Comparison of three protocols to preserve Leptospira spp. in cat urine for efficient DNA extraction and PCR amplification

Background: The pathogenic leptospira infection in mammalian species can cause a range of acute or chronic manifestations and may result in a carrier state. Previous studies have suggested that cats were resistant to acute leptospirosis however, the description of some clinical cases suggests that Leptospira spp. may also be pathogenic to this species. Recentstudies have shown that leptospires may be shed in the urine of infected cats. Endogenous substances present in urine mayinhibit PCR and allow leptospires to evade detection. This study aims to compare three protocols for sample processingto optimize the detection of pathogenic leptospires in cat urine.Materials, Methods & Results: Three protocols to optimize the detection of pathogenic leptospires in cat urine were tested.Aliquots of standard concentration of L. interrogans serovar Canicola culture were added to urine samples to achieveconcentrations of 1×105 to 1×102 leptospires/mL for each protocol. In protocols A and B the urine was neutralized by theaddition of phosphate-buffered saline (PBS), pH 7.4, in a proportion of 1 PBS: 2.5 urine (v/v). In protocol A, PBS wasadded to neutralize the urine pH for the leptospiral organisms immediately after addition of leptospires. In protocol B,PBS was added just before DNA extraction. In protocol C, no PBS was added. DNA extraction was performed at 4, 24and 48 h after addition of the leptospires using a modified protocol. Samples were incubated at 37ºC for 10 min. Sampleswere then centrifuged (850 g) for 15 min, at 25ºC. The supernatants were transferred to another tube, and the pellets werediscarded. The supernatants were centrifuged (16060 g) for 20 min at 4ºC. The supernatants were then discarded, and thepellets resuspended and washed with 1000 µL of PBS. All the samples were centrifuged at 16060 g for an additional 20min at 25ºC. The supernatants were discarded and the pellets were resuspended in 100 µL of PBS and incubated at 94ºCfor...(AU)

Comparison of three protocols to preserve Leptospira spp. in cat urine for efficient DNA extraction and PCR amplification

Background: The pathogenic leptospira infection in mammalian species can cause a range of acute or chronic manifestations and may result in a carrier state. Previous studies have suggested that cats were resistant to acute leptospirosis however, the description of some clinical cases suggests that Leptospira spp. may also be pathogenic to this species. Recentstudies have shown that leptospires may be shed in the urine of infected cats. Endogenous substances present in urine mayinhibit PCR and allow leptospires to evade detection. This study aims to compare three protocols for sample processingto optimize the detection of pathogenic leptospires in cat urine.Materials, Methods & Results: Three protocols to optimize the detection of pathogenic leptospires in cat urine were tested.Aliquots of standard concentration of L. interrogans serovar Canicola culture were added to urine samples to achieveconcentrations of 1×105 to 1×102 leptospires/mL for each protocol. In protocols A and B the urine was neutralized by theaddition of phosphate-buffered saline (PBS), pH 7.4, in a proportion of 1 PBS: 2.5 urine (v/v). In protocol A, PBS wasadded to neutralize the urine pH for the leptospiral organisms immediately after addition of leptospires. In protocol B,PBS was added just before DNA extraction. In protocol C, no PBS was added. DNA extraction was performed at 4, 24and 48 h after addition of the leptospires using a modified protocol. Samples were incubated at 37ºC for 10 min. Sampleswere then centrifuged (850 g) for 15 min, at 25ºC. The supernatants were transferred to another tube, and the pellets werediscarded. The supernatants were centrifuged (16060 g) for 20 min at 4ºC. The supernatants were then discarded, and thepellets resuspended and washed with 1000 µL of PBS. All the samples were centrifuged at 16060 g for an additional 20min at 25ºC. The supernatants were discarded and the pellets were resuspended in 100 µL of PBS and incubated at 94ºCfor...

