Diploid Sperm Research Articles

Macrocephaly in Bull Spermatozoa Is Associated with Nuclear Vacuoles, Diploidy and Alteration of Chromatin Condensation

Spermatozoa from 2 dairy AI (artificial insemination) bulls (A and B), identified by their abnormal spermiogram with cells depicting frequent macrocephaly, double tails and nuclear vacuoles, were case-investigated and compared to normal spermatozoa from a control AI sire (C). Head sizes were measured and morphological abnormalities scored using brightfield and differential interference contrast microscopy. The degree of sperm maturation and of resistance to acid-induced DNA denaturation in situ were determined after uploading of acridine orange using flow cytometry of 5,000 cells/sample. Nuclear fragmentation, i.e. the ratio of red to total (red + green) fluorescence, reached 7.1% and 31% in bulls A and B, compared to 2% in bull C. The proportion of immature spermatozoa, i.e. those with incomplete histone-protamine exchange and depicting higher green fluorescence compared to the main population of the control bull, reached 9.54% in A and 7.75% in B, compared to only 0.47% in the control. In the second part of this study the previously unknown chromosomal constitution of large-headed spermatozoa of bull A was investigated by fluorescence in situ hybridization using an X-Y painting probe set. The 7.5% XY-bearing cells and the presence of diploid spermatozoa detected by flow cytometry indicate a meiotic arrest in the first division in bull A, becoming the first proven case of association of macrocephaly and M1 diploidy. The diverse approaches used for the investigation of spermatozoal DNA provide insights into the etiology of macrocephaly.

The influence of ploidy level on ultrastructure and motility of tench Tinca tinca (L.) spermatozoa

The ultrastructure of diploid and triploid tench Tinca tinca (L.) spermatozoa were examined using electron microscopy focusing on parameters that influence movement. Triploid tench were produced artificially using a cold shock. Spermatozoa of triploid males in comparison with diploids featured significantly larger head (P < 0.01), higher amount of mitochondria (P < 0.05), and, surprisingly larger widths of the peripheral doublets and central pair of microtubules and the single microtubule (P < 0.01). However, the diameters of the flagellum were without significant differences as well as the length of the flagellum and length and width of the midpiece. Also motility parameters of spermatozoa did not significantly differ between diploid and triploid males, but the total velocity (summary of spermatozoa velocity and duration of movement) positively correlated with the flagellum length and negatively with the head diameter of tench spermatozoa with a high significant influence (P < 0.01).

Diploid males, diploid sperm production, and triploid females in the ant Tapinoma erraticum

Under complementary sex determination (CSD), females of Hymenoptera arise from diploid, fertilized eggs and males from haploid, unfertilized eggs. Incidentally, fertilized eggs that inherit two identical alleles at the CSD locus will develop into diploid males. Diploid males are usually unviable or sterile. In a few species, however, they produce diploid sperm and father a triploid female progeny. Diploid males have been reported in a number of social Hymenoptera, but the occurrence of triploid females has hardly ever been documented. Here, we report the presence of triploid females, diploid males, and diploid sperm (produced by diploid males and stored in queen spermathecae) in the ant Tapinoma erraticum. Moreover, we show variations in the frequency of triploids among female castes: Triploid females are more frequent among workers than virgin queens; they are absent among mated, reproductive queens. The frequency of triploid workers also varies between populations and between nests within populations.

Die Pepsinvorbehandlung menschlichen Spermas als Standardmethode in der impulscytophotometrischen Analyse*

The 30 minutes pepsin incubation of human semen has proved to be a standard pretreatment procedure for impuls cytophotometry (ICP). This method, in comparison to papain treatment, shows a better preservation of the material and avoids the phenomenon of "disintegration" and reduced staining property. The CV of 1C peak is small enough. Additionally, the immature spermatozoa, earlier dissociated late elongated spermatids, diploid spermatozoa, dual forms and diploid semen cells (spermatogenetic cells, leukocytes, epithelial cells are demonstrated in small peaks, a fact which can help explain the disturbances in pathological cases.

