Systemic phaeohyphomycosis in a dog caused by Cladophialophora bantiana

Background: Cladophialophora bantiana is a dematiaceous fungus that causes phaeohyphomycosis, a generic term used to describe a variety of unusual mycoses caused by fungi that have melanin in their cell wall. C. bantiana targets the central nervous system, commonly causing localized brain infections that may result in disseminated infections. In Brazil, minimal phaeohyphomycosis data are available, and information about C. bantiana infections in animals, especially canines, is scarce. Thus, the aim of this study was to describe the clinical and pathological aspects of systemic phaeohyphomycosis caused by C. bantiana in a dog. Case: A 1-year-old female Pit Bull presented with weight loss, reduced appetite, and a history of cutaneous lesions on the right thoracic limb; however, clinical evolution was not reported. The bitch had reportedly given birth recently. Physical examination revealed thinness, pale ocular and oral mucosa, submandibular lymph nodes, and enlarged popliteal lymph nodes. The bitch died after convulsive crises during hospitalization. At necropsy, white-yellowish multifocal nodules were observed in the liver and right kidney. The brain featured left cerebral hemisphere asymmetry with blood vessel congestion in the leptomeninges and an irregular brownish focal area on the surface of the right occipital cortex. Cross-sections of the formalin-fi xed brain exhibited compression of the left lateral ventricle and the presence of grayish and friable multifocal areas in the gray matter of the left parietal and right occipital cortices. Fragments of the lesions were collected for histopathological and microbiological examination. Histologically, the lesions were similar, characterized by hepatitis, nephritis, and granulomatous and necrotizing...

Dermatite seborreica em anta albina (Tapirus terrestris) / Seborrheic dermatitis in albino lowland tapir (Tapirus terrestris) / Dermatitis seborreica en tapir albino (Tapirus terrestris)

O albinismo é uma desordem genética recessiva em que a produção de melanina é reduzida ou ausente, o que interfere na pigmentação da pele, pelos e olhos dos indivíduos. É considerada de rara ocorrência natural, principalmente em animais selvagens. Indivíduos albinos são altamente suscetíveis a injúrias na pele causadas pelo sol, já que a exposição aos raios ultravioletas (UV) de forma cumulativa pode ser prejudicial. Este relato discorre sobre a ocorrência de seborreia seca em uma anta albina (Tapirus terrestris), mantida sob cuidados humanos. A seborreia seca é uma enfermidade cutânea de caráter crônico em que ocorre falha na queratinização da epiderme, formando excessivamente escamas e alterando a oleosidade da pele e pelos. O tratamento clínico instituído foi baseado na aplicação tópica de solução dermatológica hidratante spray e em mudanças realizadas no recinto a fim de diminuir a incidência de luz solar no indivíduo. O resultado foi satisfatório e teve como auxílio um microscópio digital portátil que foi utilizado periodicamente até a recuperação do animal.(AU)

Albinism is a recessive genetic disorder in which melanin prodution is reduced or absent, interfering with pigmentation of the skin, hair and eyes of individuals. It is considered to be of rare natural occurrence, mainly in wild animals. Albino individuals are highly susceptible to skin damage caused by the sun, once cumulative exposure to ultraviolet (UV) rays can be harmful. This report discusses the occurrence of dry seborrhea in an albino lowland tapir (Tapirus terrestris), kept under human care. Dry seborrhea is a chronic skin disease in which there is a failure in the keratinization of the epidermis, forming excessive scales and altering the oiliness of the skin and hair. The clinical treatment instituted was based on topical application of a moisturizing dermatological spray solution and on changes made to the enclosure in order to reduce the incidence of sunlight on the individual. The result was satisfactory and had as na aid a portable digital microscope that was used periodically until the recovery of the animal.(AU)

El albinismo es una anomalía genética recesiva en la que la produccíon de la melanina está reducida o ausente, lo que interfiere com la pigmentación de la piel, pelaje y los ojos de los individuos. Se considera un fenómeno natural raro, principalmente en animales salvajes. Los albinos son muy susceptibles al daño de la piel por el sol, ya que la exposición acumulada a los rayos ultravioleta (UV) puede ser dañina. Este reporte discute la ocurrencia de seborrea seca en un tapir albino (Tapirus terrestris), mantenida bajo cuidado humano. La seborrea seca es una enfermedad crónica de la piel en la que falla la queratinización de la epidermis, formándose descamaciones excesivas y alterando la oleosidad de la piel y la pelaje. El tratamiento clínico instituido se basó en la aplicación tópica de uma solución dermatológica hidratante en spray y em la modificación del recinto para reducir la incidencia de la luz solar sobre el individuo. El resultado fue satisfactorio y tuvo como ayuda un miscroscopio digital portátil que se utilizó periódicamente hasta la recuperación del animal.(AU)

