inicial repetiu-se os exames ELISA (positivo) e PCR (negativo), e então o animal foi considerado curado clinicamente devido à ausência de sinais clínicos. Tendo em vista a complexidade dos fatores no ciclo de transmissão, conclui-se que as medidas em saúde ainda são insuficientes para o controle efetivo da doença. É importante o papel do Médico Veterinário na saúde pública, devido a obrigatoriedade de notificação de casos de Leishmaniose Visceral Canina, sendo necessários esforços nas diferentes áreas da saúde animal, humana e do meio ambiente, visando medidas de vigilância e controle da doença no país.

The present work aims to report a case of a Belgian Shepherd, from the municipality of Ponta Porã, Mato Grosso do Sul, positive for Visceral Leishmaniasis, treated in 2017 in a veterinary clinic located in Pedro Juan Caballero, Paraguay. The diagnosis was confirmed through the characteristic clinical signs, and the positive ELISA and PCR tests. The animal was submitted to clinical treatment for improvement of symptoms, whose initial antiparasitic treatment was performed with the association of sodium stibogluconate 75 mg/kg and allopurinol 100 mg followed by alloputinol 100mg of continuous use and use of the antileishmaniasis collar. This treatment was considered efficient, with clinical improvement of the animal. After 24 months the animal was diagnosed with a breast tumor and scrotum injury, and was submitted to clinical and surgical treatment. At 30 and 36 months from the initial diagnosis, the ELISA tests (positive) and PCR (negative) were repeated, and then the animal was considered clinically cured due to the absence of clinical signs. Considering the complexity of the factors in the transmission cycle, it is concluded that the health measures are still insufficient for the effective control of the disease. The role of the veterinarian in public health is important, due to the obligatory notification of cases of Canine Visceral Leishmaniasis, being necessary efforts in the different areas of animal health, human and environment, aiming at measures of surveillance and control of the disease in the country.

Biofilm production and antifungal susceptibility of co-cultured Malassezia pachydermatis and Candida parapsilosis isolated from canine seborrheic dermatitis.

The yeasts Malassezia (M.) pachydermatis and Candida (C.) parapsilosis are often co-isolated in case of canine seborrhea dermatitis (SD) and also are emerging as opportunistic pathogens of immunocompromised human beings. Increased information about how their relationship results in biofilm production and an antifungal response would be useful to inform treatment and control. This study was designed to investigate biofilm production derived from co-culture of M. pachydermatis and C. parapsilosis from dog skin and to determine their in vitro antifungal susceptibility. We demonstrated that regardless of yeast strain or origin all single and dual cultures produced biofilms within 24 hours, and the greatest amount was present after 72 hours. Biofilm production from mixed cultures was greater than for single strains (P < .05). All sessile forms of the single and dual cultures were resistant to the tested antifungals itraconazole and ketoconazole, whereas planktonic forms were susceptible. The study suggests that dual cultures produce stronger biofilms that are likely to enhance persistence in skin lesions in dogs and result in greater resistance to antifungal treatment.

Producción de fosfolipasa y proteinasa en cepas de Malassezia pachydermatis aisladas de perros con otitis y sin otitis / Phospholipase and proteinase production by Malassezia pachydermatis isolated in dogs with and without otitis

