Sperm chromosome analysis of two men heterozygous for reciprocal translocations: t(1;9)(q22;q31) and t(16;19)(q11.1;q13.3).

Sperm chromosome complements were analysed in two men who were heterozygous carriers of reciprocal translocations. A total of 363 sperm were karyotyped after in vitro penetration of hamster oocytes, including 180 sperm from a male with a t(1;9)(q22;q31) and 183 from a male with a t(16;19)(q11.1;q13.3). All possible 2:2 and 3:1 meiotic segregations were observed for both translocations. The frequencies of alternate, adjacent 1, adjacent 2, and 3:1 segregations were 46%, 38%, 13%, and 4% for the t(1;9) and 40%, 28%, 31%, and 1% for the t(16;19), respectively. Within the alternate segregation group, the number of normal sperm was not significantly different from the number of sperm carrying a balanced form of the translocation for either of the translocations, as expected. There was no evidence for an interchromosomal effect of either translocation, since the frequencies of numerical abnormalities unrelated to the translocation were within the normal range observed in sperm from control donors. The percentage of sperm with an unbalanced form of the translocation was 54% for the t(1;9) and 61% for the t(16;19).     

Meiotic and sperm chromosome studies in a reciprocal translocation t(1;2)(q32;q36)

Summary Meiotic and sperm chromosomes were studied in a man heterozygous for a reciprocal translocation t(1;2)(q32; q36). Forty-five meiotic metaphase I cells were obtained from semen samples: 86.6% were 22,XY,IV and 13.3% had synaptic anomalies that affected all or some of the bivalents. The quadrivalents observed had a ring configuration (92.3%) or a chain configuration (7.7%). A total of 105 sperm chromosome complements were analyzed: 41% resulted from an alternate segregation, and the percentage of unbalanced sperm was 59%; most of them (71%) resulted from an adjacent 1 segregation. The frequency of anomalies unrelated to the translocation (5.7% numerical and 14.1% structural anomalies) were within the normal range for control donors. There was a good correspondence between the percentage of cells with a ring IV (92.3%) and the proportion of 2:2 segregations (88.6%) and between the percentage of chain IV (7.7%) and the incidence of 3:1 segregations (11.4%).     

Sperm chromosome analysis of two males heterozygous for a t(2;17)(q35;p13) and t(3;8)(p13;p21) reciprocal translocation

Summary Sperm chromosome complements from two males, one heterozygous for the reciprocal translocation t(2;17)(q35;p13) (n = 18) and one for t(3;8) (p13;p21) (n = 73), were analyzed. Only 2:2 segregations were observed with t(2;17): alternate, 56%; adjacent-I, 33%; adjacent-II, 11%. Both 2:2 and 3:1 meiotic segregations occurred in t(3;8): alternate, 34.2%; adjacent-I, 43.8%; adjacent-II, 20.5% and 3:1, 1.4%. A significant excess of chromosomally normal versus balanced sperm complements was observed with both translocation heterozygotes. The frequencies of other chromosome aberrations unrelated to the translocations were 16.7% for t(2;17) and 8.2% for t(3;8). The ratio of X-bearing to Y-bearing sperm was not different from the theoretically expected ratio of 1:1.     

Analysis of segregation and aneuploidy in two reciprocal translocation carriers, t(3;9)(q26.2;q32) and t(3;9)(p25;q32), by triple-color fluorescence in situ hybridization

Meiotic segregation patterns of chromosomes 3 and 9 were analyzed in sperm of two translocation carriers (t(3;9)(q26.2;q32) and t(3;9)(p25;q32)) by triple-color fluorescent in situ hybridization (FISH) with a telomeric DNA probe in addition to two centromeric probes. The frequencies of each sperm product resulting from alternate or adjacent I, adjacent II and 3:1 segregation in a t(3;9)(q26.2;q32) translocation carrier were 88.35%, 5.44% and 5.94%, respectively. On the other hand, the frequencies of each sperm product in a t(3;9)(p25;q32) translocation carrier were 89.23%, 6.02% and 4.48%, respectively. Of all the sperm products, the frequency of normal or chromosomally balanced sperm in a t(3;9)(q26.2;q32) and a t(3;9)(p25;q32) were 52.49% and 47.25%, respectively. The frequencies of each sperm product resulting from various segregations were different between both carriers and significantly deviated from the expected frequencies. Additional dual-color and triple-color FISH were performed to analyze aneuploidy rates for chromosomes 12, 17, 18, X and Y in order to detect any interchromosomal effect; no evidence of an interchromosomal effect was found.     

