Amostragem sequencial e microssatelites na avaliação da qualidade genética em lotes de sementes de milho / Sequential sampling and microsatellites for evaluation of the genetic quality in corn seeds lots

Na produção de sementes, a pureza genética é um dos requisitos fundamentais para a comercialização. Assim, foi determinado o tamanho da amostra para a avaliação da pureza genética, avaliada a sensibilidade da técnica de microssatélites para discriminar híbridos de seus respectivos parentais e a sensibilidade em detectar misturas, quando presentes em pequenas proporções na amostra. Para a amostragem sequencial, foram tomados grupos de 40 sementes híbridas em sequência, até no máximo 400 sementes, as quais foram marcadas e misturadas ao lote de sementes, simulando contaminações de 0,25%; 0,5%; 1%; 2%; 4% e 6%. A sensibilidade da técnica de microssatélite foi avaliada misturando-se diferentes proporções de DNA dos híbridos com o de suas respectivas linhagens. Para mistura superior a 1:8 (1P1: 8P2; 8P1: 1P2), a sensibilidade do marcador variou em função do primer utilizado. Na amostragem sequencial, para detectar níveis de mistura acima de 1% no lote de sementes, com nível de risco tanto para o produtor quanto para o consumidor de 0,05, o tamanho de amostra necessário é inferior ao requerido pela amostra de tamanho fixo, o que possibilita a redução do custo e a viabilização da técnica para atestar a pureza genética de lotes de sementes de milho.

In the seed production system, genetic purity is one of the fundamental requirements for its commercialization. The present work had the goal of determined the sample size for genetic purity evaluation, in order to protect the seed consumer and the producer and to evaluate the sensitivity of microsatellite technique for discriminating hybrids from their respective relatives and for detecting mixtures when they are present in small amounts in the samples. For the sequential sampling, hybrid seeds were marked and mixed in with the seed lots, simulating the following levels of contamination: 0.25, 0.5, 1.0, 2.0, 4.0, and 6.0%. After this, groups of 40 seeds were taken in sequence, up to a maximum of 400 seeds, with the objective of determining the quantity of seeds necessary to detect the percentage of mixture mentioned above. The sensitivity of microsatellite technique was evaluated by mixing different proportions of DNA from the hybrids with their respective seed lines. For the level of mixture was higher than 1:8 (1P1:8P2; 8P1:1P2), the sensitivity of the marker in detecting different proportions of the mixture varied according to the primer used. In terms of the sequential sampling, it was verified that in order to detect mixture levels higher than 1% within the seed lot- with a risk level for both the producer and the consumer of 0.05- the size of the necessary sample was smaller than the size needed for the fixed sample size. This also made it possible to reduce costs, making it possible to use microsatellites to certify the genetic purity of corn seeds lots.

Prognostic Biomarkers and EBV Infection Research in Diffuse Large B-Cell Lymphoma of the Palatine Tonsils.

Diffuse large B-cell lymphoma represents approximately 30%-40% of all diagnoses of non-Hodgkin's Lymphoma and may represent up to 80% of all lymphomas that arise in the palatine tonsils. Several studies have attempted to correlate clinical, laboratorial, and tissue factors with the prognosis of the lymphomas, such as the International Prognostic Index, the tissue expression of some proteins, and the lymphocyte count at the time of diagnosis, as well as to correlate Epstein-Barr virus (EBV) infection with worse prognoses. Patients with palatine tonsil DLBCL, from Salvador, Bahia, Brazil, were studied in order to identify prognostic factors. Twenty-four patients with DLBCL were studied. The factors that negatively influenced the patients' survival rates were the lymphocyte count at the time of diagnosis <1.000/mm(3) and the Bcl-2 protein expression. There was no CD5 expression in these lymphomas, and neither was there an association with EBV infection.

Subcutaneous Panniculitis-Like T-Cell Lymphoma (SPTL) in a Child with Spontaneous Resolution.

Subcutaneous panniculitis-like T-cell lymphomas (SPTLs) α/ß are rare in childhood. The present report refers to a case of a 7-year-old male child presenting an extensive skin lesion that began when he was 5 years of age. Two biopsies were evaluated using the CD3, CD4, CD8, CD56, ßF1, and TIA markers. A dense infiltrate of CD3+, CD4-, CD8+, CD56-, ßF1+, and TIA+ pleomorphic lymphocytes was found in the subcutis. The previous biopsy showed cytophagic histiocytic panniculitis with a small focus on CD8+ and ßF1+ malignant cells. The lesion regressed spontaneously. This case shows that prognosis may be excellent in SPTL (α/ß). On the other hand, it also serves as an alert that a biopsy performed in an area of cytophagic panniculitis may lead to misdiagnosis.

