Comprehensive meiotic segregation analysis of a 4-breakpoint t(1;3;6) complex chromosome rearrangement using single sperm array comparative genomic hybridization and FISH.

Complex chromosomal rearrangements (CCR) represent rare structural chromosome abnormalities frequently associated with infertility. In this study, meiotic segregation in spermatozoa of an infertile normospermic carrier of a 4-breakpoint t(1;3;6) CCR was analysed. A newly developed array comparative genomic hybridization protocol was used, and all chromosomes in 50 single sperm cells were simultaneously examined. Three-colour FISH was used to analyse chromosome segregation in 1557 other single sperm cells. It was also used to measure an interchromosomal effect; sperm chromatin structure assay was used to measure chromatin integrity. A high-frequency of unbalanced spermatozoa (84%) was observed, mostly arising from the 3:3 symmetrical segregation mode. Array comparative genomic hybridization was used to detect additional aneuploidies in two out of 50 spermatozoa (4%) in chromosomes not involved in the complex chromosome rearrangement. Significantly increased rates of diploidy and XY disomy were found in the CCR carrier compared with the control group (P < 0.001). Defective condensation of sperm chromatin was also found in 22.7% of spermatozoa by sperm chromatin structure assay. The results indicate that the infertility in the man with CCR and normal spermatozoa was caused by a production of chromosomally unbalanced, XY disomic and diploid spermatozoa and spermatozoa with defective chromatin condensation.

Balanced chromosomal translocations in men: relationships among semen parameters, chromatin integrity, sperm meiotic segregation and aneuploidy.

PURPOSE: To analyse relationships between semen parameters, sperm chromatin integrity and frequencies of chromosomally unbalanced, disomic and diploid sperm in 13 Robertsonian and 37 reciprocal translocation carriers and to compare the results with data from 10 control donors. METHODS: Conventional semen analysis, Sperm Chromatin Structure Assay and FISH with probes for chromosomes involved in the individual translocations and for chromosomes X, Y, 7, 8, 13, 18 and 21. RESULTS: Normal semen parameters were found in 30.8 % of Robertsonian and 59.5 % of reciprocal translocation carriers. The rates of unbalanced sperm were 12.0 % in Robertsonian and 55.1 % in reciprocal translocation carriers with no difference between normospermic patients and those showing altered semen parameters. Significantly increased frequencies of spermatozoa showing defects in chromatin integrity and condensation, aneuploidy for chromosomes not involved in a translocation and diploidy were detected in translocation carriers with abnormal semen parameters. Normospermic reciprocal translocation carriers showed an increase in chromosome 13 disomy compared to the control group. There was no relationship between gametic and somatic aneuploidy in 12 translocation carriers studied by FISH on sperm and lymphocytes. The frequency of motile sperm was negatively correlated with the frequency of sperm showing disomy, diploidy and defective chromatin condensation. CONCLUSIONS: Abnormal semen parameters can serve as indicators of an additional risk of forming spermatozoa with defective chromatin and aneuploidy in translocation carriers.

der(4)t(Y;4): Three-generation transmission and sperm meiotic segregation analysis.

We present a family where five members (three males and two females) are carriers of der(4)t(Y;4)(q11.23;p16.3). The adult carriers are phenotypicaly normal and fertile; the boy shows macrocephaly, psychomotor retardation, and atypical autism. The FISH on cultured lymphocytes confirmed that the redundant Yq heterochromatin was attached to the 4p-subtelomeric region maintained on the der(4). Sperm FISH analysis performed in a normospermic der(4) carrier showed a significant distortion of the expected 1:1 ratio of the X- and Y-bearing spermatozoa in favor of the X chromosome and significant lack of Y,der(4)spermatozoa. The overall lack of Y spermatozoa was not balanced even by a relative excess of Y,4 sperm. The analysis of X, Y, 7, 8, 18, and 21 sperm disomy and diploidy did not indicate any interchromosomal effect. The chromosome 4 disomy was significantly increased but still very low to be of considerable reproductive significance. The neurodevelomental phenotype of the boy was probably caused by a gene mutation. The coincidental occurrence of such chromosomal aberration and boy's phenotype might lead to misinterpretation of the causal relationship between these findings. It is necessary to consider the results of chromosomal analysis and clinical records of relatives for provide genetic counseling in such families.