Cytogenetic analysis of 400 sperm from three translocation heterozygotes

Summary Sperm chromosome complements were studied in three men who carried reciprocal translocations. A total of 400 sperm were karyotyped after in vitro penetration of hamster eggs: 217 sperm from t(2;9) (q21;p22), 164 from t(4;6) (q28;p23) and 19 from t(7;14) (q21;q13). All possible 22 and 31 meiotic segregations were observed for t(2;9) and t(4;6); for t(7;14) only 22 segregations were observed. For alternate segregations, the number of normal sperm was not significantly different from the number of sperm carrying a balanced form of the translocation in any of the translocations, as theoretically expected. The percentage of sperm with an unbalanced form of the translocation was 57% for t(2;9), 54% for t(4;6) and 47% for t(7;14). There was no evidence for an interchromosomal effect in any of the translocations since the frequencies of numerical abnormalities (unrelated to the translocation) were within the normal range of control donors. The frequencies of X- and Y-bearing sperm did not differ significantly from 50%. Results from a total of 17 reciprocal translocations studied by sperm chromosomal analysis were reviewed.     

Meiotic segregation, recombination, and gamete aneuploidy assessed in a t(1;10)(p22.1;q22.3) reciprocal translocation carrier by three- and four-probe multicolor FISH in sperm.

Meiotic segregation, recombination, and aneuploidy was assessed for sperm from a t(1;10)(p22.1;q22.3) reciprocal translocation carrier, by use of two multicolor FISH methods. The first method utilized three DNA probes (a telomeric and a centromeric probe on chromosome 1 plus a centromeric probe on chromosome 10) to analyze segregation patterns, in sperm, of the chromosomes involved in the translocation. The aggregate frequency of sperm products from alternate and adjacent I segregation was 90.5%, and the total frequency of normal and chromosomally balanced sperm was 48.1%. The frequencies of sperm products from adjacent II segregation and from 3:1 segregation were 4.9% and 3.9%, respectively. Reciprocal sperm products from adjacent I segregation deviated significantly from the expected 1:1 ratio (P < .0001). Our assay allowed us to evaluate recombination events in the interstitial segments at adjacent II segregation. The frequencies of sperm products resulting from interstitial recombination in chromosome 10 were significantly higher than those resulting from interstitial recombination in chromosome 1 (P < .006). No evidence of an interchromosomal effect on aneuploidy was found by use of a second FISH method that simultaneously utilized four chromosome-specific DNA probes to quantify the frequencies of aneuploid sperm for chromosomes X, Y, 18, and 21. However, a significant higher frequency of diploid sperm was detected in the translocation carrier than was detected in chromosomally normal and healthy controls. This study illustrates the advantages of multicolor FISH for assessment of the reproductive risk associated with translocation carriers and for investigation of the mechanisms of meiotic segregation of chromosomes.     

Sperm chromosome complements from two human reciprocal translocation heterozygotes

Summary Using the hamster oocyte/human sperm fusion technique, we studied sperm chromosome complements in two male reciprocal translocation heterozygotes, 46,XY,t(11;17)(p11.2;q12.3) and 46,XY,t(1;11) (p36.3;q13.1). For the t(11;17) carrier, 202 sperm chromosome complements were obtained, but 18 karyotypes were not included in the segregation data because of multiple breaks and rearrangements. The alternate and adjacent I types, adjacent II, and 31 segregations accounted for 38.6%, 32.1%, 26.6%, and 2.7% of the sperm analyzed from the t(11;17) carrier. A total of 575 sperm chromosome complements was obtained using sperm from the t(1;11) heterozygote, and 27 karyotypes were excluded from the segregation data because of multiple breaks and rearrangements. For the t(1;11) carrier, the alternate and adjacent I types, adjacent II, and 31 segregations were responsible for 31.4%, 42.9%, 15.9%, and 8.0% of the analyzed sperm chromosome complements. Chromosomal abnormalities unrelated to the translocation, particularly the conservative estimate of aneuploidy frequency, were within the range observed in normal men. Hence, there was no evidence for an interchromosomal effect causing meiotic nondisjunction, despite the large sample sizes studied.     