Melanose talâmica em caprinos / Thalamic melanosis in goats

Background: Melanosis is a blackened pigmentation resulting from the accumulation of melanocytes in tissues that are not normally pigmented. This change in the color of the organs occurs due to the agglomeration of melanocytes originating from abnormal migration during embryogenesis and does not cause dysfunction to the affected organ. Although melanosis frequently occurs in several species and affects several organs such as the brain and spinal cord leptomeninges, involvement in the thalamus region is unusual. The objective of this work was to report 2 cases of thalamic melanosis in goats, determining the pathological and histochemical aspects that assist in the diagnosis of this condition. Cases: Two cases of thalamic melanosis in goats were diagnosed. In both cases, the animals had no nervous history disease and clinical signs. The cause of death in cases 1 and 2 was established based on anatomopathological findings and clinical signs being diagnosed with mycoplasmosis and asphyxia, respectively. After fixing and making cross-sections of the brain, a focal lengthy blackened area was observed on the thalamus surface in both cases. Microscopically, lesions in the brain were similar in both cases and exclusively affected the thalamus. These cells had abundant cytoplasm, well delimited with brownish granular pigment. The nuclei were difficult to visualize and in some cells, it was rounded, well-defined, morphologically compatible with melanocytes. Melanocytes were mainly distributed around neurons and often distended the perivascular space of multiple blood vessels. In Fontana Masson staining, the granules in the cytoplasm of these cells stained strongly black. The Prussian Blue, Periodic Acid- Schiff's, Von Kossa, and Giemsa stains were negative, and the pigment remained brown. In the unstained slides, assembled after the deparaffinization and clarification process, it was observed the permanence of cells with blackish-brown pigment in the cytoplasm. In immunohistochemistry, strong immunostaining of pigmented cells with the Anti-MelanA antibodies was observed in both cases. Discussion: The diagnosis of thalamic melanosis in goats was carried out based on the characteristic pathological findings, in which melanin pigments were demonstrated and identified through HE, Fontana-Masson staining, and unstained slides and confirmed by the IHC. The use of complementary histochemical techniques was fundamental for the classification of the pigment as melanin, demonstrating to be an accessible and reliable tool for the diagnosis of pathological processes that lead to the accumulation of pigments and or material in the tissues. The occurrence of melanin in the thalamus may be associated with a failure in the migration of melanoblasts, which would go to the optical pathways or to the thalamus. This erratic migration of melanoblasts can be explained by the fact that the forebrain is the embryogenic origin of the optic and diencephalon pathways. Macroscopically, thalamic melanosis must be differentiated mainly from neoplastic processes such as melanoma and hemangiosarcoma, pigmented fungus infections, Phalaris angusta poisoning, listeriosis, neurocutaneous melanosis, and neuromelanin. It was concluded that thalamic melanosis is an uncommon alteration in goats and although it has been diagnosed as an incidental necropsy finding, should be included in the differential diagnosis of diseases that affect the central nervous system, especially those that have a color change associated with the deposition of pigments in the tissues.

Use of Giemsa staining for the immunohistochemical counterstaining in canine melanomas: an "old and forgotten" method

Melanoma is a neoplasm originating from melanocytes and represents 7% of skin tumors and the most common in oral cavity of dogs. Melanoma may present melanocytic pigment in its cytoplasm in varying quantity and its characterization by immunohistochemistry (IHC) is challenging due to the use of chromogen diaminobenzidine (DAB) which itself produces a brown product what makes difficult to distinguish from melanin pigment in this technique. To demonstrate a reliable technique, the use of the Giemsa counterstaining was performed in the IHC of melanomas with different degrees of pigmentation for different cytoplasmic, membrane and nuclear markers. The modification in the IHC technique by the counterstaining of Giemsa allows observable differences under the microscope between melanic pigment(in a blue-green stain), while the DAB chromogen will be observed in brown. With this technique, the prognostic and predictive interpretation of markers in canine melanomas may be more reliable in definitions of clinical behaviors and inexperimental analyzes.(AU)

Use of Giemsa staining for the immunohistochemical counterstaining in canine melanomas: an "old and forgotten" method

Melanoma is a neoplasm originating from melanocytes and represents 7% of skin tumors and the most common in oral cavity of dogs. Melanoma may present melanocytic pigment in its cytoplasm in varying quantity and its characterization by immunohistochemistry (IHC) is challenging due to the use of chromogen diaminobenzidine (DAB) which itself produces a brown product what makes difficult to distinguish from melanin pigment in this technique. To demonstrate a reliable technique, the use of the Giemsa counterstaining was performed in the IHC of melanomas with different degrees of pigmentation for different cytoplasmic, membrane and nuclear markers. The modification in the IHC technique by the counterstaining of Giemsa allows observable differences under the microscope between melanic pigment(in a blue-green stain), while the DAB chromogen will be observed in brown. With this technique, the prognostic and predictive interpretation of markers in canine melanomas may be more reliable in definitions of clinical behaviors and inexperimental analyzes.

Regression of dark color in subterranean fishes involves multiple mechanisms: response to hormones and neurotransmitters

Organisms with source-populations restricted to the subterranean biotope (troglobites) are excellent models for comparative evolutionary studies, due to their specialization to permanent absence of light. Eye and dark pigment regression are characteristics of most troglobites. In spite of the advance in knowledge on the mechanisms behind eye regression in cave fishes, very little is known about pigmentation changes. Studies were focused on three species of the genus Pimelodella. Exemplars of the troglobitic P. spelaea and P. kronei were compared with the epigean (surface) P. transitoria, putative sister-species of the latter. Melanophore areas and densities are significantly lower in the troglobitic species. Evaluating the in vitro response of these cells to adrenaline, acetylcholine and MCH, we observed a reduced response in both troglobites to adrenaline. The same trend was observed with MCH, but not statistically significant. No response to acetilcholine was detected in all the three. Contrary to expectations, even though eye-regression in P. spelaea was much lower than in P. kronei, pigmentation regression was more advanced. Multiple mechanisms of loss showing a mosaic of traits in troglobitic fishes are discussed here.(AU)