Chromosome Doubling in Early Spermatogonia Produces Diploid Spermatozoa in a Natural Clonal Fish1

The natural clonal loach Misgurnus anguillicaudatus (Teleostei: Cobitidae) is diploid (2n = 50) and produces genetically identical unreduced eggs, which develop into diploid individuals without any genetic contribution from sperm. Artificially sex-reversed clones created by the administration of 17alpha-methyltestosterone produce clonal diploid sperm. In metaphase spreads from testicular cells of the sex-reversed clones, spermatocytes had twice the normal number of chromosomes (50 bivalents) compared with those of normal diploids (25 bivalents). Thus, the production of unreduced diploid spermatozoa is initiated by premeiotic endomitosis (or endoreduplication), chromosome doubling before meiosis, and is followed by two quasinormal divisions. Larger nuclei in the germ cells were observed in all stages of type B spermatogonia in the testes of the sex-reversed clones. In contrast, besides having larger type A spermatogonia, the sex-reversed clones also had the type A spermatogonia that were the same size as those of normal diploids. It follows that chromosome duplication causing unreduced spermatogenesis occurred in the type A spermatogonia. The presence of tetraploid type A and early type B spermatogonia, identified by labeling with antispermatogonia-specific antigen 1, was verified using DNA content flow cytometry. These results support the conclusion that chromosome doubling occurs at the type A spermatogonial stage in diploid spermatogenesis in the clonal fish.

Chromosomal unbalancements in sperm and oocytes of two Italian cattle breeds as determined by dual color fluorescentin situ hybridization (FISH)

Aneuploidy is one of the most important causes of embryonic and foetal mortality in mammals. In order to assess the possible risk of chromosomal abnormalities in germ cells of domestic animals we investigated the aneuploidy rates on partially decondensed sperm and in vitro matured oocytes in two cattle breeds, Italian Friesian (I.F.) and Italian Brown (I.B.), by using FISH with chromosome-specific painting probes (chromosomes X-Y for sperm and chromosomes X-5 for oocytes). For each bull, more than 5,000 sperm were analyzed, for a total of 52,586 and 51,342 sperm cells for the two breeds, respectively. Aneuploid and diploid sperm had, respectively, a frequency of 0.110% and 0.050% in the I.F. and 0.078% and 0.062% in the I.B. breeds. Out of 100 in vitro matured oocytes for each breed, on the average, diploidy affected 11.2% and 18.4% in the I.F. and I.B., respectively, whereas disomy for chromosome X-5 had a frequency of 2% in the I.F. and 2.5% in the I.B. breeds. Further studies are needed to expand our knowledge on frequency of aneuploidy in sperm and oocytes of domestic animals, in order to assess their impact on productive and reproductive efficiency, also in relation to climatic changes and environmental hazards.

Can IMSI be efficient in situation of ICSI failure and teratospermy, with special study of macrocephaly?

OBJECTIVE: Intracytoplasmic morphologically selected sperm injection (IMSI) is proposed in 2 situations: previous ICSI failure and severe teratozoospermia. IMSI allows to select, at high magnification”top”spermatozoa (normal size, normal shape, normal morphology) and those”top”injected spermatozoa would allow better development. Macrocephalic sperm head syndrome is known to be associated with aneuploid and polyploid sperm content. IMSI could allow to select spermatozoa with normal chromosomal content. DESIGN: To evaluate the efficiency of IMSI, when ICSI either would not be performed or had been done inefficiently. MATERIALS AND METHODS: We performed IMSI on 27 cases: 13 for major teratozoospermia (11 cases <4% of sperm with typical form, and 2 cases with respectively 94 and 96% macrocephaly and no normal form = group 1 and 14 for failure of 2 ICSI =group 2). The mean female ages were similar (34.3 in group 1, 31.6 in group 2), as the type of infertility, and the basal hormonal levels. The efficiency of IMSI selection for macrocephaly was evaluated by FISH:”ICSI like” (x400) and “IMSI like” (x6000) “normal” spermatozoa were aspirated, fixed, and FISH analysed with X, Y and 18 probes. RESULTS: Fertilization rates were similar (52/92) and (53/89), as well as pregnancy rates (3/14 and 5/11) in the 2 groups. Concerning the FISH results, before selection, patient 1 had only 10/1012 (1.0%) normal spermatozoa, 14 (1.4%) were haploid but aneuploid, 200 (19.8%) diploid and 788 (77.8%) polyploid. Among the 31 “ICSI like selected” spermatozoa, only 1 (3.2%) was normal, 15 (48.4%) were haploid but aneuploid and 16 (51.6%) diploid. All the 6 “IMSI like” selected top spermatozoa were haploid but aneuploid. For patient 2, before selection, only 3 (0.3%) spermatozoa were normal, 26 (2.6%) haploid but aneuploid, 103 (10.3%) diploid and 870 (86.8%) polyploid. Among the 8 “ICSI like” selected spermatozoa, only 1 was normal, 4 (50.0%) were haploid but aneuploid, 1 (12.5%) diploid and 2 (25.0%) polyploid, no “top spermatozoa” was found at X6000. CONCLUSIONS: Polyploid or diploid spermatozoa can be eliminated by high magnification sperm selection (p<0.05), but no euploïd spermatozoa was found by IMSI of the macrocephalic sperms, thus IMSI is not the solution.The other results are encouraging but randomized studies are needed to confirm the better efficiency of IMSI compared to ICSI in case of previous ICSI failure or severe teratozoospermia.