FISH on sperm heads allows the analysis of chromosome segregation and interchromosomal effects in carriers of structural rearrangements: results in a translocation carrier, t(5;8)(q33;q13)

Using three-color fluorescence in situ hybridization (FISH) with specific DNA probes, we have determined the chromosome segregation pattern of the translocated chromosomes in a human male carrier of a reciprocal translocation, t(5;8)(q33;q13). At the same time, we have assessed the possible interchromosomal effect on pair 21 using dual-color FISH. The segregation results showed that a 45.12% of the spermatozoa analyzed resulted from alternate segregation, 38.31% resulted from adjacent I, 6.97% from adjacent II, and 6.56% from 3:1 segregation. Finally, 1.23% could be either diploid sperm or 4:0 segregation. In both types of adjacent segregations, an excess of products containing short translocated segments (adjacent I) and interstitial regions (adjacent II) were found. Products resulting from the presence of an interstitial chiasma in pair 5 (1.26%), were found much more frequently (P < 0.0001) than those resulting from an interstitial chiasma in pair 8 (0.13%) (evaluated after adjacent II segregation). In 3:1 segregation, the products containing one chromosome were observed more frequently than those containing three chromosomes (P < 0.0001). No evidence of an interchromosomal effect on chromosome 21 was detected, the percentage of disomy 21 being similar to that in the controls (0.23% vs. 0.37%). However, the percentage of diploid sperm (1.18%) was significantly higher (P < 0. 0001) than that in the controls (0.27%). FISH therefore appears to be a useful technique for assessing the percentage of abnormal sperm in translocation carriers. Their application in assisted reproduction centers could offer patients more accurate genetic counseling.     

Familial translocation t(10;21)(q22;q22)

Summary A family is described with a translocation t(10;21)(q22;q22) transmitted through three generations. This family was studied for the apparition of several miscarriages and two sisters with multiple malformations. Both children had a probably partial trisomy of chromosome 10 and a monosomy of chromosome 21 due to a maternal adjacent-2 meiotic segregation.     

Meiotic segregation analysis of a 14;21 Robertsonian translocation carrier by fluorescence in situ hybridization

Meiotic segregation of chromosomes 14 and 21 in sperm from a 14;21 Robertsonian translocation carrier was analyzed with dual-color FISH using two locus-specific DNA probes (Tel 14q and LSI 21). The frequency of normal or chromosomally balanced sperm, resulting from alternate segregation, was 88.42%. The frequency of unbalanced sperm, resulting from adjacent segregation, was 11.25%. These observed frequencies deviated significantly from the theoretical frequencies (33.33% and 66.67%, respectively) based on random chromosome segregation, with sperm resulting from alternate segregation being preferentially produced in the translocation carrier. With respect to the chromosomally unbalanced sperm, the frequency of 21q disomic sperm was 2.45%, which is in agreement with the frequencies of unbalanced fetuses or offspring at the time of amniocentesis or at term (0-4.3%) reported by others. Although the frequency of 14 or 21 nullisomic sperm should be theoretically equal to that of 14q or 21q disomic sperm in both the carrier and controls, the frequency of nullisomic sperm was significantly higher than that of disomic sperm in the carrier (P=0.0009 for chromosome 14, P<0.0001 for chromosome 21) but not in the controls (P=0.091 for chromosome 14, P=0.74 for chromosome 21). This evidence suggests the occurrence of maturation arrest during spermatogenesis of the carrier.     