Antecedentes. Malassezia pachydermatis forma parte de la microbiota cutánea de perros y gatos. M. pachydermatis se ha asociado frecuentemente a otitis externa y dermatitis seborreicas, sobre todo en el perro, y con menor frecuencia en el gato. M. pachydermatis podría actuar como patógeno cuando existen alteraciones en los mecanismos físicos, químicos o inmunológicos de la piel. Se han identificado diversos factores de virulencia como la capacidad de producir estearasas, lipasas, lipooxigenasas, proteinasas, condroitinsulfatasas e hialuronidasas. Objetivos. Se ha estudiado la actividad fosfolipasa medida a pH 6,3 y la actividad proteinasa medida a pH 6,3 y pH 6,8 (pH de oídos de perros con otitis) de cepas de M. pachydermatis aisladas de perros con otitis y sin otitis. Métodos. Se ha estudiado la actividad fosfolipasa mediante un método semicuantitativo con yema de huevo y la actividad proteinasa mediante un método semicuantitativo con agar albúmina sérica bovina. Se ha realizado el estudio en 96 aislamientos de M. pachydermatis, 43 de ellos aislados de perros sin sintomatología clínica de otitis y 52 aislados de perros con otitis. Resultados. Se observó que el 75,8% de los aislamientos presentaron actividad fosfolipasa a pH 6,3 y el 81% presentaron actividad proteinasa medida a pH 6,3, y el 97,9% a pH 6,8. Se detectó una mayor actividad fosfolipasa en cepas aisladas de perros con otitis. Con respecto a la actividad proteinasa, esta fue mayor a pH 6,8. Conclusiones. Estos hallazgos sugieren que la actividad fosfolipasa podría jugar un papel importante en la invasión de los tejidos del hospedador, por lo menos en la otitis crónica canina. Con respecto a la actividad proteinasa, estos hallazgos podrían ayudar a mejorar la terapéutica de la otitis cuando está implicada M. pachydermatis en el proceso, ajustando a pH bajos los tratamientos aplicados (AU)

Antifungal agent susceptibilities and interpretation of Malassezia pachydermatis and Candida parapsilosis isolated from dogs with and without seborrheic dermatitis skin.

Malassezia pachydermatis and Candida parapsilosis are recognized as commensal yeasts on the skin of healthy dogs but also causative agents of eborrheic dermatitis, especially in atopic dogs. We determined and compared the susceptibility levels of yeasts isolated from dogs with and without seborrheic dermatitis (SD) using the disk diffusion method (DD) for itraconazole (ITZ), ketoconazole (KTZ), nystatin (NYS), terbinafine (TERB) and 5-fluorocytosine (5-FC) and the broth microdilution method (BMD) for ITZ and KTZ. The reliability between the methods was assessed using an agreement analysis and linear regression. Forty-five M. pachydermatis and 28 C. parapsilosis isolates were identified based on physiological characteristics and an approved molecular analysis. By DD, all tested M. pachydermatis isolates were susceptible to ITZ, KTZ, NYS and TERB but resistant to 5-FC. Only 46 - 60% of the tested C. parapsilosis isolates were susceptible to KTZ, TERB and 5-FC, but ITZ and NYS were effective against all. By BMD, over 95% of M. pachydermatis isolates were susceptible to KTZ and ITZ with an MIC90 < 0.03 and 0.12 µg/ml, respectively. The frequency of KTZ- and ITZ-resistant C. parapsilosis was 29% and 7%, and the MIC90 values were 1 µg/ml and 0.5-1 µg/ml, respectively. Regarding the agreement analysis, 2.2% of minor errors were observed in M. pachydermatis and 0.2-1% of very major errors occurred among C. parapsilosis. There were no significant differences in the yeast resistance rates between dogs with and without SD. KTZ and ITZ were still efficacious for M. pachydermatis but a high rate of KTZ resistant was reported in C. parapsilosis.

Skin lipid profiling in normal and seborrhoeic shih tzu dogs.