Organismos com populações-fonte restritas ao biótopo subterrâneo (troglóbios) são excelentes modelos para estudos evolutivos comparativos, devido à especialização resultante do isolamento sob um regime seletivo particular, com ênfase na permanente falta de luz. A regressão do olho e da pigmentação são características dos troglobites. Apesar do avanço do conhecimento sobre os mecanismos subjacentes à regressão ocular em peixes de caverna, pouco se sabe sobre mudanças de pigmentação. Os estudos foram focados em três espécies do gênero Pimelodella. Exemplares das espécies troglóbias P. spelaea e P. kronei foram comparados com a epígea P. transitoria, provável espécie-irmã dessa última. As áreas e densidades dos melanóforos são significativamente menores nas espécies troglóbias. Avaliando a resposta in vitro dessas células à adrenalina, acetilcolina e MCH, observamos uma resposta reduzida em ambos os troglóbios à adrenalina. A mesma tendência foi observado com o MCH, mas não estatisticamente. Nenhuma resposta à acetilcolina foi detectada três. Contrariamente às expectativas, embora a regressão ocular em P. spelaea seja bem menor do que em P. kronei, a regressão na pigmentação foi mais acentuada. Múltiplos mecanismos de regressão, mostrando um mosaico de características em peixes troglóbios, são discutidos aqui.(AU)

Regression of dark color in subterranean fishes involves multiple mechanisms: response to hormones and neurotransmitters

ABSTRACT Organisms with source-populations restricted to the subterranean biotope (troglobites) are excellent models for comparative evolutionary studies, due to their specialization to permanent absence of light. Eye and dark pigment regression are characteristics of most troglobites. In spite of the advance in knowledge on the mechanisms behind eye regression in cave fishes, very little is known about pigmentation changes. Studies were focused on three species of the genus Pimelodella. Exemplars of the troglobitic P. spelaea and P. kronei were compared with the epigean (surface) P. transitoria, putative sister-species of the latter. Melanophore areas and densities are significantly lower in the troglobitic species. Evaluating the in vitro response of these cells to adrenaline, acetylcholine and MCH, we observed a reduced response in both troglobites to adrenaline. The same trend was observed with MCH, but not statistically significant. No response to acetilcholine was detected in all the three. Contrary to expectations, even though eye-regression in P. spelaea was much lower than in P. kronei, pigmentation regression was more advanced. Multiple mechanisms of loss showing a mosaic of traits in troglobitic fishes are discussed here.

RESUMO Organismos com populações-fonte restritas ao biótopo subterrâneo (troglóbios) são excelentes modelos para estudos evolutivos comparativos, devido à especialização resultante do isolamento sob um regime seletivo particular, com ênfase na permanente falta de luz. A regressão do olho e da pigmentação são características dos troglobites. Apesar do avanço do conhecimento sobre os mecanismos subjacentes à regressão ocular em peixes de caverna, pouco se sabe sobre mudanças de pigmentação. Os estudos foram focados em três espécies do gênero Pimelodella. Exemplares das espécies troglóbias P. spelaea e P. kronei foram comparados com a epígea P. transitoria, provável espécie-irmã dessa última. As áreas e densidades dos melanóforos são significativamente menores nas espécies troglóbias. Avaliando a resposta in vitro dessas células à adrenalina, acetilcolina e MCH, observamos uma resposta reduzida em ambos os troglóbios à adrenalina. A mesma tendência foi observado com o MCH, mas não estatisticamente. Nenhuma resposta à acetilcolina foi detectada três. Contrariamente às expectativas, embora a regressão ocular em P. spelaea seja bem menor do que em P. kronei, a regressão na pigmentação foi mais acentuada. Múltiplos mecanismos de regressão, mostrando um mosaico de características em peixes troglóbios, são discutidos aqui.

Identification and characterisation of alternative splice variants of HOXB9 and their correlation with melanogenesis in the black-boned chicken

Homeobox B9 (HOXB9) gene has been demonstrated to be associated with melanogenesis in chicken plumage by high-throughput RNA sequencing. In this study, we cloned and characterised HOXB9 in black-boned chickens. Two alternative splice variants (HOXB9-1 and HOXB9-2) were identified in chicken feather bulbs. Expression analysis of HOXB9 in 11 different chicken tissues by RT-PCR indicated that the two transcripts were only expressed in the kidney, abdominal fat, feather bulbs, skin, and small intestine. No HOXB9-1 or HOXB9-2 transcripts were detected in the breast muscle or the ovary. The two HOXB9 variants were expressed at significantly different levels in black feather bulbs and white feather bulbs (p<0.01), and in black skin compared with white skin (p<0.01). The results indicate that HOXB9-1 and HOXB9-2 may be involved in melanin formation in the plumage and skin. The expression of HOXB9-2 was higher in white than in black muscles (p<0.05), HOXB9-2 may play a role in muscle melanogenesis. Our results suggest that HOXB9, which is partially regulated by alternative splicing, may be involved in the process of melanogenesis in the black-boned chicken.(AU)

Identification and characterisation of alternative splice variants of HOXB9 and their correlation with melanogenesis in the black-boned chicken

Homeobox B9 (HOXB9) gene has been demonstrated to be associated with melanogenesis in chicken plumage by high-throughput RNA sequencing. In this study, we cloned and characterised HOXB9 in black-boned chickens. Two alternative splice variants (HOXB9-1 and HOXB9-2) were identified in chicken feather bulbs. Expression analysis of HOXB9 in 11 different chicken tissues by RT-PCR indicated that the two transcripts were only expressed in the kidney, abdominal fat, feather bulbs, skin, and small intestine. No HOXB9-1 or HOXB9-2 transcripts were detected in the breast muscle or the ovary. The two HOXB9 variants were expressed at significantly different levels in black feather bulbs and white feather bulbs (p<0.01), and in black skin compared with white skin (p<0.01). The results indicate that HOXB9-1 and HOXB9-2 may be involved in melanin formation in the plumage and skin. The expression of HOXB9-2 was higher in white than in black muscles (p<0.05), HOXB9-2 may play a role in muscle melanogenesis. Our results suggest that HOXB9, which is partially regulated by alternative splicing, may be involved in the process of melanogenesis in the black-boned chicken.

