Euchromatic variant 16p+. Implications in prenatal diagnosis.

BACKGROUND: Euchromatic imbalances at the cytogenetic level are usually associated with phenotypic consequences. Among the exceptions are euchromatic variants of chromosome 16 (16p+) with normal phenotype. There is a growing list of euchromatic duplications and deletions involving both G-positive and G-negative bands that seem to be phenotypically neutral, but these euchromatic variants are rare. OBJECTIVE: The aim of this report is to describe a new familial case of euchromatic variant 16p+ and to emphasise the misinterpretation of these rare euchromatic variants particularly when ascertained at prenatal diagnosis. METHODS AND RESULTS: Fluorescence in situ hybridisation with clone RP11-261A7 showed an amplified signal in the larger chromosome 16. This clone contains FLJ43855 gene, similar to sodium- and chloride-dependent creatine transporter. CONCLUSION: So, this 16p+ variant that involves amplification of pseudogenetic sequences is considered a polymorphism in normal individuals.

Automated FISH spot counting in interphase nuclei: statistical validation and data correction.

The evaluation of an automated system for Fluorescence In Situ Hybridization (FISH) spot counting in interphase nuclei is presented in this paper. Different types of experiments have been performed with samples from known populations. In all of them the goal is to detect mosaicism of chromosome X in leukocytes from mixtures in known proportions of healthy male and female blood. First the initial results from the automatic FISH analysis system were obtained and evaluated. Then the analysis was modified to reduce systematic errors, so that the results are closer to what an experienced human operator would have obtained (system calibration step). Finally, an additional control probe of chromosome Y was used to detect and discard cells where incorrect hybridization or other abnormal situations had occurred. In each step the system sensitivity was determined by the use of two statistical validation tests, so that the improvement brought about by the correction methods could be assessed. The results obtained in the study showed that, using both corrections, the system is able to detect 10% monosomies with a significance level alpha = 0.1%.

Applying watershed algorithms to the segmentation of clustered nuclei.

Cluster division is a critical issue in fluorescence microscopy-based analytical cytology when preparation protocols do not provide appropriate separation of objects. Overlooking clustered nuclei and analyzing only isolated nuclei may dramatically increase analysis time or affect the statistical validation of the results. Automatic segmentation of clustered nuclei requires the implementation of specific image segmentation tools. Most algorithms are inspired by one of the two following strategies: 1) cluster division by the detection of internuclei gradients; or 2) division by definition of domains of influence (geometrical approach). Both strategies lead to completely different implementations, and usually algorithms based on a single view strategy fail to correctly segment most clustered nuclei, or perform well just for a specific type of sample. An algorithm based on morphological watersheds has been implemented and tested on the segmentation of microscopic nuclei clusters. This algorithm provides a tool that can be used for the implementation of both gradient- and domain-based algorithms, and, more importantly, for the implementation of mixed (gradient- and shape-based) algorithms. Using this algorithm, almost 90% of the test clusters were correctly segmented in peripheral blood and bone marrow preparations. The algorithm was valid for both types of samples, using the appropriate markers and transformations.

Delineation of a duplication map of chromosome 3q: a new case confirms the exclusion of 3q25-q26.2 from the duplication 3q syndrome critical region.

We report on the clinical, cytogenetic, and molecular characterization of a propositus and his mother with a duplication of 3q25-q26, minor anomalies, and mental retardation. The duplication, detected by cytogenetic analysis, was confirmed and delineated by comparative genomic hybridization and fluorescence in situ hybridization using probes previously mapped to the region. Comparison of the mapping data obtained in these patients and those obtained in patients that present with a typical dup(3q) syndrome phenotype shows that the segment duplicated in these patients lies proximally to the reported dup(3q) syndrome critical region, thus explaining the absence in our patients of the characteristic phenotype of dup(3q) syndrome patients. Accumulation of mapping data in patients with segmental duplications of 3q will eventually allow us to build a duplication map of the region and a genotype-phenotype correlation.

Cytogenetic study of neuroendocrine carcinoma of Merkel cells.

We describe the cytogenetic study of a neuroendocrine tumor of Merkel cells which appeared in a patient following a heart transplant. An abnormal karyotype was observed in a metastatic lymph node. The abnormality includes two markers derived from the long arm of chromosome 1, while maintaining two normal chromosomes 1.

Chromosome painting in biological dosimetry: assessment of the ability to score stable chromosome aberrations using different pairs of paint probes.

We exposed human peripheral lymphocytes in vitro to 0.3 and 1 Gy of 60Co gamma rays to evaluate whether the ability and sensitivity to detect chromosomal aberrations by chromosome painting is independent or not to the specific paint probes. To detect structural aberrations (translocations), we painted chromosome spreads simultaneously with two whole-chromosome libraries for chromosomes 1, 2, 3, 4, 5, 6, 7, 11, 13, 16, and 18. To compare the rate of chromosome translocations detected by the different pairs of chromosomes, data were normalized according to the fraction of genome painted and evaluated by unconditional logistic regression. Our results show that any combination of paint probes can be used to score induced chromosomal aberrations. We observed that the amounts of translocations are dose dependent and quite homogeneous within each dose of radiation, independently of chromosomes painted. However, the use of small chromosome probes is not recommended because of the high number of cells to be analyzed due to the small amount of genome painted and because it is more difficult to detect translocations in small chromosomes.

Genetic counselling in a prenatal marker chromosome identified as an i (18p) by in situ hybridization.

The origin of a de novo metacentric small marker chromosome found in a cytogenetic study in amniotic fluid was determined by fluorescence in situ hybridization (FISH) using chromosome specific probes. It was identified as an isochromosome 18p. We want to emphasize that when an extra chromosome is found in prenatal diagnosis and it cannot be identified by conventional cytogenetics banding, FISH should be applied in order to give real risks for fetal anomalies and an accurate genetic counselling.

Burkitt lymphoma with a duplication of der(8)t(2;8). Interpretation of a complex karyotype by chromosome painting.

A 51-year-old male patient was diagnosed with Burkitt lymphoma 3 months after cardiac transplantation. The bone marrow karyotype was very complex, and to better define the complex karyotype we used the in situ suppression hybridization technique. Previously we interpreted this karyotype to be: 48,XY,t(2;8)(p11;q24), +der(2)t(2;8)(p11;q24),del(2)(q23), +7, +der(8)t(2;8)(p11;q24), +12, -13, -18, by G banding techniques, with a duplication of the t(2;8) derivatives. After in situ hybridization we changed to a: 48,XY,t(2;8)(p11;q24),t(2;18)(q23;q22), +7, +der(8)t(2;8)(p11;q24), +12, -13, which implies duplication of only one t(2;8) derivative.