BACKGROUND: Seborrhoea is a clinical condition resulting in excessive lipid and/or scale on the skin and is a common and important skin disease of dogs. However, there is little information on the skin surface lipid composition of dogs with seborrhoea. HYPOTHESIS/OBJECTIVES: To compare skin surface lipid profiles in normal and seborrhoeic shih tzu dogs. METHODS: Fourteen client-owned dogs (seven seborrhoeic and seven normal) were investigated. Lipids in sebaceous glands (SGs) were extracted from homogenized tissues of SG hyperplasia. Surface lipid was collected by tape stripping [stratum corneum (SC)-enriched fraction] and acetone-wetted cotton swab (acetone-extracted fraction). Lipids in SGs, SC-enriched fractions and acetone-extracted fractions were evaluated by high-performance thin-layer chromatography. RESULTS: Lipids in SGs mainly consisted of cholesterol esters, wax esters and triglycerides, whereas lipids in the SC-enriched fraction mainly consisted of ceramides. The acetone-extracted fraction contained a mixture of lipid classes recognized in SG- and SC-enriched fractions. In seborrhoeic dogs, concentrations of wax esters and triglycerides in the acetone-extracted fraction were significantly higher than in control dogs (P = 0.0285). Amounts of total ceramides (in micrograms) per milligram of SC were not significantly different between the two groups (P = 0.5204). Interestingly, two unknown ceramide fractions, which accounted for 20% of the total ceramides, were recognized exclusively in seborrhoeic dogs. CONCLUSIONS AND CLINICAL IMPORTANCE: These results provide evidence that the skin surface lipid profiles are altered in shih tzu dogs with seborrhoea.

Investigation of the aetiology of udder-thigh dermatitis in French dairy cattle.

Udder-thigh dermatitis (UTD) is a common disease in dairy cattle. The aim of this study was to define UTD incidence, its risk factors and the involved pathogens. Of 74 respondents, 72 (97%) reported having had the disease. On those farms the incidence was 5.3 cases per 100 cow years. The odds ratio (OR) of UTD in primiparous compared to multiparous cows was 23.4 (95% CI 17.3-33.8). Compared to tied stalls, the ORs of UTD were 0.65 (95% CI 0.45-0.92) and 0.43 (95% CI 0.24-0.71) for free stalls and straw yards, respectively. Udder oedema was reported in 98.3% of cows with UTD. The most common bacteria isolated from affected skin were Fusobacterium spp. (12/14 cases). This study suggests that UTD management should focus on local treatment, reducing udder oedema and increasing exercise.

Putative Malassezia dermatitis in six goats.

Histopathology submissions from 28 goats with dermatological disease were identified in an archival search of pathology files. Microscopic sections of skin biopsy specimens were examined for the presence of Malassezia spp. organisms. Six cases with many Malassezia yeasts were identified histopathologically. Based on the extent of clinical disease, three cases were regarded as localized and three were generalized infections. Clinical findings included alopecia with dry seborrhoea (four cases), greasy seborrhoea (one case), and no clinical findings specific to localized Malassezia infection when concurrent bacterial infection was present (one case). Mild pruritus was reported in two cases of generalized infection. No breed predilection was apparent. Three cases were male and three were female. Malassezia dermatitis occurred in goats from 10 months to 13 years of age. Three of six cases had concurrent bacterial infection. Skin lesions resolved following topical antifungal therapy in the two goats that were treated. Histopathological findings in all cases were severe follicular and epidermal orthokeratotic hyperkeratosis with minimal epithelial change and mild superficial perivascular to interstitial nonsuppurative inflammation. Numerous budding yeasts were visible within the stratum corneum of all cases; however, Malassezia was not isolated in the three cases in which culture was attempted. Based upon these findings, the authors suggest that the diagnosis Malassezia dermatitis in goats is most likely to be made by cytological examination of skin impressions or by examination of skin biopsy samples.

Comparative analysis of the frequency, distribution and population sizes of yeasts associated with canine seborrheic dermatitis and healthy skin.

The purpose of this study was to investigate the diversity of yeast associated with the degree of canine seborrheic dermatitis (SD) by anatomical sites. Fifty-seven samples were divided as 17 healthy skin, 20 with primary seborrheic dermatitis (PSD), and 20 with secondary seborrheic dermatitis (SSD). Yeast isolation and characterization were carried out based on microscopical features and biochemical properties. DNA analysis at the internal transcribed spacer I of 26S rDNA region was utilized for species confirmation. Four species of yeast consisting Malassezia pachydermatis, Malassezia furfur, Candida parapsilosis and Candida tropicalis recovered from examined dogs. M. pachydermatis and C. parapsilosis were isolated from all dogs, but C. tropicalis and M. furfur were recovered from 3 healthy dogs and one diseased dog, respectively. The number of M. pachydermatis and C. parapsilosis in diseased dogs was higher than that of healthy specimens (P<0.01). High frequency and population size of C. parapsilosis were closely associated to PSD, while those of M. pachydermatis were associated with both PSD and SSD (P<0.01). C. parapsilosis were predominant at the perianal area. This study demonstrated the co-colonization of M. pachydermatis and C. parapsilosis in large amounts and frequency associated with stage of disease and anatomical site.

