In vitro study of cyclosporine A 0.05 % on primary and recurrent pterygium fibroblasts.

To compare the cyclosporine 0.05 % exposure effect on fibroblasts from primary and recurrent pterygium. Primary culture of fibroblasts from primary and recurrent pterygium was performed until the third passage, which was exposed to cyclosporine 0.05 % in a group and the other remaining unexposed (control group), in triplicates. After 3, 6, 12, and 17 days of exposure the viable cell counting was performed by hemocytometer. The results were statistically analyzed using the technique of analysis of non-parametric variance model for repeated measures with three factors. There was a significant reduction in both fibroblast proliferation, in primary as in the recurrent pterygium cultures exposed to cyclosporine when compared not exposed cultures, with statistical significance (P < 0.05). Comparing primary and recurrent pterygium that received the drug, there was no significant difference in cell proliferation in relation to primary or recurrent pterygium. Cyclosporine 0.05 % is effective in inhibiting fibroblast proliferation in culture, both in primary and as in recurrent pterygium.

Pesquisa de Salmonella e das condições sanitárias em frangos e lingüiças comercializados na cidade de Botucatu / Research of Salmonella and sanitary conditions in poultry and sausages retailed in Botucatu

O objetivo do trabalho foi avaliar as condições sanitárias de frango e diversos tipos de lingüiças comercializados na cidade de Botucatu,São Paulo, pela determinação do número mais provável de coliformes a 45ºC/g além da pesquisa de Salmonella pela metodologia tradicional e pela PCR. Foram coletadas 50 amostras de carcaça de frango e 75 de lingüiças frescais, procedentes de nove estabelecimentos diferentes da cidade, no período de abril a novembro de 2006. Das 50 amostras de frango, 35 (70%) estavam fora dos parâmetros microbiológicos, segundo a RDC n°12 da Anvisa (>104 coliformes a 45ºC/g). Embora nessa Resolução, a pesquisa de Salmonella não seja exigida, 4 amostras(8%) apresentaram o patógeno pela metodologia tradicional. Essa presença foi confirmada pela PCR, que também foi positiva para mais 23 amostras, num total de 27 positivas (54%). Dentre as 75 amostras de lingüiças, 30 (40%) estavam fora dos limites permitidos, com 7 amostras positivas para Salmonella, pela metodologia tradicional. Entretanto, se considerar-se a pesquisa pela PCR, o número de amostras positivas aumenta para 42 (56%). Somando-se a taxa de freqüência de Salmonella aos limites microbiológicos para coliformes a 45ºC, 86,7% das lingüiças analisadas estavam impróprias para o Consumo.

In the present investigation were evaluated the sanitary conditions of poultry and several types of sausages retailed in Botucatu, São Paulo, Brazil for the determination of the most probable number of coliforms at 45ºC/g besides the research of Salmonella using traditional methodology and PCR. In order to do so, 50 samples of poultry and 75 of sausages were collected from nine different establishments in the city, in the period of April to November of 2006. Of the 50 samples of chicken meat, 35 (70%) were out of the microbiologic parameters, according to Brazilian Sanitary Resolution RDC n°12 of Anvisa (>104 coliforms at 45ºC/g). In this Resolution, the research of Salmonella is not demanded, but 4 samples (8%) presented the pathogen using the traditional methodology. That presence was confirmed by PCR, which was also positive in another 23, in a total of 27 positive samples (54%). Among 75 samples of sausages, 30 (40%)were out of the allowed limits, with 7 positive samples for Salmonella,using traditional methodology. However, if we consider PCR test, the number of positive samples increases to 42 (56%). Adding this number to coliforms microbiological limits, 86.7% of the analyzed sausages were inappropriate for the consumption.

Laryngeal papillomatosis: morphological study by light and electron microscopy of the HPV-6.