Analysis of crossing-over in a family with translocation 9;10 involving a chromosome 9 with a pericentric inversion

Summary The presence of two markers on chromosome 9, both a balanced reciprocal translocation and an inversion, allows morphologic demonstration of recombination between the normal and rearranged homologues. In the family under discussion 50% of the progeny studied (two of four) received a translocated 9 without the inversion from a parent with a translocated and inverted 9, indicating crossing-over between members of the chromosome 9 pair. Thus the morphology of the chromosomes allows a recombinat event which is normally invisible to be seen cytologically. Theoretically after crossing-over the balanced reciprocal translocation heterozygote results from adjacent-1 segregation and unbalanced derivative chromosome combinations from alternate segregation. Therefore it cannot be assumed that the balanced progeny necessarily result from alternate segregation and the unbalanced from adjacent-1. The prenatal diagnostic studies presented in this report also show that chromosome analysis of other family members is required when the recombination between homologues produces differences in chromosome morphology between parent and fetus.     

Meiotic products of two reciprocal translocations studied by multicolor fluorescence in situ hybridization

The sperm products of two male carriers of reciprocal translocations were studied by fluorescence in situ hybridization (FISH) using a combination of three probes for each translocation. One patient carried a t(2;18)(p21;q11.2), the other a t(8;9)(q24.2;q32). The probes selected included a centromeric marker for each chromosome involved in the translocation plus a third probe distal to the translocation breakpoint of one of the translocation chromosomes. This assay identifies alternate, adjacent 1, adjacent 2, and 3:1 types of meiotic products. It allows the identification of recombination events and also estimation of the frequency of diploidy. For the t(2;18), the frequency of normal and balanced sperm and of adjacent 1, adjacent 2, and 3:1 products was 43.6%, 29. 8%, 10.5%, and 12.8%, respectively. Similar segregation patterns had been reported for this donor by direct sperm karyotyping of sperm cells. For the t(8;9), the frequency of normal and balanced sperm and of adjacent 1, adjacent 2, and 3:1 products was 44.4%, 41%, 3.1%, and 9.4%, respectively. The frequency of complementary adjacent 1 products was statistically different in both the t(2;18) (P < 0. 0001) and the t(8;9) (P < 0.0001) carrier. When the number of adjacent 2 products with one translocation chromosome (regardless of normal or derivative) was compared to the number of adjacent 2 products with the second translocation chromosome (again, regardless of normal or derivative), no statistical difference was noted for either the t(2;18) (P = 0.32) or the t(8;9) (P = 0.69). Recombination events within the interstitial segment of chromosome 2 were statistically higher than those seen in chromosome 18 (P < 0. 0001), whereas in chromosomes 8 and 9, recombination in the interstitial segments was similar (P = 0.64). The rate of diploidy was similar in both the t(2;18) (0.5%) and the t(8;9) (0.6%). Thus, FISH provides chromosome information on the sperm products produced by translocation carriers, although it cannot provide an assessment of the full chromosome complement of the spermatozoon.     

Human sperm chromosome studies in a reciprocal translocation t(2;5)

Summary Sperm chromosome complements have been studied in a man heterozygous for a reciprocal translocation t(2;5)(p11;q15). Human sperm chromosomes were obtained after fertilization of zona-free hamster eggs. A total of 75 human sperm metaphases were analysed. Of the complements studied, 59 (78.6%) resulted from a 2:2 segregation and 16 (21.3%) from a 3:1 segregation, 4:0 segregation was not observed. Our results indicate that at least 36% of sperm complements were unbalanced with respect to the translocation. The frequency of other chromosome anomalies unrelated to the translocation was 16%.     