Malassezia pachydermatis and M nana predominate amongst the cutaneous mycobiota of Sphynx cats.

Carriage of Malassezia species yeasts in healthy Sphynx cats was compared with that in Devon Rex cats (DRC), Cornish Rex cats (CRC) and domestic shorthair (DSH) cats. Swab samples from the external ear, anus and claw folds, and contact plate samples from the axillae and groins, were incubated on modified Dixon's agar at 32°C for 7 days. Malassezia species were isolated from all 18 Sphynx cats; M pachydermatis accounted for 118/140 isolates. Of 20 isolates of M nana, 16 were recovered from the ear canal. M slooffiae was isolated from the claw fold of one cat and the left groin of another. The high counts of M pachydermatis obtained from the axillae, groins and claw folds of the Sphynx cats exceeded those of healthy DSH, CRC and DRC; axillary populations were comparable to those of seborrhoeic DRC. These data support recent reports of high Malassezia species colonisation in Sphynx cats.

Localized seborrhoeic dermatitis with hyperhidrosis due to mite infestation in an Iranian cross-breed ram.

A 3-years-old Iranian cross-breed ram with history of repeated local sweating, severe pruritus of body surface was referred to the veterinary clinic. On clinical examination wetness, warmness, pruritus and thickness of affected area were observed. In affected area, hair coat was staring and draggy. Body temperature, heart and respiratory rates were 40.4 degrees C, 120 beat min(-1) and 40 min(-1), respectively. Hematologic indices including packed cell volume, total and differential white blood cell (WBC) and total red blood cell (RBC) were normal. Laboratory examinations of skin scrapings confirmed infestation with Psoroptes ovis. Histopathologic findings included dilation of sweat glands, hyperplasia of sebaceous glands, hyperkeratosis, ulcer and scab formation and eosinophilic dermatitis. History and clinical findings association with the skin scraping and histopathologic findings indicated localized seborrhoeic dermatitis with hyperhidrosis. After treatment with ivermectin at the dose rate of 0.2 mg kg(-1), all clinical signs subsided. This confirmed that the cause of seborrhic dermatitis and hyperhidrosis was mite infestation and other possible causes were ruled out. So this is the first report of localized seborrhoeic dermatitis with hyperhidrosis due to mite infestation in animals.

Cutaneous carriage of Malassezia species in healthy and seborrhoeic Sphynx cats and a comparison to carriage in Devon Rex cats.

Cutaneous carriage of Malassezia species yeast was investigated in 32 Sphynx cats, and in 10 domestic shorthair (DSH) cats. Samples for mycological culture were taken using contact plates and swabs at seven sites in each cat (left and right axillae and groin, left ear, claw fold on left front paw and the interdigital palmar web of the left front paw). Malassezia species were isolated from 26/32 Sphynx cats (81%) and from 0/10 DSH control cats. In five cases Malassezia species yeasts were isolated at a single site, in the remaining 21 Sphynx cats at multiple sites. A total of 73 Malassezia species isolates were made, of which 68 were Malassezia pachydermatis and five were lipid-dependent Malassezia. Five out of the 32 Sphynx had greasy seborrhoea, and all seborrhoeic cats had M pachydermatis isolated from their skin, at multiple sites. None of the 32 Sphynx had Malassezia species isolated from the ears. The difference in population sizes between Sphynx and DSH cats was significant (P

Carriage of Malassezia spp. yeasts in Cornish Rex, Devon Rex and Domestic short-haired cats: a cross-sectional survey.