UNLABELLED: Laryngeal papillomatosis is the most frequent benign neoplasia in children. It is caused by HPV 6 and 11. The lesions are exophytic and highly recurrent, compromising the airway mucosa, mainly the larynx. Study design--clinical prospective. AIMS: To show morphologic alterations of the epithelium (light and electron microscopy) in the HPV-6 lesions. METHODS: Specimens of laryngeal lesions obtained during surgery of four children (1 male, 3 female) were submitted to HPV typing (PCR), light microscopy and electron microscopy. RESULTS: In all specimens, HPV type 6 was found. Epithelial projections were found by electron microscopy with superficial cells in desquamation. Light microscopy showed exophytic projections of the keratinized stratified squamous epithelium overlying a fibrovascular core. Koilocytes (vacuolated cells), suggesting the viral infection by HPV, were identified. No alterations were seen in the basement membrane and corion. Ultraestrutural analysis showed vacuolated cells with clear cytoplasmic inclusions, intercellular injuries and widening intercellular spaces. CONCLUSIONS: morphologic alterations of the epithelium in the HPV-6 lesions are superficial, and additional studies including the others HPV types are needed to show the more aggressive and extensive aspect of the disease.

Papilomatose laríngea: análise morfológica pela microscopia de luz e eletrônica do HPV-6 / Laryngeal papillomatosis: morphological study by light and electron microscopy of the HPV-6

Papilomatose laríngea é neoplasia benigna mais freqüente nas crianças, causada pelo HPV, principalmente subtipos 6 e 11 e caracteriza-se pela presença de lesões proliferativas exofíticas e recidivantes sobre a mucosa das vias aérea, em especial na laringe. Forma de Estudo: Clínico prospectivo. OBJETIVOS: Demonstrar alterações epiteliais morfológicas (pela microscopia de luz e eletrônica) em lesões papilíferas casadas pelo HPV-6. MATERIAL E MÉTODOS: Fragmentos de lesões de papilomatose laríngea, colhidos durante procedimento cirúrgico de quatro crianças (1 masculino, 3 femininas), foram submetidos à tipagem do HPV (por método de PCR), análise pela microscopia de luz e microscopia eletrônica (varredura e transmissão). RESULTADOS: Na tipagem, todos os papilomas eram do subtipo 6. A microscopia de varredura identificou projeções epiteliais de vários tamanhos, com células superficiais em descamação. A microscopia de luz demonstrou lesões exofíticas, revestidas por epitélio hiperplásico com coilócitos e binucleações, característicos do HPV. A membrana basal e o córion adjacente estavam íntegros. À microscopia eletrônica de transmissão identificou-se vacuolização perinuclear e alargamento das junções intercelulares. CONCLUSÕES: As alterações morfológicas apresentadas pelo HPV-6 demonstram o caráter não-invasivo da lesão, sendo necessário estudos morfológicos adicionais relacionando os outros tipos de HPV, considerados mais agressivos, com os achados ultra-estruturais.

Laryngeal papillomatosis is the most frequent benign neoplasia in children. It is caused by HPV 6 and 11. The lesions are exophytic and highly recurrent, compromising the airway mucosa, mainly the larynx. Study design - clinical prospective. AIMS: to show morphologic alterations of the epithelium (light and electron microscopy) in the HPV-6 lesions. METHODS: specimens of laryngeal lesions obtained during surgery of four children (1 male, 3 female) were submitted to HPV typing (PCR), light microscopy and electron microscopy. RESULTS: in all specimens, HPV type 6 was found. Epithelial projections were found by electron microscopy with superficial cells in desquamation. Light microscopy showed exophytic projections of the keratinized stratified squamous epithelium overlying a fibrovascular core. Koilocytes (vacuolated cells), suggesting the viral infection by HPV, were identified. No alterations were seen in the basement membrane and corion. Ultraestrutural analysis showed vacuolated cells with clear cytoplasmic inclusions, intercellular injuries and widening intercellular spaces. CONCLUSIONS: morphologic alterations of the epithelium in the HPV-6 lesions are superficial, and additional studies including the others HPV types are needed to show the more aggressive and extensive aspect of the disease.