Triple genetic identities for the complete hydatidiform mole, placenta and co-existing fetus after transfer of a single in vitro fertilized oocyte: Case report and possible mechanisms

We found different genotypes for the complete hydatidiform mole (CHM), placenta and co-existing fetus derived from a single in vitro fertilized human oocyte by the analysis of short tandem repeat (STR) DNA markers. The molar tissue was found to be heterozygously androgenetic. The fetus and placenta contained identical maternal, but different paternal genomes. Two models were proposed to account for the identification of triple genetic identities in a single fertilized oocyte. In the first model, the oocyte was fertilized by a diploid sperm, resulting in diandric triploidy. Premature cytokinesis resulted in early splitting of a cytoplasmic fragment with one copy of the replicated sperm chromosome, which developed into a heterozygous CHM. The bipolar spindle in syngamy pulled the other copy of sperm chromosomes and replicated oocyte chromosomes to form two blastomeres, which develop into the fetus and placenta, respectively. In the second model, the oocyte was fertilized by two haploid sperms, followed by tripolar spindle formation. Whatever is the mechanism, this case provides direct evidence that CHM can be derived from an oocyte containing an intact maternal genome.

DNA flow cytometry of human spermatozoa: Consistent stoichiometric staining of sperm DNA using a novel decondensation protocol

Rapid flow cytometric measurement of the frequency of aneuploid human sperms is in increasing demand but development of an exploitable method is hindered by difficulties of stoichiometric staining of sperm DNA. An aggressive decondensation protocol is needed after which cell integrity still remains intact. We used flow cytometry to examine the effect of lithium diiodosalicylate (LIS, chaotropic agent) on fluorescence intensity of propidium iodide-treated human spermatozoa from 10 normozoospermic men. When flow cytometric identification of diploid spermatozoa was achieved, validation was performed after sorting by three-color FISH. In contrast with the extremely variable histograms of nondecondensed sperms, consistent identification of haploid and diploid spermatozoa was possible if samples were decondensed with LIS prior to flow cytometry. A 76-fold enrichment of diploid sperms was observed in the sorted fractions by FISH. A significant correlation was found between the proportion of sorted cells and of diploid sperms by FISH. Application of LIS during the preparation of sperm for flow cytometry appears to ensure the stoichiometric staining of sperm DNA, making quantification of aneuploid sperm percentage possible. To our knowledge this is the first report in terms of separating spermatozoa with confirmedly abnormal chromosomal content. High correlation between the proportion of cells identified as having double DNA content by flow cytometry and diploid sperm by FISH allows rapid calculation of diploidy rate.

Ploidy manipulation using diploid sperm in the loach,Misgurnus anguillicaudatus: a review

Summary This paper assesses the present state of the art of ploidy manipulation in the loach, Misgurnus anguillicaudatus (Teleoste: Cobitidae). Diploid sperm can be obtained from natural tetraploid individuals with four sets of homologous chromosomes. Using diploid sperm, various polyploids and androgenetic diploids have been produced. Cryptic clonal lineages are also recognized in wild populations of the loach. They produce unreduced diploid eggs genetically identical to somatic cells of the mother fish and most diploid eggs develop gynogenetically as a member of the clone. However, some eggs develop to triploid and/or diploid-triploid mosaic individuals by incorporation of sperm nucleus. Diploid-triploid mosaic males exclusively generate fertile diploid sperm with clonal genotypes. Such diploid sperm can also be obtained from artificially sex-reversed clonal individuals. Recent population studies suggested that Japanese M. anguillicaudatus might not be a single species, but a complex involving cryptic species, because wild populations exhibited genetic differentiation at interspecific level. This implies possible relationship between atypical reproduction and natural hybridization in the loach.