Monozygotic twins with discordant karyotypes following preimplantation genetic screening and single embryo transfer: case report.

PURPOSE: to report a case of monozygotic monochorial diamniotic twins with discordant karyotypes. METHODS AND RESULTS: the pregnancy was achieved following a treatment cycle with intracytoplasmic sperm injection (ICSI) and preimplantation genetic screening (PGS) for chromosomes X, Y, 13, 16, 18, 21, 22. One embryo euploid for studied chromosomes was transferred. Prenatal ultrasonography revealed monozygotic twins. One fetus had growth retardation, multiple organ abnormalities and polyhydramnion. The other twin had normal ultrasound appearance. Delivery on week 29 of gestation resulted in the birth of two females, a stillborn twin with karyotype 45,XX,-13[12]/46,XX,r(13)[3] and a healthy twin with normal karyotype. CONCLUSIONS: the discordance in the twins' karyotypes originated from a mosaic embryo. Structural chromosomal abnormality of the affected twin could not be revealed using standard PGS investigation. Embryo splitting occurred probably due to apoptotic process in an early stage of embryo development. Apoptosis represents one of the possible mechanisms which can explain the embryo twinning process globally.

Sperm fluorescence in situ hybridization study of meiotic segregation and an interchromosomal effect in carriers of t(11;18).

BACKGROUND: Alanced translocations are associated with infertility, spontaneous abortions and birth defects. METHODS: We report the analysis, by multicolour fluorescence in situ hybridization (FISH), of meiotic segregation and aneuploidy of chromosomes X, Y, 7, 8 and 21 in sperm from three men who are carriers of two different translocations involving chromosomes 11 and 18. A control group comprised ten young, healthy normospermic men. RESULTS: The higher prevalence of alternate segregation followed by adjacent 1, adjacent 2 and 3:1, and other segregants was observed in all three patients. Two carriers of the same translocation differed only in the frequency of adjacent 2 segregation (P < 0.01). The carrier of the other translocation showed significantly higher frequency of alternate (P < 0.01) and less adjacent 1 and 3:1 segregation products (P < 0.01). An increased frequency of XY (P < 0.01), YY (P < 0.05) and diploid (P < 0.01) sperm was also detected in the group of translocation carriers compared with the control group. This difference was caused by elevated frequencies of disomy and diploidy in two of our carriers. CONCLUSIONS: The incidence of chromosomally unbalanced or aneuploid gametes varies in the individual translocation carriers even if the same chromosomes are included in the translocation. FISH analysis provides information useful for genetic counseling and assisted reproduction.

Hybridization of the 18 alpha-satellite probe to chromosome 1 revealed in PGD.

Although the chromosome 18 alpha-satellite probe is considered to have a very low polymorphism rate, the routine use of this probe in prenatal diagnosis revealed rare variants in size and copy number of these sequences. A polymorphic signal was detected in preimplantation genetic diagnosis (PGD) for aneuploidy, in a patient with repeated early miscarriages. A third small signal of chromosome 18 alpha-satellite probe was observed in two of four evaluated embryos. Hybridization to the woman's metaphasic lymphocytes revealed that the small signal was localized in the pericentromeric region of chromosome 1. Reanalysis of blastomeres with telomeric probes for chromosome 18q confirmed the presence of only two copies of chromosome 18. Options for verifying PGD analysis results, to prevent misdiagnosis in cases of suspected polymorphism, are discussed. Although some authors speculate about a possible role of heterochromatin polymorphism in infertility, this rare polymorphism of 18 alpha-satellite sequences is in itself probably a normal variant. This is the third report of a cross-hybridization of the chromosome 18 alpha-satellite probe and the first report of the localization of the polymorphic 18 alpha-satellite signal to chromosome 1.

Aneuploidy detection in pigs using comparative genomic hybridization: from the oocytes to blastocysts.

