Fourteen marker chromosomes were studied by FISH (fluorescence in-situ hybridization) in cytogenetic preparations from 13 patients. The derived markers were identified as one isodicentric bisatellited mar(22), one fragment sized r(X), one fragment sized r(Y), one i(18p), small autosomal ring markers in three different patients derived from chromosomes 2, 8, and 8, a marker comprised of 9p and part of 9qh, and 3 bisatellited apparently monocentric markers; one of each from chromosomes 13 or 21, 14 or 22, and 15. Two fragment sized small ring markers in one patient and a small ring marker in another were negative with all twenty-two different probes used. In addition, the small ring marker Y chromosome that was found in a boy with karyotype 46,X,-Y,+mar was negative with both pDXZ1 and pDYZ3. This anomaly of negative results with the battery of centromeric alphoid probes can be explained if one breakpoint for some small ring markers is very near to or within the centromere. Only some of the pericentromeric repetitive sequences in the normal chromosome would be represented in the chromosome specific alphoid probes, and presumably those corresponding to the currently available probes are truncated during the formation of the unidentified markers. In three of the small ring markers the FISH signal on the marker was much stronger than on the normal homologues in various proportions of cells, and this may indicate that some of the fragment sized small rings were multicentric. The literature was reviewed for Distamycin A/DAPI negative small ring markers that were present as extra chromosomes. There were only single published cases of most small rings but there were three r(8) cases, two r(1) cases, two r(12) cases, and two r(20) cases, uncomplicated by the presence of other chromosome abnormalities. Most cases with similar small rings were quite dissimilar phenotypically and syndrome identification was not possible, but in pooled data, 18/23 (about 80%) were developmentally and/or phenotypically abnormal. Some patients (5/23, about 20%) with small rings were dysmorphic without intellectual handicap. Of 28 such patients with small ring markers (Distamycin/Dapi negative) in pooled data there are 6 (about 20%) with multiple markers mostly derived from different chromosomes. This is a very high figure and would suggest that the ring formation events, although involving different chromosomes, must be related and must be an indicator of the mechanism of origin of this group of markers.

Seven supernumerary small ring marker autosomes were studied. The pantelomere probe (Oncor) in conjunction with scoring for dicentric rings was used to confirm ring morphology. The small rings were identified mainly by FISH with chromosome probe arrays (Cytocell) containing representations from all 24 chromosomes and the rings were derived from chromosomes 7, 8 (three cases), 11, 12, and 14. The effectiveness of the array methodology in identifying markers was tested. Microsatellite DNA data showed biparental disomy (BPD) was present for the rings from chromosomes 7 and 14 thereby excluding UPD, both were de novo but the ring 14 was of paternal origin. The literature on supernumerary small ring autosomes was reviewed excluding chromosome 15. The grade and distribution of mosaicism was invoked as the major determinant of the differences in phenotype and, in addition, variation was attributed to the possibility of different contributions from each chromosome arm. There are 88 published supernumerary small ring cases in total, with phenotypic data attributable to the respective rings in 77 cases and all chromosomes being represented except chromosome 17. Of the prenatally ascertained cases, where there was adequate phenotypic data, 30% had an abnormal phenotype attributable to the ring, and there were 44% familial cases in this group. Of the postnatally ascertained small rings, 75% had an abnormal phenotype attributable to the ring and there were 13% familial cases. This higher abnormality rate is concordant with the considerable ascertainment bias of this latter group and the prenatal data are recommended for genetic counseling. Although data are small there were some differences between the rings derived from different chromosomes. Chromosomes 3 and 8 demonstrate the extremes. Of the supernumerary small r(8) cases reviewed including the three presently described, 8/11 had an abnormal phenotype attributable to the marker but of the small r(3) cases, only 1/6 had an abnormal phenotype. Two of the present r(8) were studied with the GATA4 probe at 8p23.1. The r(8) in case 2 (patient moderately retarded) was comprised mostly of an intact 8p whereas the larger r(8) in case 3 (normal phenotype) was missing 8p23.1 --> pter and had more of 8q contributing to the ring. In other supernumerary rings postnatally ascertained, there is mostly insufficient data but there is an abnormal phenotype in 8/11 cases with multiple small rings, in 5/6 cases with r(20), and in 5/10 with r(1). A novel origin for supernumerary small rings is proposed: that they may originate from incompletely digested superfluous (haploid) pronuclei. The small rings presumptively so formed may occasionally be transfected into the zygote nucleus. The high proportion ( approximately 12.5%) of cases with multiple supernumerary small rings almost always of different centromeric origin is consistent with this concept.

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Two cases of submicroscopic recombinants of intrachromosomal transposition of telomeres, one each from chromosome 1 and 2 are described. Meiotic crossing-over would generate the recombinants from these reciprocal rearrangements. In both cases, which were detected by FISH with subtelomeric probes, there is a minute deletion of the qter region and a second presence of the pter subtelomeric region on the respective qter, i.e., a duplication of 1pter or 2pter respectively. The deletion on 2qter (case 2) was confirmed by microsatellite inheritance and was of paternal origin, but in case 1 there was no detectable 1q deletion other than of the subtelomeric probe, and parental origin could not be determined. The present case 2 with del(2qter)/dup(2pter) shares many features with reported cases of simple deletion (2qter) but did not have features of Albright hereditary osteodystrophy, which are seen in half of such deletion patients. The clinical features present in case 1 were similar to those of the previously reported case of a submicroscopic 1qter deletion but also to cases with microscopically visible 1qter deletions, presumably because of gene enrichment in subtelomeric regions. Recombinants of such intrachromosomal subtelomere transpositions detected by subtelomeric probes may comprise up to 10% of submicroscopic pter or qter deletion cases. Other cases of this unusual mechanism may be detected with more common use of subtelomeric probes. It is suggested the bouquet associations of telomeres in early meiosis may facilitate such unusual rearrangements.

We report three new cases of chromosome 13 derived marker chromosomes, found in unrelated patients with dysmorphisms and/or developmental delay. Molecular cytogenetic analysis was performed using fluorescence in situ hybridization (FISH) with chromosome-specific painting probes, alpha satellite probes, and physically mapped probes from chromosome 13q, as well as comparative genomic hybridization (CGH). This analysis demonstrated that these markers consisted of inversion duplications of distal portions of chromosome 13q that have separated from the endogenous chromosome 13 centromere and contain no detectable alpha satellite DNA. The presence of a functional neocentromere on these marker chromosomes was confirmed by immunofluorescence with antibodies to centromere protein-C (CENP-C). The cytogenetic location of a neocentromere in band 13q32 was confirmed by simultaneous FISH with physically mapped YACs from 13q32 and immunofluorescence with anti-CENP-C. The addition of these three new cases brings the total number of described inv dup 13q neocentic chromosomes to 11, representing 21% (11/52) of the current overall total of 52 described cases of human neocentric chromosomes. This higher than expected frequency suggests that chromosome 13q may have an increased propensity for neocentromere formation. The clinical spectrum of all 11 cases is presented, representing a unique collection of polysomy for different portions of chromosome 13q without aneuploidies for additional chromosomal regions. The complexity and variability of the phenotypes seen in these patients does not support a simple reductionist view of phenotype/genotype correlation with polysomy for certain chromosomal regions.