Complete hydatidiform mole with coexisting dichorionic diamniotic twins following testicular sperm extraction and intracytoplasmic sperm injection

We present the first report of complete hydatidiform mole (HM) with coexisting dichorionic diamniotic twins. This pregnancy was achieved after testicular sperm extraction and intracytoplasmic sperm injection (ICSI) for azoospermia in the woman's husband. Standard in vitro fertilization may cause multisperm fertilization and increase triploid partial HM and complete HM, which arise from dispermic fertilization. In contrast, ICSI can avoid multisperm fertilization. In our case, paternal isodisomy in the molar tissue was confirmed by microsatellite analysis suggesting that it resulted from duplication of a haploid paternal genome following monospermic fertilization of an inactivated oocyte or from monospermic fertilization of an inactivated oocyte with a diploid sperm. Although the patient was eager to continue the pregnancy, the size of the HM component increased rapidly and termination of the pregnancy was required for pre-eclampsia-like symptoms at 15 weeks of gestation. After the operation, chemotherapy was initiated for persistent trophoblastic disease.

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

Alanced translocations are associated with infertility, spontaneous abortions and birth defects. 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. 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. 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.

The chemotherapeutic agents nocodazole and amsacrine cause meiotic delay and non-disjunction in spermatocytes of mice

Aneuploidy of germ cells contributes to reduced fertility, foetal wastage and genetic defects. The possible risk of aneuploidy induction by the cancer chemotherapeutic drugs amsacrine (AMSA) and nocodazole (NOC) was investigated in male mice. Two molecular cytogenetic approaches were used: (1) the BrdU-incorporation assay to test the altered duration of meiotic divisions and (2) the sperm-FISH assay to determine aneuploidy induction during meiosis by observing hyperhaploid and diploid sperm. Sperm were sampled from the Caudae epididymes of treated and solvent control males. Single intraperitoneal injections with NOC (35 mg/kg) and AMSA (15 mg/kg) caused a meiotic delay of 24 h. The timing of sperm sampling for the sperm-FISH assay was adjusted accordingly, i.e. 23 days after treatment. Mice were treated with 18, 35 and 50 mg/kg of NOC, or 5, 10, 15 and 20 mg/kg of AMSA. Significant dose-dependent increases above the concurrent controls in the frequencies of hyperhaploid sperm were found with both agents. Significant increases in the frequencies of diploid sperm were found only with AMSA. These results provide a basis for genetic counselling of patients under AMSA or NOC chemotherapy. During a period of 3–4 months after the end of chemotherapy, they may stand a higher risk of siring chromosomally abnormal offspring.

Mechanisms giving rise to triploid zygotes during assisted reproduction

To review information on the origin of triploid zygotes as gathered from assisted reproduction techniques. Identification of relevant literature by a MEDLINE search and own experience on the basis of cytogenetic studies of abnormally fertilized oocytes. None. None. None. None. Penetration of two haploid spermatozoa or of a single diploid spermatozoon into the oocyte causes diandric triploidy. The first case can be discerned by formation of a total of three pronuclei, whereas the second process will remain undetected, because it involves a female and a single but diploid male pronucleus. Digynic triploidy after intracytoplasmic sperm injection is characterized by nonextrusion of the second polar body and formation of three pronuclei. Digyny can also result from the fertilization of diploid giant oocytes. Depending on how maturation of these gametes proceeds, three or only two pronuclei will be observed. Thus, the size of the pronuclear stage must be considered for a successful identification of the abnormality. Endoreduplication within the female pronucleus is not detectable and may represent another, albeit rare, origin of digynic triploidy. Routine inspection of the number of pronuclei is not an absolutely reliable tool for excluding the development of triploid embryos. Observations during assisted reproduction may yield valuable information on the origin of triploidy.