Flower cells in patients with infective dermatitis associated with HTLV-1.

BACKGROUND: Infective dermatitis associated with HTLV-1 (IDH) is a severe childhood form of eczema that may progress to adult T-cell leukemia/lymphoma (ATL). OBJECTIVE: In this study, the presence of clinical and laboratory parameters suggestive of ATL was evaluated in a cohort of 30 patients with IDH. STUDY DESIGN: Over a period of 33 months, the patients were submitted to three-monthly clinical evaluations, routine laboratory exams, full blood count and blood smears, and to six-monthly blood sampling for HTLV-1 proviral load determination. HTLV-1 proviral load was quantified using real-time TaqMan PCR assay. RESULTS: Abnormal cells (Ably) were found in the peripheral blood smears of nine patients (30%), flower cells being detected in five of these cases (16.6%). The presence of Ably and flower cells was not associated with a higher proviral load in those patients. CONCLUSIONS: This is the first report on the presence of flower cells in HTLV-1-infected children and adolescents. Furthermore, these cells have not previously been reported in IDH patients. The cases with flower cells probably represent precursory ATL cases, these patients being at a greater risk of developing ATL.

Adult T-cell leukemia/lymphoma (ATL) presenting in the skin: clinical, histological and immunohistochemical features of 52 cases.

BACKGROUND: Adult T-cell leukemia/lymphoma (ATL) is a severe disease caused by HTLV-I. This paper describes the clinicopathological and immunohistochemical findings of 52 cases of ATL with skin involvement and investigates whether there is any relationship between median survival time (MST) and histological patterns, primary cutaneous involvement and CD8 positivity. MATERIAL AND METHODS: All cases were HTLV-I+ and HIV- and were clinically classified. HTLV-I proviral integration was investigated in atypical cases. Immunohistochemistry was performed using CD3, CD4, CD5, CD7, CD8, CD20, CD25, CD30 and CD45RO markers. Ki-67 was used to evaluate the proliferative index. RESULTS: Twenty-seven cases were primary, while 25 were secondary. Monoclonal viral integration was demonstrated in all atypical cases. Patterns resembling mycosis fungoides (MF) were found in 19 cases and anaplastic large-cell lymphoma (ALCL) in two cases. Fifteen cases had an atypical immunophenotype and expressed CD8. Primary cutaneous ATL had a longer MST (48 months) than the secondary cutaneous ATL (7 months) and the difference was statistically significant, but no statistically significant difference was found between the MST of CD8-positive and negative cases. CONCLUSIONS: It is important to differentiate between primary and secondary cutaneous ATL and classify the cases histologically in order to better evaluate the prognosis. The two forms of primary cutaneous ATL, primary cutaneous smoldering and primary cutaneous tumoral (PCT), should also be identified. The smoldering type presented a longer survival (58 months) and histological aspects suggestive of better prognosis in contrast to the PCT type that had a shorter survival (20 months) and histological characteristics suggestive of worse outcome.

Adult-onset infective dermatitis associated with HTLV-I. Clinical and immunopathological aspects of two cases.

Infective dermatitis associated with HTLV-I (IDH) is a chronic, infected childhood eczema. Two adult-onset cases of IDH were studied, one of which was associated with HAM/TSP. The patients were submitted to dermatological, neurological and pathological examination. Immunohistochemical studies were made using CD3, CD4, CD8, CD20, CD79a, and CD57 antibodies. Cytotoxic granules were investigated using granzyme B, perforin, and TIA. The patients presented infected erythematous, scaly lesions with mild itching and a good response to sulfamethoxazole/ trimethoprim. A differential diagnosis with atopic dermatitis (AD) and seborrheic dermatitis (SD) was made, based on: the distinctive morphology and distribution of the lesions, presence of exudative and infected lesions, and mild pruritus. The inflammatory infiltrate was composed predominantly of CD8+ lymphocytes that did not present cytotoxic granules. We concluded that IDH can begin in adulthood and may be associated with HAM/TSP. The immunohistochemical findings were different from those observed in AD and SD.

