A newly recognised microdeletion syndrome involving 2p15p16.1: narrowing down the critical region by adding another patient detected by genome wide tiling path array comparative genomic hybridisation analysis

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Abstract
After routine cytogenetic analysis on cultured peripheral blood lymphocytes, a normal male karyotype was found and subsequent analysis of the subtelomeric regions by multiplex ligase-dependent probe amplification (MLPA) on DNA extracted from uncultured peripheral blood cells did not reveal any abnormalities as well. The patient’s DNA was then subjected to array based comparative genomic hybridisation (CGH) analysis using our tiling path array that encompasses 32 447 bacterial artificial chromosome (BAC) clones covering the entire human genome as previously described.1 The genome profile (fig 2A) revealed a significant loss within the short arm of chromosome 2. A total of 40 BACs spanning a region of 3.9 Mb from Mb-positions 57.7 (2p16.1) to 61.6 (2p15) (fig 2B) had a significantly lowered test over reference log2 intensity ratio with RP11-426N8 and RP11-470L12 being the telomeric and centromeric breakpoint clones, respectively. The loss was validated and confirmed by region-specific MLPA analysis with probes designed in VRK2 and USP34 (fig 2C). According to ISCN 2005 nomenclature, the combined karyotype is described as follows: 46,XY.arr cgh 2p16.1p15(RP11-426N8→RP11-470L12)x1. Please note that the aberrant region detected by array CGH is indicated from pter to qter in the karyotype, whereas a similar deletion detected by fluorescence in situ hybridisation (FISH) would be indicated with the breakpoint more proximal to the centromere specified first—that is, 46,XY.ish del(2)(p15p16.1)(FANCL-,REL-). Next, four BAC clones from the aberrant region were selected and labelled for validation by FISH on metaphase spreads from our patient. All four probes were hybridised individually and on separate slides. With each probe, only one signal was observed in 20/30 cells, whereas the remainder of the cells revealed a normal pattern with two signals, one on each 2p (data not shown). We conclude from this that the microdeletion was present in mosaic in our patient with both normal cells and abnormal cells. Parental samples were subsequently analysed by routine chromosome analysis and by FISH analysis with the aforementioned probes. In both parents, a normal karyotype and a normal FISH pattern was observed. The microdeletion in our patient has occurred de novo, which is to be expected considering the fact that it appeared to be present in mosaic. In addition to other reports describing the detection of even low level mosaics by array CGH analysis,2 3 this is yet another example demonstrating the strength of array CGH not only to reliably detect imbalances smaller than 10 Mb, but also the ability to detect unbalanced chromosome abnormalities present in mosaic. Convincing data are accumulating that seem to confirm that aberrant cells may be underrepresented in the PHA stimulated, cultured T lymphocytes that are used for routine cytogenetic analysis. The aberrant cells are more difficult to stimulate to divide and are therefore more likely to be detected with array CGH analysis using DNA isolated directly from whole blood samples.3

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