Sperm chromosome analysis in a man heterozygous for a paracentric inversion of chromosome 7 (q11q22)

Summary Human sperm chromosomes were studied in a man heterozygous for a paracentric inversion of chromosome 7 (q11q22). The pronuclear chromosomes were analysed after in vitro penetration of golden hamster (Mesocricetus auratus) eggs. Ninety-four sperm chromosome spreads were examined, of which 34 contained the normal number 7 chromosome and 59 the inverted 6. This segregation was significantly different from the expected 1:1 ratio. The number of X- to Y-bearing sperm was 48 and 46 respectively. No sperm contained a recombinant chromosome caused by a crossover within the inversion. The frequency of chromosomal abnormalities in other chromosomes was 9.6%, which is not significantly different from the frequency observed in normal donors (8.9%) in our laboratory. These result suggest that the risk of chromosomally unbalanced sperm is not high for this paracentric inversion.     

Segregation analysis in a man heterozygous for a pericentric inversion of chromosome 7 (p13;q36) by sperm chromosome studies.

We have analyzed 140 sperm chromosome complements from a subfertile man heterozygous for an inv(7)(p13;q36). Seventy-five percent of the chromosome complements were not recombinant: 37.9% contained the normal chromosome 7, and 37.1% contained the inverted chromosome 7. Twenty-five percent of the 140 were recombinant: 7.1% carried a recombinant chromosome 7 with a duplication p and deletion q, 17.1% carried a recombinant chromosome 7 with a duplication q and deletion p, and 0.7% carried both recombinant chromosomes. The frequency of structural chromosomal aberrations unrelated to the inversion was 11.4%, and the frequency of aneuploidy was 2.9%. Both frequencies were not significantly different from those in control donors. Two sperm complements with a second independent, contiguous inversion involving one of the original breakpoints (q36) were observed (1.4%). The risk of producing chromosomally abnormal offspring or spontaneous abortions would be 34.3%. The proportion of X-bearing and Y-bearing sperm was 46.8% and 53.2%, respectively, not significantly different from the expected 1:1 ratio.     

Detection of structural abnormalities in spermatozoa of a translocation carrier t(3;11)(q27.3;q24.3) by triple FISH

Structural chromosome abnormalities in spermatozoa represent an important category of paternally transmittable genetic damage. A couple was referred to our centre because of repetitive abortions and the man was found to be a carrier of a reciprocal translocation t(3;11)(q27.3;q24.3). A tailored fluorescence in situ hybridisation (FISH) approach was developed to study the meiotic segregation patterns in spermatozoa from this translocation carrier. A combination of three DNA probes was used, a centromeric probe for chromosome 11, a cosmid probe for chromosome 11q and a YAC probe for chromosome 3q. The frequency of spermatozoa carrying an abnormal chromosome constitution was compared with baseline frequencies in control semen specimens and it was found that a significantly higher percentage of spermatozoa carried an abnormal constitution for the chromosomes involved in the translocation. A normal or balanced chromosome constitution was found in 44.3% of the analysed spermatozoa, while the remainder exhibited an abnormal chromosome constitution reflecting different modes of segregation (15.9% adjacent I segregation, 6.5% adjacent II segregation, 28.9% 3 : 1 segregation, 0.8% 4 : 0 segregation, 3.6% aberrant segregation). The frequency of aneuploidy for chromosomes X, Y, 13 and 21 was assessed using specific probes but there was no evidence of interchromosomal effects or variations in the sex ratio in spermatozoa from the translocation carrier. In conclusion, structural aberrations can be reliably assessed in interphase spermatozoa using unique DNA probe cocktails, and this method provides insight into the genetic constitution of germ cells and enables evaluation of potential risks for the offspring. Received: 19 September 1997 / Accepted: 27 October 1997     

Multicolor fluorescence in situ hybridization analysis of the spermatozoa of a male heterozygous for a reciprocal translocation t(11;22)(q23;q11)