Carriage of Malassezia spp. yeasts in healthy Cornish Rex cats (CRC) was compared with that in Devon Rex (DRC) and Domestic short-haired (DSH) cats. Samples obtained from the left external ear canal, anus and claw fold of digit III of the left fore foot by swabbing, and the axilla and groin using contact plates, were incubated for yeasts on modified Dixon's agar at 32 degrees C for 7 days. Malassezia species were isolated from 90% of the DRC, but from only 39% of the CRC and 50% of the DSH cats. M. pachydermatis accounted for 121 of 141 Malassezia spp. isolates. Five CRC were colonized by M. pachydermatis alone, one CRC yielded only M. nana, and one cat yielded only M. slooffiae, whereas five CRC were colonized by both M. pachydermatis and M. nana and another yielded M. pachydermatis, M. slooffiae and M. nana. M. nana was primarily isolated from the ear canal, whereas M. slooffiae was most often isolated from the claw. Both the frequencies of isolation and the population sizes of M. pachydermatis at all sites sampled in the CRC were comparable to those of 10 healthy DSH cats. Populations of M. pachydermatis in the left axilla and left and right groin in the CRC were significantly lower when compared with counts in a group of 21 healthy DRC, a breed with very similar coat characteristics but prone to seborrheic dermatitis caused by M. pachydermatis.

Carriage of Malassezia spp. yeasts in healthy and seborrhoeic Devon Rex cats.

Skin and anal mucosal carriage of Malassezia spp. yeasts was investigated in 21 healthy Devon Rex cats (DRC) and in 9 seborrhoeic DRC using swabs and contact plates. M. pachydermatis was isolated from 26 cats and lipid-dependent Malassezia spp. isolates were recovered from the claw fold of 5 healthy and 3 seborrhoeic DRC. The frequencies of isolation and population sizes of M. pachydermatis in the axillae, left groin and claw fold in seborrhoeic DRC significantly exceeded (P<0.05) those of healthy animals. The frequencies of isolation and population sizes of M. pachydermatis in the axillae and groin in both groups of DRC, and the frequencies of isolation and population sizes of M. pachydermatis in the claw fold of the seborrhoeic DRC, exceeded those of healthy Domestic short-haired cats. Using polymerase chain reaction--restriction enzyme analyses (PCR-REA) based on amplification of the large subunit rRNA gene, all eight lipid-dependent isolates had profiles that were indistinguishable from that of M. slooffiae CBS 7956. These data indicate that DRC are frequently colonized by M. pachydermatis and that the claw folds may also be colonized by M. slooffiae. The pathogenic significance of the high Malassezia spp. counts in the seborrhoeic DRC should now be determined.

Treatment of Malassezia pachydermatis-associated seborrhoeic dermatitis in Devon Rex cats with itraconazole--a pilot study.

Treatment of Malassezia pachydermatis-associated seborrhoeic dermatitis with oral itraconazole was investigated in six Devon Rex cats (DRC). The cutaneous populations of Malassezia were determined using contact plates and a swab-wash method before and after 21 days of pulse treatment with itraconazole (5 mg kg-1 once daily, 7 days on, 7 days off, 7 days on). Before treatment, all cats had greasy seborrhoeic dermatitis involving the axillae, groin, claw folds and palmar and plantar interdigital skin, and two had similar lesions on the ventral neck. After treatment, there was a significant (P<0.05) reduction in overall clinical scores and in scores at all individual sites assessed, except for the interdigital skin (P=0.068). Population sizes of M. pachydermatis in the left and right axillae, left and right groin and palmar interdigital skin were significantly (P<0.05) reduced, whereas the reduction in claw fold counts did not reach significance (P=0.068). The dramatic reduction in yeast counts and an associated marked clinical improvement of the seborrhoeic dermatitis provide important pilot data on the potential value of oral itraconazole in the management of seborrhoeic dermatitis associated with M. pachydermatis in DRC.