The formation of improved tetraploid population of red crucian carp × common carp hybrids by androgenesis

Bisexual fertile diploid androgenetic individuals (A(0)) (2n=100) were formed by androgenesis. In this way, the diploid spermatozoa from male allotetraploid hybrids (AT) (4n=200) of red crucian carp (Carassius auratus red var.) (female) x common carp (Cyprinus carpio L.) (male) were used to fertilize the UV-treated haploid eggs of goldfish (Carassius auratus), and living androgenetic diploid fish were developed. The A(0) became sexually mature at the age of 2 years, and they fertilized with each other to form their offspring (A(1)). In this study, we observed the chromosomal number, gonadal structure and appearance of A(1) fish. The results are as follows: (1) In A(1), there were 85% tetraploids (A(1)-4n), 10% triploids (A(1)-3n) and 5% diploids (A(1)-2n), suggesting that diploid A(0) could produce diploid gametes. It was concluded that the formation of diploid gametes generated from diploid A(0) was probably related to the mechanism of pre-meiotic endoreduplication. (2) Among A(1), only A(1)-4n possessed normal ovaries and testes. The mature males of A1-4n produced white semen. Under the electron microscope, the head of diploid sperm generated by A(1)-4n was bigger than that of haploid sperm generated by red crucian carp. In the testes of the A(1)-4n, there were many mature normal spermatozoa with a head bearing plasma membrane and a tail having the typical structure of "9+2" microtubules. Between the head and the tail, there were some mitochondria. The ovaries of A(1)-4n developed well and mainly contained II, III and IV-stage oocytes. The IV-stage oocytes were surrounded by inner and outer follicular cells. The micropyle was observed on the oolemma of follicular cells. There were abundant yolks and plenty of endoplasmic reticulum in the cytoplasm of IV-stage oocytes. Because A(1)-2n and A(1)-3n were distant crossing diploid hybrids and triploid hybrids respectively, they possessed abnormal gonads, and no mature semen and eggs were observed. (3) Compared with allotetraploids, the A(1)-4n fish not only had advantages such as fast growth rate and strong resistibility but also showed some new good performances such as high ratio of body width to body length, smaller heads and shorter tails. These results indicated that androgenesis could produce bisexual fertile tetraploids and improve the shape of allotetraploid hybrids as well, which will be of great significance in both the cell genetics research and fish breeding.

Diploid sperm produced by artificially sex‐reversed Clone loaches

Clone loaches reproduce unisexually in a wild population of Hokkaido Island, Japan. These clone loaches produce genetically identical unreduced eggs which develop to diploid individuals without any genetic contribution of sperm donors. In the present study, sex reversal of clone loaches was attempted and the reproductive potential of resultant clone males was examined. Clone loaches administered 0.5 ppm of 17-alpha methyltestosterone (MT) for 30 days from 1 month after hatching differentiated into physiological males. These sex-reversed clone males produced fertile spermatozoa with a diploid DNA content. Diploid spermatozoa had significantly larger heads than normal haploid sperm, but had a normal shape showing a head, mid-piece, and tail. The motility of diploid spermatozoa was low after ambient water was added. Concentration of diploid spermatozoa per unit of sperm was lower than that of control haploid spermatozoa. Microsatellite genotyping revealed that triploid progeny from the cross between a normal diploid female and a sex-reversed clone male had two alleles specific to the diploid clone male and one allele of the mother loach. These results indicated that the sex-reversed clone males produced fertile diploid spermatozoa genetically identical to the clone lineage.

Induced interspecific androgenesis using diploid sperm from allotetraploid hybrids of common carp×red crucian carp

In the present study, viable diploid androgenetic individuals (AT-ag) (2 n = 100) have been developed in UV-radiated eggs of goldfish (GF) ( Carassius auratus) following fertilization with diploid spermatozoa from male allotetraploid hybrid (AT) F 13 (4 n = 200) of red crucian carp ( C. auratus red var.) (♀) × common carp ( Cyprinus carpio L.) (♂). In the experimental groups, eggs were inactivated by UV-rays, the range of UV-radiation dose tested varied from 240 to 600 mJ/cm 2, the relative high hatching rate (4.1 ± 0.4%) and survival from hatching to start of feeding (41.3 ± 2.6%) were given at UV dose of 300 mJ/cm 2. The androgenetic nature was confirmed by chromosome observation and DNA content measurement of somatic cells by a flow cytometer. The AT-ag were diploid (2 n = 100) whereas the controls (GF × AT) were triploids (3 n = 150). The diploid AT-ag grew slower than the triploid GF × AT. The AT-ag and GF × AT were also morphologically distinguishable. The AT-ag began to sexually mature at the age of 2 years, and their fertility further increased at the age of 3 years. The tetraploids were obtained in the offspring (F 1ag × ag ) produced by mating the males with females of AT-ag and in the progeny (AT-ag × AT) generated by crossing the males of AT with the females of AT-ag, indicating that diploid AT-ag could produce diploid gametes. All the sex ratios of males to females in AT-ag, F 1ag × ag , AT-ag × AT were not significant ( P > 0.05) from the ratio of 1:1, suggesting the XXXY sex-determining mechanism in the males of AT. The present study indicated that bisexual fertile diploid AT-ag were successfully induced from diploid spermatozoa of AT.