Data on the frequency of aneuploidy in farm animals are lacking and there is the need for a reliable technique which is capable of detecting all chromosomes simultaneously in a single cell. With the employment of comparative genomic hybridization coupled with the whole genome amplification technique, this study brings new information regarding the aneuploidy of individual chromosomes in pigs. Focus is directed on in vivo porcine blastocysts and late morulas, 4.7% of which were found to carry chromosomal abnormality. Further, ploidy abnormalities were examined using FISH in a sample of porcine embryos. True polyploidy was relatively rare (1.6%), whilst mixoploidy was presented in 46.8% of embryos, however it was restricted to only a small number of cells per embryo. The combined data indicates that aneuploidy is not a prevalent cause of embryo mortality in pigs.

Sperm and embryo analysis of similar t(7;10) translocations transmitted in two families.

OBJECTIVE: To compare the sperm meiotic segregation profiles in two men from families with similar t(7;10) translocations and determine the frequency of unbalanced sperm and preimplantation embryos in one couple. DESIGN: Analysis of sperm nuclei and blastomeres by fluorescence in situ hybridization (FISH). SETTING: Research institute. PATIENT(S): Carriers of balanced translocations t(7;10)(q34;q24) and t(7;10)(q36;q24.3). INTERVENTION(S): Multicolor FISH using probes for chromosomes 7, 10, 8, 18, 21, X, and Y on sperm and preimplantation genetic diagnosis (PGD) of blastomeres. MAIN OUTCOME MEASURE(S): Frequencies of meiotic segregation products in sperm and blastomeres and sperm aneuploidy of chromosomes 8, 18, 21, X, and Y. RESULT(S): Similar meiotic segregation patterns, with preferential alternate segregation (50.6% in patient P1, 48.1% in P2) followed by adjacent 1, adjacent 2 and 3:1 segregations, were observed in the sperm of the two carriers. An interchromosomal effect on the sex chromosomes was found when compared with disomy frequencies reported in control donors. The results of preimplantation genetic diagnosis in the first couple are roughly consistent with the sperm analysis results. CONCLUSION(S): Carriers of similar translocations show similar segregation profiles. The in vitro fertilization method accompanied by preimplantation genetic diagnosis increases the chance of translocation carriers fathering a healthy child.

Sperm meiotic segregation and aneuploidy in a 46,X,inv(Y),t(10;15) carrier: case report.

OBJECTIVE: To analyze the meiotic segregation and an interchromosomal effect in sperm of an inv(Y) (p11.1;q11.2),t(10;15) (q25.2;q12) carrier. DESIGN: Case report. SETTING: Research institute. PATIENT(S): Man with a karyotype 46,X,inv(Y),t(10;15), normal sperm parameters, and secondary infertility. INTERVENTION(S): Multicolor fluorescence in situ hybridization using probes for chromosomes 10, 15, 8, 18, 21, X, and Y. MAIN OUTCOME MEASURE(S): Frequencies of meiotic segregation products and aneuploidy of chromosomes 8, 18, 21, X, and Y. RESULT(S): The most frequent type of meiotic segregation was the alternate (40.82%), followed by the adjacent 1 (28.09%), adjacent 2 (16.33%), and 3:1 (9.91%) segregations. Neither deviation from the expected 1:1 ratio of the X- and Y-bearing spermatozoa nor any evidence of an interchromosomal effect on aneuploidy of chromosomes X, Y, 8, 18, and 21 and diploidy was observed in the carrier compared with control donors. The disomies of chromosomes 8 and 21 were equally frequent in X- and Y-bearing spermatozoa of the carrier. CONCLUSION(S): The fluorescence in situ hybridization analysis of meiotic segregation and aneuploidy helps to personalize the reproductive risk in carriers of balanced structural chromosomal aberrations. Complete information concerning the quality of spermatogenesis is necessary in all donors (both translocation carriers and controls) implicated in interchromosomal effect studies.

Sperm and embryo analysis in a carrier of supernumerary inv dup(15) marker chromosome.

We identified a small, paternally inherited, supernumerary marker chromosome, inv dup(15), in a phenotypically normal and normozoospermic male from a couple with reproductive problems. Sperm analysis by fluorescence in situ hybridization (FISH) showed that the marker was present in 26% of sperm nuclei. The disomy 15 was 10 times higher than in normal control donors. FISH analysis for aneuploidies of the other chromosomes showed an increase in nondisjunction of chromosome 21. We also examined 24 embryos by preimplantation genetic diagnosis, and 10 embryos (41.7%) contained the marker. This report provides information about inheritance of inv dup(15) from a male carrier.