Dimorphism, thermal tolerance, virulence and heat shock protein 70 transcription in different isolates of Paracoccidioides brasiliensis.

The mycelia-to-yeast (M-Y) transition, thermal tolerance and virulence profiles were evaluated for nine isolates of Paracoccidioides brasiliensis, including samples from two of the three recently discovered cryptic species, as well as their relation to the partial sequence and transcription of the hsp70 gene. The isolates Bt84 and T10 (from PS2 species) took more time to convert to yeast form and presented elongated yeast cells at 36 degrees C. Arthroconidia production was also observed during the M-Y transition for some isolates. Our data confirm that the hsp70 transcription may be associated with thermal tolerance, but this does not seem to be directly related to high virulence profiles. The partial sequencing of this gene allowed the separation of our isolates into two clusters that correspond to the two sympatric cryptic species occurring in an area hyperendemic for PCM (Botucatu, SP, Brazil).

Detecção e genotipagem de papilomavírus humano em lesões de queratoacantoma solitário de pacientes imunocompetentes / Detection and genotyping of human papillomavirus in solitary keratoacanthoma lesions of immunocompetent patients

FUNDAMENTOS: O queratoacantoma é neoplasia cutânea benigna que incide preferencialmente em indivíduos de pele clara, faixa etária elevada, acometendo áreas fotoexpostas. Além da exposição à radiação ultravioleta, sua etiologia é relacionada a diversos carcinógenos, entre eles a infecção pelo papilomavírus humano (HPV). OBJETIVOS: Avaliar a prevalência do DNA do HPV, bem como seus genótipos, em lesões de queratoacantoma solitário de pacientes imunocompetentes. MÉTODOS: Foram estudados queratoacantomas de pacientes sem evidências de imunocomprometimento, excisados entre 1996 e 2000 em hospital universitário. Realizaram-se cortes histológicos, desparafinização e extração de DNA desses fragmentos. Os espécimes positivos para DNA de HPV foram submetidos ao seqüenciamento gênico, para determinação do genótipo. RESULTADOS: Foram estudados 58 pacientes com idade média de 64,5±13,8 anos. A proporção entre os sexos foi semelhante, e as localizações mais comuns foram os membros superiores (50 por cento) e a face (27,6 por cento). Detectou-se DNA de HPV em 48 (82,7 por cento) fragmentos de queratoacantomas, sendo os genótipos 6, 11 e 16 os prevalentes. CONCLUSÕES: A alta prevalência do achado de DNA de HPV em lesões de queratoacantoma solitário pode sugerir a participação viral em sua oncogênese.

BACKGROUND - Keratoacanthoma is a benign cutaneous neoplasm that preferentially affects fair skin individuals of older age groups and involves sun-exposed areas. Apart from ultraviolet radiation exposure, its etiology is related to several carcinogens, including human papillomavirus (HPV) infection. OBJECTIVES: To assess the prevalence of HPV DNA and its genotypes in solitary keratoacanthoma lesions of immunocompetent patients. Methods: Keratoacanthoma lesions of patients with no evidence of immunological involvement, excised from 1996 to 2000, at a university hospital, were assessed. Histological aspects, deparafinization and DNA extraction of these lesions were evaluated. The specimens positive for HPV DNA were submitted to gene sequencing to determine the genotype. RESULTS: Fifty-eight patients were studied, mean age of 64.5±13.8 years. Both sexes were similarly affected, and the most common sites were upper limbs (50 percent) and face (27.6 percent). HPV DNA was detected in 48 (82.7 percent) keratoacanthoma fragments, and the genotypes 6, 11 and 16 were the most prevalent. CONCLUSIONS: The high prevalence of HPV DNA in solitary keratoacanthoma lesions may suggest viral participation in its oncogenesis.

Lack of papillomavirus (HPV) in pterygia of a Brazilian sample.