A reciprocal translocation between chromosomes 11 and 22 is a site-specific translocation that has been seen in many families with no common ancestry. This translocation is of particular interest because balanced carriers have a 0.7–3.7% risk of having children with the supernumerary der(22), resulting from a 3:1 segregation. We have used a three color fluorescence in situ hybridization (FISH) with specific DNA probes to determine the chromosome segregation pattern of a male carrier of a translocation t(11;22)(q23;q11). The probes selected included a centromeric marker for chromosome 11, a marker closely linked to the centromere of chromosome 22, and a third probe distal to the translocation breakpoint of chromosome 22. The results showed that 3 : 1 segregation is preferential in this patient, with 40.1% of spermatozoa belonging to this segregation type. Alternate segregation followed with 27.4% of analyzed spermatozoa; 17.6% resulted from adjacent 1 and 12.5% resulted from adjacent 2 segregation. We detected 0.5% of presumably diploid spermatozoa. Complementary adjacent 1 products were observed at statistically different frequencies (P = 0.02). Complementary adjacent 2 products without recombination in the interstitial segments were also seen at different frequencies (P = 0.002). In 3 : 1 segregation, the products containing one chromosome were observed more frequently than those with three chromosomes (P = 0.0001). The 24,+der(22) gamete was seen more frequently than all of the other gametes combined which had 24 chromosomes resulting from 3 : 1 segregation. The results of this study demonstrate that in this t(11;22) carrier, 3 : 1 segregation is preferential but not exclusive. Received: 9 December 1998 / Accepted: 1 March 1999     

Interchange trisomy 21 by t(1;21)(p22;q22)mat

Interchange trisomy 21 by t(1:21)(p22:q22)mat: Interchange trisomy 21 by t(1;21)(p22;q22)mat was identified in a sporadic patient with Down syndrome. With a 21q22 specific probe, we observed signals on both normal 21 chromosomes and on the der. We reviewed the 23 published reports of families with reciprocal translocations leading to viable offspring with interchange trisomy 21. The breakpoints in chromosome 21 were mainly located in 21q (19/24 instances, including the present report) and in 19/23 cases the other chromosome involved in the translocation was (pairs 1-12). The underlying 3:1 segregation occurred mainly in carrier mothers; only one patient presented a de novo imbalance and in another case the father was the carrier. In addition, there were 4 instances of concurrence with another unbalanced segregation (adjacent-1 or tertiary trisomy) and 3 families with recurrence of interchange trisomy 21. The mean age of 14 female carriers at birth of interchange trisomy 21 offspring (24.8 yr) was lower that the mean (28.3 yr) found in a larger sample of mothers of unbalanced offspring due to 3:1 segregation (mostly tertiary trisomics) and was not increased with respect to the general population average. Overall, these data agree with previous estimates regarding recurrence risk (9-15%) and abortion rate (about 28%) in female carriers ascertained through an interchange trisomic 21 child.     

Cytogenetics of human sperm: meiotic segregation in two translocation carriers.

Meiotic segregation products were studied in sperm from two men heterozygous for the reciprocal translocations t(8;15)(p22;q21) and t(3;16)(p23;q24). A total of 226 and 201 sperm complements, respectively, were analyzed. In each translocation, 63% of complements were unbalanced, and alternate and adjacent 1 percentages were similar. The 3:1 segregation frequencies produced by the two translocations were 3.5% and 5.0%.     

Analysis of sperm chromosome complements from a man heterozygous for a pericentric inversion of chromosome 1

Human sperm chromosomes were studied in a man heterozygous for a pericentric inversion of chromosome (1)(p31q12). Q-banded pronuclear chromosomes were analyzed after in vitro penetration of golden hamster oocytes. A total of 159 sperm were examined: 54% bearing the inverted chromosome 1 and 46% the normal chromosome 1. These frequencies are not significantly different from the theoretical 11 ratio. There were no recombinant sperm with duplications or deficiencies, suggesting that a pairing loop failed to form or that crossing-over was suppressed. The frequency of abnormalities unrelated to the inversion was 5% for numerical, 8.8% for structural, 2.5% for numerical and structural, values not significantly different from control donors studied in our lab. The frequencies of X- and Y-bearing sperm were 46% and 54%, respectively, not significantly different from the expected value of 50%. This is the fifth pericentric inversion studied by human sperm chromosome analysis; recombinant chromosomes have been observed in two of the five cases. Some of the factors associated with an increased risk of recombinant sperm appear to be inversion size greater than 30% of the chromosome and chromosome breakpoints in G-light bands.