Melanose multicêntrica em frango de corte: relato de caso / Multicentric melanosis in broiler: case report

Melanoblastos podem migrar de forma errática durante a embriogênese, gerando um quadro conhecido como melanose. São raros os estudos envolvendo melanose com acometimento múltiplo dos órgãos. Objetivou-se descrever um caso de melanose multicêntrica em frango de corte que gerou condenação do animal ao abate. Foram encaminhadas ao Laboratório de Patologia Animal da Universidade Federal do Espírito Santo para avaliação histopatológica amostras de diversos órgãos de uma ave de corte da linhagem Cobb de 48-49 dias de idade. Esse animal foi condenado na linha de inspeção por apresentar áreas multifocais enegrecidas. Ao exame microscópico, observou-se melanina multifocal variando de moderada a intensa quantidade em todos os órgãos acometidos com lesões enegrecidas na macroscopia. As áreas pigmentadas foram negativas para a coloração especial de azul da Prússia e não foram encontradas células neoplásicas. A pigmentação da pele em aves comercializadas no Ocidente é rara, pois a característica de pele clara foi selecionada por meio de melhoramento genético, mas pouco se sabe sobre quais mutações desencadeiam melanose nas linhagens selecionadas para não apresentar pigmentação. A ave apresentou melanose multicêntrica e tal alteração não compromete a função dos órgãos acometidos nem representa risco para a saúde humana, no entanto, devido ao aspecto macroscópico, os órgãos que apresentam tal lesão foram condenados para consumo humano.(AU)

Melanoblasts can enter an erratic migratory pathway during embryogenesis and it creates a clinical condition known as melanosis. Studies involving melanosis in multiple organs are rare. The aim of this paper is to describe a case of multicentric melanosis in a broiler resulting in animal condemnation at slaughter. Samples from diverse organs originating from a Cobb broiler, 48-49 days of age, were sent to Laboratório de Patologia Animal from Universidade Federal do Espírito Santo for histopathological avaliation. This animal was condemned in line inspection due to multifocal black areas of pigmentation. At microcopic avaliation, multifocal melanin in moderate-intense amount in all the organs stricken with macroscopic black lesions was noted. Sections of pigmented areas were negative for Prussian Blue and no neoplasic cells were found. Pigmentation of the skin in a broiler from the Ocident is rare because the white/yellow skin was selected through genetic enhancement, but there is some data about which mutations unleash melanosis in lineages selected to not have pigmentation. The broiler had multicentric melanosis and this alteration doesn't change functional activity of the stricken organs and doesn't represent any risk for human health, but the organs with this lesion were condemned for human consumption due to their macroscopic appearance.(AU)

Melanose multicêntrica em frango de corte: relato de caso / Multicentric melanosis in broiler: case report

Melanoblastos podem migrar de forma errática durante a embriogênese, gerando um quadro conhecido como melanose. São raros os estudos envolvendo melanose com acometimento múltiplo dos órgãos. Objetivou-se descrever um caso de melanose multicêntrica em frango de corte que gerou condenação do animal ao abate. Foram encaminhadas ao Laboratório de Patologia Animal da Universidade Federal do Espírito Santo para avaliação histopatológica amostras de diversos órgãos de uma ave de corte da linhagem Cobb de 48-49 dias de idade. Esse animal foi condenado na linha de inspeção por apresentar áreas multifocais enegrecidas. Ao exame microscópico, observou-se melanina multifocal variando de moderada a intensa quantidade em todos os órgãos acometidos com lesões enegrecidas na macroscopia. As áreas pigmentadas foram negativas para a coloração especial de azul da Prússia e não foram encontradas células neoplásicas. A pigmentação da pele em aves comercializadas no Ocidente é rara, pois a característica de pele clara foi selecionada por meio de melhoramento genético, mas pouco se sabe sobre quais mutações desencadeiam melanose nas linhagens selecionadas para não apresentar pigmentação. A ave apresentou melanose multicêntrica e tal alteração não compromete a função dos órgãos acometidos nem representa risco para a saúde humana, no entanto, devido ao aspecto macroscópico, os órgãos que apresentam tal lesão foram condenados para consumo humano.(AU)

Melanoblasts can enter an erratic migratory pathway during embryogenesis and it creates a clinical condition known as melanosis. Studies involving melanosis in multiple organs are rare. The aim of this paper is to describe a case of multicentric melanosis in a broiler resulting in animal condemnation at slaughter. Samples from diverse organs originating from a Cobb broiler, 48-49 days of age, were sent to Laboratório de Patologia Animal from Universidade Federal do Espírito Santo for histopathological avaliation. This animal was condemned in line inspection due to multifocal black areas of pigmentation. At microcopic avaliation, multifocal melanin in moderate-intense amount in all the organs stricken with macroscopic black lesions was noted. Sections of pigmented areas were negative for Prussian Blue and no neoplasic cells were found. Pigmentation of the skin in a broiler from the Ocident is rare because the white/yellow skin was selected through genetic enhancement, but there is some data about which mutations unleash melanosis in lineages selected to not have pigmentation. The broiler had multicentric melanosis and this alteration doesn't change functional activity of the stricken organs and doesn't represent any risk for human health, but the organs with this lesion were condemned for human consumption due to their macroscopic appearance.(AU)