Histopathological and immunohistochemical studies of infective dermatitis associated with HTLV-I.

Infective dermatitis associated with HTLV-I (IDH) is a chronic, recurrent, exudative eczema occurring in childhood which is considered to be a risk factor for the development of lymphoma and HTLV-I-associated myelopathy/tropical spastic paraparesis. Skin biopsies from 19 patients with IDH were studied histologically and immunohistochemically using the following antibodies: anti-CD3, CD45RO, CD20, CD79a, CD4, CD8, CD56, CD57, TIA-1, granzyme-B, and perforin. A chronic dermatitis similar to atopic and seborrheic dermatitis was observed in 15 cases, whereas architectural aspects mimicking mycosis fungoides were observed in the remaining four. The infiltrate consisted predominantly of CD8+ lymphocytes and of CD57+ cells in the dermis and epidermis. TIA-1 and granzyme-B were expressed in 15/18 cases and 5/19 cases at the proportion of < or = 15% and < or = 3%, respectively. All cases were negative for perforin and CD56. Like other dermatites, histologically IDH may represent a benign simulator of mycosis fungoides. IDH shows a predominance of CD8+ cells and a low percentage of cells with cytotoxic granules, indicating that most CD8+ lymphocytes are not activated. These findings differ from the immunohistochemical pattern of atopic and seborrheic dermatitis, possibly representing additional means of differentiation between IDH and these dermatites. The distribution of CD57+ cells suggests that they play a role in the inflammatory process.

Burkitt's lymphoma of the scapula.

Burkitt's lymphoma may involve the maxilla or mandible, but to date, there has been no reference in the literature to scapular involvement by this tumor. This article describes the case of a 9-year-old child who presented with a huge tumor involving the right shoulder with osteolytic and sclerotic lesions in the scapula. The histopathological findings were suggestive of Burkitt's lymphoma, and the immunohistochemical findings discard a lymphoblastic lymphoma or plasmablastic lymphoma. In addition, in situ hybridization for Epstein-Barr virus encoded small nuclear RNA (EBER) was positive. Although the child also presented a cervical lymphadenopathy, the fact that Burkitt's lymphoma is generally extranodal, the marked swelling of the shoulder, and the extensive involvement of the bone strongly argue that this tumor had its origin in the scapula. This case demonstrates the importance of including Burkitt's lymphoma in the differential diagnosis of lymphoma involving bones in children.

Fatores preditivos de recidiva do carcinoma mamário axila-negativo / Predictive factors for recurrence in axillary node-negative breast carcinoma

Metástase em gânglios linfáticos axilares é um fator prognóstico importante para o carcinoma mamário invasivo. Contudo, cerca de 10 por cento a 20 por cento das pacientes com carcinoma axila-negativo apresentam recidiva da neoplasia. Para avaliar os possíveis fatores preditivos de recidiva, foram revisados os prontuários de 738 casos de pacientes com câncer de mama tratados cirurgicamente no Hospital Aristides Maltez entre 1995-97, encontrando-se 271 pacientes livres de metástases axilares, das quais 32 (11,8 por cento) recidivaram. A média de idade da paciente ao diagnóstico foi de 60 anos. Compararam-se os Grupos 1 (com recidiva) e 2 (sem recidiva) considerando-se idade do paciente ao diagnóstico, tamanho tumoral e grau histológico do tumor (Scarff-Bloom-Richardson). Calcularam-se os riscos relativos (RR) e intervalos de confiança de 95 por cento. Observou-se um RR=3,45 (1,65; 7,22) para recidiva e idade inferior a 40 anos. Com relação ao grau histológico encontrou-se RR=3,08 (1,52; 6,24) de recidiva em pacientes com grau histológico III, comparando-se àquelas com graus I e II. Foram observados um RR=1,13 (IC 95 por cento=0,55; 2,33) para tumores maiores que 2cm e um RR=1,75 (IC 95 por cento=0,74; 4,14) para tumores maiores que 5cm. Concluiu-se que a idade da paciente ao diagnóstico abaixo de 40 anos e grau histológico III são importantes fatores preditivos de recidiva do carcinoma mamário axila-negativo. Estes achados têm fundamento na literatura e podem justificar a adoção de terapêutica complementar para este grupo de pacientes.