Evidence for recombination of mitochondrial DNA in triploid crucian carp.

In this study, we report the complete mitochondrial DNA (mtDNA) sequences of the allotetraploid and triploid crucian carp and compare the complete mtDNA sequences between the triploid crucian carp and its female parent Japanese crucian carp and between the triploid crucian carp and its male parent allotetraploid. Our results indicate that the complete mtDNA nucleotide identity (98%) between the triploid crucian carp and its male parent allotetraploid was higher than that (93%) between the triploid crucian carp and its female parent Japanese crucian carp. Moreover, the presence of a pattern of identity and difference at synonymous sites of mitochondrial genomes between the triploid crucian carp and its parents provides direct evidence that triploid crucian carp possessed the recombination mtDNA fragment (12,759 bp) derived from the paternal fish. These results suggest that mtDNA recombination was derived from the fusion of the maternal and paternal mtDNAs. Compared with the haploid egg with one set of genome from the Japanese crucian carp, the diploid sperm with two sets of genomes from the allotetraploid could more easily make its mtDNA fuse with the mtDNA of the haploid egg. In addition, the triple hybrid nature of the triploid crucian carp probably allowed its better mtDNA recombination. In summary, our results provide the first evidence of mtDNA combination in polyploid fish.

A human tetraploid pachytene spermatocyte as the possible origin of diploid sperm: a case report

Diploid spermatozoa represent 0.2-0.3% of all spermatozoa in the normal population and cause 8.3% of diandric triploids. Errors in meiosis I and II are the most common mechanisms by which diploid spermatozoa are produced. Endoreduplication before meiosis has been suggested as a possible origin for tetraploid meiocytes, which might, in turn, produce diploid sperm. Synaptonemal complex (SC) spreads of a fertile man were immunolabelled (SCP3, MLH1 and CENP) and hybridized with subtelomere-specific multiplex fluorescent in situ hybridization (stM-FISH) assay for SCs identification. The unexpected finding of a tetraploid pachytene cell and the identification of all of its SCs demonstrate that synapsis and crossover events can occur in human tetraploid cells. Moreover, it indicates that diploid sperm may also originate from mitotic errors (endoreduplication) occurring before meiosis.

Sperm nuclei analysis of 1/29 Robertsonian translocation carrier bulls using fluorescence in situ hybridization

In 1964, Gustavsson and Rockborn first described the 1/29 Robertsonian translocation in cattle. Since then, several studies have demonstrated the negative effect of this particular chromosomal rearrangement on the fertility of carrier animals. During the last decade, meiotic segregation patterns have been studied on human males carrying balanced translocations using FISH on decondensed sperm nuclei. In this work, we have applied the ‘Sperm-FISH’ technique to determine the chromosomal content of spermatozoa from two bulls heterozygous for the 1/29 translocation and one normal bull (control). 5425 and 2702 sperm nuclei were scored, respectively, for the two heterozygous bulls, using whole chromosome painting probes of chromosomes 1 and 29. Very similar proportions of normal (or balanced) spermatozoa resulting from alternate segregation were observed (97.42% and 96.78%). For both heterozygous bulls, the proportions of nullisomic and disomic spermatozoa did not follow the theoretical 1:1 ratio. Indeed, proportions of nullisomic spermatozoa were higher than those of disomic sperma tozoa (1.40% vs 0.09% (bull 1) and 1.29% vs 0.15% (bull 2) for BTA1, and 0.65% vs 0.40% (bull 1) and 1.11% vs 0.63% (bull 2) for BTA29). The average frequencies of disomic and diploid spermatozoa in the normal bull were 0.11% and 0.05%, respectively.