Análise histoquímica da língua de morcegos frugívoros neotropicais (Mammalia - chiroptera) com relato de pigmentação epitelial / Histochemical analysis of the tongue in neotropical frugivorous bats (Mammalia - chiroptera) with report of epithelial pigmentation

The tongue is regarded as a key organ for bats, since the specialized characters are used for taxonomic and systematic purposes, cladistic studies, and elucidation of patterns of eating habits. This study aimed to characterize morphologically the tongue in the species Artibeus lituratus, Artibeus planirostris, Artibeus obscurus, and Dermanura cinerea, and to report the first evidence of melanin in the lingual epithelium of bats. We used 20 specimens collected in the municipalities of Vitória de Santo Antão and Sirinhaém, in Pernambuco State, Brazil. The tongues were extracted and submitted to routine histological techniques, stained with hematoxylin and eosin (HE) and Fontana-Masson. In the four species, the keratinized stratified pavement epithelium was observed, but with the presence of melanin granules on the basal layer only in the species A. lituratus, A. planirostris, and A. obscurus, evidenced through the Fontana-Masson staining technique. The melanin found in the tongue of these species is characterized as the first record for the order Chiroptera and it may be related to oral asepsis, and/or inhibition of toxic substances observed in plants. Therefore, there is a need for further studies, aiming to improve our knowledge of the function of pigment in this organ.

A língua é considerada um órgão-chave para os morcegos, pois os caracteres especializados são utilizados para fins taxonômicos e sistemáticos, estudos cladísticos e elucidação dos padrões dos hábitos alimentares. Objetivou-se caracterizar morfologicamente a língua das espécies Artibeus lituratus, Artibeus planirostris, Artibeus obscurus e Dermanura cinerea e relatar a primeira evidência de melanina no epitélio lingual de morcegos. Foram utilizados 20 exemplares coletados nos municípios de Vitória de Santo Antão e Sirinhaém, em Pernambuco. As línguas foram extraídas e submetidas a técnicas histológicas de rotina, coradas com hematoxilina e eosina (HE) e Fontana-Masson. Nas quatro espécies, foi observado o epitélio estratificado pavimentoso queratinizado, porém com presença de grânulos de melanina na camada basal somente nas espécies A. lituratus, A. planirostris e A. obscurus, evidenciados por meio da técnica de coloração por Fontana-Masson. A melanina encontrada na língua dessas espécies caracteriza-se como o primeiro registro para a ordem Chiroptera e pode estar relacionada à assepsia oral e/ou à inibição de substâncias tóxicas presentes em vegetais. Portanto, há necessidade de novos estudos, visando aprimorar o conhecimento acerca da função do pigmento nesse órgão.

Análise histoquímica da língua de morcegos frugívoros neotropicais (Mammalia - chiroptera) com relato de pigmentação epitelial / Histochemical analysis of the tongue in neotropical frugivorous bats (Mammalia - chiroptera) with report of epithelial pigmentation

The tongue is regarded as a key organ for bats, since the specialized characters are used for taxonomic and systematic purposes, cladistic studies, and elucidation of patterns of eating habits. This study aimed to characterize morphologically the tongue in the species Artibeus lituratus, Artibeus planirostris, Artibeus obscurus, and Dermanura cinerea, and to report the first evidence of melanin in the lingual epithelium of bats. We used 20 specimens collected in the municipalities of Vitória de Santo Antão and Sirinhaém, in Pernambuco State, Brazil. The tongues were extracted and submitted to routine histological techniques, stained with hematoxylin and eosin (HE) and Fontana-Masson. In the four species, the keratinized stratified pavement epithelium was observed, but with the presence of melanin granules on the basal layer only in the species A. lituratus, A. planirostris, and A. obscurus, evidenced through the Fontana-Masson staining technique. The melanin found in the tongue of these species is characterized as the first record for the order Chiroptera and it may be related to oral asepsis, and/or inhibition of toxic substances observed in plants. Therefore, there is a need for further studies, aiming to improve our knowledge of the function of pigment in this organ.(AU)

A língua é considerada um órgão-chave para os morcegos, pois os caracteres especializados são utilizados para fins taxonômicos e sistemáticos, estudos cladísticos e elucidação dos padrões dos hábitos alimentares. Objetivou-se caracterizar morfologicamente a língua das espécies Artibeus lituratus, Artibeus planirostris, Artibeus obscurus e Dermanura cinerea e relatar a primeira evidência de melanina no epitélio lingual de morcegos. Foram utilizados 20 exemplares coletados nos municípios de Vitória de Santo Antão e Sirinhaém, em Pernambuco. As línguas foram extraídas e submetidas a técnicas histológicas de rotina, coradas com hematoxilina e eosina (HE) e Fontana-Masson. Nas quatro espécies, foi observado o epitélio estratificado pavimentoso queratinizado, porém com presença de grânulos de melanina na camada basal somente nas espécies A. lituratus, A. planirostris e A. obscurus, evidenciados por meio da técnica de coloração por Fontana-Masson. A melanina encontrada na língua dessas espécies caracteriza-se como o primeiro registro para a ordem Chiroptera e pode estar relacionada à assepsia oral e/ou à inibição de substâncias tóxicas presentes em vegetais. Portanto, há necessidade de novos estudos, visando aprimorar o conhecimento acerca da função do pigmento nesse órgão.(AU)