PURPOSE: Because the influence of papillomavirus (HPV) in pterygium pathogenesis is controversial, the aim of this study was to identify whether it is present in the pterygia lesions in our region. METHODS: Thirty-six patients with unilateral primary pterygia were submitted to excision of pterygia and a sample of normal conjunctiva. Tissues were submitted to polymerase chain reaction (PCR) evaluation for papillomavirus DNA detection. RESULTS: We were unable to detect any HPV DNA in all studied specimens. CONCLUSION: According to our results papillomarivus is not important for pterygium formation.

Lack of papillomavirus (HPV) in pterygia of a Brazilian sample / Ausência de papilomavirus (HPV) em pterígios de uma amostra brasileira

PURPOSE: Because the influence of papillomavirus (HPV) in pterygium pathogenesis is controversial, the aim of this study was to identify whether it is present in the pterygia lesions in our region. METHODS: Thirty-six patients with unilateral primary pterygia were submitted to excision of pterygia and a sample of normal conjunctiva. Tissues were submitted to polymerase chain reaction (PCR) evaluation for papillomavirus DNA detection. RESULTS: We were unable to detect any HPV DNA in all studied specimens. CONCLUSION: According to our results papillomarivus is not important for pterygium formation.

OBJETIVO: Existem controvérsias a respeito da influência do papilomavirus (HPV) no desenvolvimento do pterígio. Assim, este estudo foi elaborado com o objetivo de verificar se o papilomavirus está presente na lesão. MÉTODOS: Trinta e seis portadores de pterígio unilateral foram operados, preparando-se o tecido removido e uma amostra de conjuntiva normal para exame de reação em cadeia da polimerase (PCR) para detecção de DNA. RESULTADOS: Em todas as amostras do pterígio e da conjuntiva normal a pesquisa do DNA-papilomavirus por PCR resultou negativa. CONCLUSÃO: Segundo nossos resultados, o papilomavirus não é importante para o desenvolvimento do pterígio.

Human pythiosis, Brazil.

Pythiosis, caused by Pythium insidiosum, occurs in humans and animals and is acquired from aquatic environments that harbor the emerging pathogen. Diagnosis is difficult because clinical and histopathologic features are not pathognomonic. We report the first human case of pythiosis from Brazil, diagnosed by using culture and rDNA sequencing.

Molecular detection of Paracoccidioides brasiliensis in soil.

The objective of the present study was to evaluate different techniques for the detection of Paracoccidioides brasiliensis in soil, e.g., culture, animal inoculation and specific DNA amplification by Nested PCR. We designed species-specific inner primers derived from rDNA regions (ITS, 5.8S gene) and found their sensitivity to be higher than culture and animal inoculation. In addition, the sensitivity of these primers was higher than p27-gene primers developed for detection of P. brasiliensis in soil in a previous study. DNA from P. brasiliensis was detected in soil artificially seeded with the fungus (positive soil control) and from environmental samples collected in an armadillo burrow.

Human respiratory syncytial virus detection in children admitted at a community hospital in Botucatu, SP, Brazil

O Vírus Respiratório Sincicial Humano (VRSH) é descrito como o mais importante patógeno viral causador de doenças respiratórias agudas das vias respiratórias inferiores em crianças. Neste estudo 84 amostras de crianças com idade abaixo dos dois anos apresentando sintomas de doença respiratória aguda, foram obtidas no período de setembro de 2000 a novembro de 2001. Analise por imunofluorescência indireta e transcrição reversa seguida de PCR, revelou que 18 per center (15/84) das amostras foram positivas, sendo que em 80 per center (12/15) dos casos a detecção de VRSH foi observada em crianças abaixo dos seis meses, e também que os subgrupos A e B co-circularam. Estes são os primeiros dados obtidos para a cidade de Botucatu, sendo que a sazonalidade mostrou-se evidente pela maior circulação desse vírus entre os meses de maio e julho.