Dog factor differences in Can f 1 allergen production.

BACKGROUND: The clinical importance of dog allergy is well known, but it is unknown if all types of dogs represent the same risk for allergic patients. The purpose of this work was to evaluate among 288 healthy dogs if the levels of Can f 1 on fur vary between breeds (German Shepherd, Pyrenean Shepherd, Poodle, Cocker spaniel, Spaniel, Griffon, Labrador retriever and Yorkshire terrier), gender, hormonal status, hair length, and according to the presence of seborrhea. METHODS: Each dog was shaved in a limited area and Can f 1 concentrations were measured in mug/g fur by ELISA. The results (geometric mean values and 95% confidence intervals) were analyzed using analysis of variance and with nonparametric tests. RESULTS: A wide variability in Can f 1 levels was found between dog breeds, from Labradors [1.99 (0.03-129.91)] to Yorkshires [16.72 (3.67-76.16)] and Poodles [17.04 (2.79-103.94)] but only the Labrador levels were significantly different from each other breed. Males produced more Can f 1 than females, 11.75 (1.27-108.40) vs 8.89 (0.91-86.39). No difference was found according to hair length or hormonal status. The seborrheic status highly (P = 0.0019) influenced the presence of Can f 1 on hair: 16.66 (1.59-173.96) vs 9.40 (1.03-85.70). CONCLUSION: Breeds (Labrador retriever), sex and seborrhea seem to influence the levels of Can f 1 on fur.

Seborrhoeic dermatitis in a goat due to Malassezia pachydermatis.

A 6-year-old female goat was presented with a seborrhoeic dermatosis of 5 months duration. The condition developed following a severe enteritis associated with weight loss. Dermatological examination showed a generalized greasy seborrhoeic dermatosis, which spared the head and extremities of the limbs. Microscopic examination of impression smears of lesional skin revealed numerous yeasts typical of Malassezia sp. Culture on Sabouraud's dextrose agar yielded Malassezia pachydermatis growth. Histopathological examination of haematoxylin/eosin and safranin (HES) stained sections of biopsies showed mild lymphocytic superficial perivascular hyperplastic dermatitis. Numerous budding yeasts were visible both on the surface, and follicular keratin, in HES and periodic acid Schiff (PAS) stained sections. A dramatic response was observed after 1 week of a topical anti-Malassezia treatment, and the resolution of the condition was complete after 4 weeks.

Psoriasiform-lichenoid-like dermatosis in three dogs treated with microemulsified cyclosporine A.

Cyclosporine has been reported to be effective for the treatment of various cutaneous autoimmune disorders in dogs. Adverse reactions have generally been limited to gastrointestinal tract disturbances and cutaneous eruptions. The article describes antimicrobial-responsive cutaneous reactions in 3 dogs being treated with microemulsified cyclosporine A because of various dermatologic conditions. Cutaneous reactions in these dogs were similar to psoriasiform-lichenoid dermatitis and may represent an atypical staphylococcal infection.

Humoral and cell-mediated responses to Malassezia pachydermatis in healthy dogs and dogs with Malassezia dermatitis.

The in vitro proliferative responses of peripheral blood mononuclear cells from seven healthy basset hounds exposed to Malassezia pachydermatis antigen (500 micrograms/ml) exceeded (P < 0.05) those of seborrhoeic basset hounds with high populations of M pachydermatis and eight Irish setters with gluten-sensitive enteropathy. The stimulation indices in the latter two groups and in eight healthy beagles were comparable. The stimulation indices of the four groups after exposure to phytohaemaglutinin did not differ significantly. The serum titres of M pachydermatis-specific IgG and IgA measured by enzyme-linked immunosorbent assays (ELISA) in 21 seborrhoeic basset hounds and 11 affected dogs of various breeds exceeded those of 14 healthy basset hounds and eight healthy beagles (P < 0.01 for IgG, P < 0.05 for IgA). Total serum IgA concentrations measured by ELISA in the affected dogs were not lower than those of healthy dogs.