ANÁLISE HISTOQUÍMICA DA LÍNGUA DE MORCEGOS FRUGÍVOROS NEOTROPICAIS (MAMMALIA - CHIROPTERA) COM RELATO DE PIGMENTAÇÃO EPITELIAL

Abstract The tongue is regarded as a key organ for bats, since the specialized characters are used for taxonomic and systematic purposes, cladistic studies, and elucidation of patterns of eating habits. This study aimed to characterize morphologically the tongue in the species Artibeus lituratus, Artibeus planirostris, Artibeus obscurus, and Dermanura cinerea, and to report the first evidence of melanin in the lingual epithelium of bats. We used 20 specimens collected in the municipalities of Vitória de Santo Antão and Sirinhaém, in Pernambuco State, Brazil. The tongues were extracted and submitted to routine histological techniques, stained with hematoxylin and eosin (HE) and Fontana-Masson. In the four species, the keratinized stratified pavement epithelium was observed, but with the presence of melanin granules on the basal layer only in the species A. lituratus, A. planirostris, and A. obscurus, evidenced through the Fontana-Masson staining technique. The melanin found in the tongue of these species is characterized as the first record for the order Chiroptera and it may be related to oral asepsis, and/or inhibition of toxic substances observed in plants. Therefore, there is a need for further studies, aiming to improve our knowledge of the function of pigment in this organ.

Resumo A língua é considerada um órgão-chave para os morcegos, pois os caracteres especializados são utilizados para fins taxonômicos e sistemáticos, estudos cladísticos e elucidação dos padrões dos hábitos alimentares. Objetivou-se caracterizar morfologicamente a língua das espécies Artibeus lituratus, Artibeus planirostris, Artibeus obscurus e Dermanura cinerea e relatar a primeira evidência de melanina no epitélio lingual de morcegos. Foram utilizados 20 exemplares coletados nos municípios de Vitória de Santo Antão e Sirinhaém, em Pernambuco. As línguas foram extraídas e submetidas a técnicas histológicas de rotina, coradas com hematoxilina e eosina (HE) e Fontana-Masson. Nas quatro espécies, foi observado o epitélio estratificado pavimentoso queratinizado, porém com presença de grânulos de melanina na camada basal somente nas espécies A. lituratus, A. planirostris e A. obscurus, evidenciados por meio da técnica de coloração por Fontana-Masson. A melanina encontrada na língua dessas espécies caracteriza-se como o primeiro registro para a ordem Chiroptera e pode estar relacionada à assepsia oral e/ou à inibição de substâncias tóxicas presentes em vegetais. Portanto, há necessidade de novos estudos, visando aprimorar o conhecimento acerca da função do pigmento nesse órgão.

MELANOGÊNESE E SENSIBILIDADE DE CEPAS DO COMPLEXO DE ESPÉCIES Cryptococcus neoformans/Cryptococcus gattii IN VITRO: UMA MODULACÃO COM INIBIDORES DA BOMBA DE PRÓTONS, CATECOLAMINAS E PARAQUAT

Cryptococcus spp. são agentes etiológicos da criptococose. Para estabelecer a infecção, essas leveduras expressam fatores de virulência como a melanina e o biofilme. Esta tese foi dividida em três objetivos: 1. Avaliar os efeitos dos inibidores da bomba de prótons (IBPs) sobre o crescimento e a produção de melanina de Cryptococcus spp.; 2. Investigar a produção de melanina pelo complexo de espécies C. neoformans/C. gattii na presença de diferentes precursores da melanogênese, avaliando o efeito da melanização na sensibilidade antifúngica; 3. Avaliar a atividade de paraquat (PQ) contra Cryptococcus spp. nas formas planctônica e biofilme, bem como o efeito protetor de agentes antioxidantes sobre essa atividade. Buscou-se avaliar ainda a cinética da produção de melanina diante o PQ. As concentrações inibitórias mínimas (CIMs) de IBPs foram determinadas por microdiluição. O efeito dos IBPs na produção de melanina foi avaliado na presença de sulfato de cobre ou glutationa. Foram utilizados como substratos para a melanogênese, a epinefrina, norepinefrina, dopamina, ácido cafeico e L-dopa, sendo em seguida avaliada a sensibilidade in vitro. A atividade da anfotericina B em cepas melanizadas foi investigada no ensaio de time kill. A sensibilidade ao PQ foi avaliada e, em seguida, as cepas expostas ao PQ foram cultivadas em meio com ou sem ácido ascórbico (AA) e glutationa (GSH). Ademais, a produção de melanina foi avaliada na presença de L-dopa + PQ. Os IBPs mostraram CIMs variando de 125-1000 g/mL e redução da melanização em células de Cryptococcus. O sulfato de cobre e a glutationa restauraram a melanogênese das cepas na presença de IBPs. Adicionalmente, a presença de IBPs diminuiu a toxicidade do sulfato de cobre (1 mM). As cepas produziram melanina, em escalas diferenciada, de acordo com o substrato testado. As células melanizadas foram mais tolerantes à anfotericina B. No tocante ao efeito do PQ, observou-se CIM variando de 8-256 g/mL e uma inibição na formação do biofilme. O efeito inibitório planctônico foi restaurado por AA e GSH. Além disso, L-dopa + PQ alterou a cinética da produção de melanina. Em conclusão, observou-se que IBPs inibem o crescimento planctônico e a melanogênese em Cryptococcus spp., sendo este último efeito dependente da quelação do cobre ou inibição do transporte de cobre ATPase. Ademais, os dados em conjunto, evidenciaram que a melanina contribui para a maior resiliência de Cryptococcus spp. a drogas antifúngicas, como a anfotericina B, e agentes agressores ambientais, como o paraquat.