Demodiciose felina - revisão de literatura / Feline demodicosis review

A demodiciose felina é uma dermatopatia parasitária rara, causada principalmente por dois ácaros, oDemodex cati e o Demodex gatoi. O D. cati é comensal da pele dos gatos, causa lesões alopécicas não pruriginosase, eventualmente, generalizadas, e pode ser associado a doenças imunodepressoras como a FIV eFeLV. O D. gatoi, por outro lado, é infeccioso, causa lesões pruriginosas que podem mimetizar dermatopatiasalérgicas ou psicogênicas e acomete gatos anteriormente saudáveis. (AU)

The feline demodicosis is a rare parasitic skin disease, mainly caused by two mites, Demodex cati and Demodexgatoi. D. cati is found on healthy hair follicles but it may cause generalized non pruritic alopecic lesions on hostsand it is associated with immunosuppressive diseases like FIV and FeLV, meanwhile. D. gatoi is infectious, causes pruritic lesions that may mimic allergic or psychogenic injuries and may affect otherwise healthy cats. (AU)

Demodiciose felina - revisão de literatura / Feline demodicosis – review

A demodiciose felina é uma dermatopatia parasitária rara, causada principalmente por dois ácaros, oDemodex cati e o Demodex gatoi. O D. cati é comensal da pele dos gatos, causa lesões alopécicas não pruriginosase, eventualmente, generalizadas, e pode ser associado a doenças imunodepressoras como a FIV eFeLV. O D. gatoi, por outro lado, é infeccioso, causa lesões pruriginosas que podem mimetizar dermatopatiasalérgicas ou psicogênicas e acomete gatos anteriormente saudáveis.

The feline demodicosis is a rare parasitic skin disease, mainly caused by two mites, Demodex cati and Demodexgatoi. D. cati is found on healthy hair follicles but it may cause generalized non pruritic alopecic lesions on hostsand it is associated with immunosuppressive diseases like FIV and FeLV, meanwhile. D. gatoi is infectious, causes pruritic lesions that may mimic allergic or psychogenic injuries and may affect otherwise healthy cats.

Pododermatite plasmocítica felina relato de caso / Feline plasma cellpododermatitis case report

A pododermatite plasmocítica felina é uma enfermidade incomum que acomete os coxins plantares e/ ou palmares. Sua patogenia é desconhecida, porém, acredita-se tratar de uma doença imunomediada. O presente trabalho relata o caso de um gato, fêmea, SRD, de cinco anos de idade, ovariohisterectomizada,com histórico de emagrecimento, claudicação e edemaciação dos coxins plantares e palmares, com duas semanas de evolução. De acordo com os sinais clínicos suspeitou-se de pododermatite plasmocítica felina, a qual foi confirmada com o exame histopatológico de um coxim afetado. O tratamento foi realizado com uso de glicocorticóide em dose imunossupressora associado à ciclosporina, mostrando-se eficaz. (AU)

Feline plasma cell pododermatitis is an uncommon disease that affects the paw pads. The pathogenesis is unknown, but is believed to treat a immunomediated disease. This study reports a cat, female,domestic shorthair, five years old, spayed, with a weight loss history, lameness and swelling in pawpads. Duration of lesions prior to consultation ranged from two weeks. According to clinical signs, wassuspected of feline plasma cell pododermatitis, which was confirmed by histopathology of a paw padaffected. The treatment was performed with use of immunosuppressive glucocorticoids and cyclosporine,proving to be effective. (AU)

Pododermatite plasmocítica felina – relato de caso / Feline plasma cellpododermatitis – case report

A pododermatite plasmocítica felina é uma enfermidade incomum que acomete os coxins plantares e/ ou palmares. Sua patogenia é desconhecida, porém, acredita-se tratar de uma doença imunomediada. O presente trabalho relata o caso de um gato, fêmea, SRD, de cinco anos de idade, ovariohisterectomizada,com histórico de emagrecimento, claudicação e edemaciação dos coxins plantares e palmares, com duas semanas de evolução. De acordo com os sinais clínicos suspeitou-se de pododermatite plasmocítica felina, a qual foi confirmada com o exame histopatológico de um coxim afetado. O tratamento foi realizado com uso de glicocorticóide em dose imunossupressora associado à ciclosporina, mostrando-se eficaz.

Feline plasma cell pododermatitis is an uncommon disease that affects the paw pads. The pathogenesis is unknown, but is believed to treat a immunomediated disease. This study reports a cat, female,domestic shorthair, five years old, spayed, with a weight loss history, lameness and swelling in pawpads. Duration of lesions prior to consultation ranged from two weeks. According to clinical signs, wassuspected of feline plasma cell pododermatitis, which was confirmed by histopathology of a paw padaffected. The treatment was performed with use of immunosuppressive glucocorticoids and cyclosporine,proving to be effective.