Cryptococcus spp. are etiological agents of cryptococcosis. To establish infection, these yeasts produce virulence factors such as melanin and biofilm. This thesis was divided into three aims: 1. Evaluate the effects of proton pump inhibitors (PPIs) on growth and melanin production by Cryptococcus spp.; 2. Investigate the production of melanin by the C. neoformans/C. gattii species complex in the presence of different precursors of melanogenesis, evaluating the effect of melanization on antifungal susceptibility. 3. Evaluate the activity of paraquat (PQ) against Cryptococcus spp. in planktonic and biofilm forms, as well as the protective role of antioxidant agents against PQ. We also sought to evaluate the kinetics of melanin production in the presence of PQ. Minimum inhibitory concentrations (MICs) of PPIs were determined by microdilution. The effect of PPIs on melanin production was evaluated in the presence of copper sulfate or glutathione. As substrates for melanogenesis, epinephrine, norepinephrine, dopamine, caffeic acid and L-dopa were used, then, in vitro susceptibility was evaluated. Amphotericin B activity against melanized strains was investigated by the time kill assay. The susceptibility to PQ was evaluated and the strains exposed to the PQ were cultivated in medium with or without ascorbic acid (AA) and glutathione (GSH). Furthermore, melanin production was evaluated in the presence of L-dopa + PQ. PPIs showed MICs ranging from 125-1000 g/mL and reduced melanization of Cryptococcus cells. Copper sulfate and glutathione restored the melanogenesis of the strains in the presence of PPIs. Additionally, the presence of PPIs decreased copper sulphate toxicity (1 mM). The strains produced melanin, at different levels, according to the tested substrate. Melanized cells were more tolerant to amphotericin B. Regarding the effect of PQ, MICs ranged from 8-256 g/mL and biofilm formation was inhibited. The inhibitory effect of PQ was hampered by AA and GSH. Furthermore, L-dopa + PQ altered the kinetics of melanin production. In conclusion, it was observed that PPIs inhibit planktonic growth and melanogenesis of Cryptococcus spp., with the latter being dependent on copper chelation or inhibition of copper ATPase transport. Furthermore, altogether, the data showed that melanin contributes to reduced susceptibility of Cryptococcus spp. to the antifungal drug amphotericin B, and the environmental aggressor paraquat.

CARACTERIZAÇÃO E PRODUÇÃO DE MELANINA PELO FUNGO NEMATÓFAGO Duddingtonia flagrans E AVALIAÇÃO DA SUA PARTICIPAÇÃO NA ATIVIDADE PREDATÓRIA

Melaninas formam um grupo diverso de pigmentos sintetizados na maioria dos organismos por meio da hidroxilação e polimerização de compostos orgânicos. Nos fungos, a melanina exerce um importante papel na proteção contra as intempéries ambientais, a radiação ultravioleta, extremos de temperaturas, além de aumentar a virulência e a patogenicidade destes organismos. Assim, buscou-se descrever a síntese de melanina em Duddingtonia flagrans, fungo nematófago, e a relação da melanização sobre a cinética de predação do nematoide Panagrellus spp. Para tal foram testados diferentes meios de cultura na presença e na ausência de precursores L-DOPA, na adição de glicose e do inibidor tricyclazol. As análises foram realizadas por espectroscopia de infravermelho, ressonância paramagnética eletrônica (RPE) e microscopia eletrônica de transmissão. O melhor crescimento e a melanização do fungo ocorreram em meio Sabouraud dextrose quando comparado aos demais meios utilizados, após 21 dias incubados a 30°C. O acréscimo de glicose nas concentrações de 2% a 10% (v/w) não inibiu o processo de melanização em D. flagrans. Entretanto acréscimos de 2% de glicose em pH 5,0 observou-se um aumento na coloração em relação aos demais tratamentos. Nas concentrações de glicose acima de 10% ocorreu a inibição completa do crescimento. Como conclusão pode-se confirmar a presença do pigmento melanina e atribuir os valores obtidos por RPE (g=2.0051 0.0001) a produção de feomelanina pelo fungo. Ainda, pode-se verificar a deposição dos grânulos de melanina sobre a parede celular. A síntese da melanina ocorreu em melanossomos e sua gênese é análoga a de melanossomos de células de mamíferos. Contudo, a presença de melanina não influenciou de forma significativa a cinética de predação entre o fungo e nematoide.

Melanins form a diverse group of pigments synthesized in most organisms through the hydroxylation and polymerization of organic compounds.In the fungi, melanin has an important role in protecting against environmental inclemencies, ultraviolet radiation, temperature extremes, virulence and pathogenicity of these organisms. Thus, we attempted to describe the melanin synthesis in Duddingtonia flagrans, nematophagous fungus, and the relationship between melanization and predation kinetics of Panagrellus spp. nematode. For this, differents culture media were tested in presence and in absence of L-DOPA precursors, in addition of glucose and tricyclazole inhibitor. The analysis were realized by infrared spectroscopy, electronic paramagnetic resonance (EPR) and transmission electron microscopy. The better growth and melanization of the fungus occurred in Sabouraud dextrose medium when compared to other media used after 21 days incubated at 30°C. The addition of glucose at 2% to 10% (v/w) concentrations did not inhibit the melanization process in D. flagrans. However, the addition of glucose at 2% at pH 5.0, it was observed an increase staining in relation to others treatments. In the glucose concentrations above 10% occurred complete inhibition of growth. It was confirmed the presence of melanin pigment and to assign the values obtained by RPE (g=2.0051 0.0001) to production of pheomelanin by the fungus. And it was observed the deposition of the melanin granules on the cell wall. Melanin synthesis has occurred in melanosomes and its genesis is analogous to that of mammalian cell melanosomes. However, the presence of melanin did not significantly influence the kinetics of predation between the fungus and the nematode.

Ultrastructural study of the mycelial phase of clinical isolates of Sporothrix schenckii obtained from feline, canine and human cases of sporotrichosis

Using transmission electron microscopy, we studied the presence of melanin and cell wall thickness of clinical isolates of Sporothrix schenckii obtained from cats, dogs and humans as compared to reference strains. We detected differences regarding presence of the melanin among the clinical isolates of S. schenckii and a correlation between presence of melanin and cell wall thickness.

Clareamento da melanina na epiderme do Lobo-marinho-sul-americano e sua aplicação na imunohistoquímica enzimática / Bleaching of melanin in the epidermis of South American fur seal and its application on enzyme immunohistochemistry

O Lobo-marinho-sul-americano (Arctocephalus australis) é um mamífero marinho anfíbio distribuído ao longo da Costa do Atlântico e do Pacífico da América do Sul. Esta espécie está bem adaptada a diferentes habitats devido à morfologia dos membros em forma de nadadeira e de seu sistema tegumentar. Estudos imuno-histoquímicos são importantes para avaliar os mecanismos de adaptação da pele devido a diferencial expressão dos antígenos presentes no tecido dependendo da região da superfície corporal. Entretanto, sua epiderme altamente pigmentada (melanina) impede a visualização dos marcadores cromógenos utilizados na imunohistoquímica. Neste trabalho foi desenvolvido um método de clarear a melanina para permitir a visualização dos cromógenos sem alterar a afinidade antígeno-anticorpo para a imuno-histoquímica. A análise do índice do PCNA (proliferating cell nuclear antigen) na epiderme de A. australis, com diaminobenzidina (DAB) como cromógeno foi usada para testar o método. O clareamento da melanina permitiu obter o índice de proliferação celular na epiderme e evitar resultados falso-positivos sem afetar os resultados imuno-histoquímicos.(AU)

The South American fur seal (Arctocephalus australis) is an amphibious marine mammal distributed along the Atlantic and Pacific coasts of South America. The species is well adjusted to different habitats due to the morphology of its fin-like members and due to some adaptations in their integumentary system. Immunohistochemical studies are very important to evaluate the mechanisms of skin adaptation due the differential expression of the antigens present in the tissue depending of the region of the body surface. However, its strongly pigmented (melanin) epidermis prevents the visualization of the immunohistochemical chromogens markers. In this study a melanin bleaching method was developed aimed to allow the visualization of the chromogens without interfering in the antigen-antibody affinity for immunohistochemistry. The analysis of PCNA (proliferating cell nuclear antigen) index in the epidermis of A. australis by immunohistochemistry with diaminobenzidine (DAB) as chromogen was used to test the method. The bleaching of the melanin allowed to obtain the cell proliferation index in epidermis and to avoid false positive results without affecting the immunohistochemical results.(AU)

Displasias foliculares em cães: relato de dois casos na cidade de João Pessoa - PB / Follicular dysplasia in dogs: report of two cases in the city of João Pessoa - PB

Um dos principais motivos pelos quais cães e gatos são levados ao consultório ou clínica veterinária é a percepçãopor parte dos proprietários da alopecia. Diversas dermatopatias podem levar a este sintoma, em variados graus,e, acompanhados ou não de outros tipos de lesões. O objetivo deste trabalho é relatar um caso clínico de alopeciapor diluição da cor (ADC) e um caso de displasia folicular dos pelos pretos (DFPP), duas doenças consideradasraras. Os cães foram atendidos na Clínica Veterinária Animais em Pessoa, na cidade de João Pessoa PB (AU)

One of the main reasons why dogs and cats are taken to the clinic or veterinary clinic is the perceptionby the owners of alopecia. Various skin diseases can lead to this symptom, in varying degrees, and withor without other lesions. The objective of this study is to report a clinical case of color dilution alopecia(CDA) and a case of black hair follicular dysplasia (BHFD), both considered rare diseases. The dogs weretreated at Animais em Pessoa Veterinary Clinic in the city of João Pessoa - PB. (AU)

Displasias foliculares em cães: relato de dois casos na cidade de João Pessoa - PB / Follicular dysplasia in dogs: report of two cases in the city of João Pessoa - PB

Um dos principais motivos pelos quais cães e gatos são levados ao consultório ou clínica veterinária é a percepçãopor parte dos proprietários da alopecia. Diversas dermatopatias podem levar a este sintoma, em variados graus,e, acompanhados ou não de outros tipos de lesões. O objetivo deste trabalho é relatar um caso clínico de alopeciapor diluição da cor (ADC) e um caso de displasia folicular dos pelos pretos (DFPP), duas doenças consideradasraras. Os cães foram atendidos na Clínica Veterinária Animais em Pessoa, na cidade de João Pessoa – PB

One of the main reasons why dogs and cats are taken to the clinic or veterinary clinic is the perceptionby the owners of alopecia. Various skin diseases can lead to this symptom, in varying degrees, and withor without other lesions. The objective of this study is to report a clinical case of color dilution alopecia(CDA) and a case of black hair follicular dysplasia (BHFD), both considered rare diseases. The dogs weretreated at Animais em Pessoa Veterinary Clinic in the city of João Pessoa - PB.