Detection of HIV-1 RNA in seminal plasma samples from treated patients with undetectable HIV-1 RNA in blood plasma.

Five percent of 145 HIV-1 infected men enrolled in an assisted reproductive technology (ART) program harbored detectable HIV-1 RNA in semen, although they had no other sexually transmitted disease and their blood viral load was undetectable for at least 6 months under antiretroviral treatment. This result justifies measuring HIV-1 RNA in semen before the ART process and suggests that a residual risk of transmission has to be mentioned to the patients who would like to have unprotected sexual intercourse.

Comparison of three in vitro culture systems for maturation of early preantral mouse ovarian follicles.

The aim of this study was to compare three different culture systems for in vitro follicular growth and oocyte maturation in ovarian follicles of mice in order to assess the technique with the optimal growth and improved rate of meiotic maturation. The three systems tested were culture under oil, on a hydrophobic membrane and on agar respectively. Early preantral follicles were cultured for 12 days in alpha-MEM GlutaMAX medium. Follicular growth, oocyte meiotic maturation, oocyte extrusion, atresia and estradiol production were analysed. Follicular development showed two phases in the three systems, with slow growth before day 5 and subsequent acceleration. The percentage of follicles transferred into oocyte maturation medium was significantly higher after culture under oil. The proportion of oocytes that achieved nuclear maturation (metaphase II) was higher when follicles were cultured under oil or on a hydrophobic membrane than on agar. Our results support the use of culture under oil for in vitro follicular growth from the early preantral stage in order to obtain metaphase II oocytes. Fertilization ability of these oocytes and the capacity to obtain healthy mice in a reproducible manner warrants further investigation.

Molecular cytogenetics analysis with whole chromosome paint probes of sperm nuclei from a (13;15) Robertsonian translocation carrier.

Meiotic segregation of a Robertsonian translocation (13;15) was assessed in sperm nuclei using dual-color fluorescent in situ hybridization (FISH) with whole-chromosome paint probes. Most spermatozoa in the (13;15) translocation carrier resulted from alternate segregation. Although an increased frequency of unbalanced gametes was observed, spontaneous pregnancy led to the birth of a boy with a normal karyotype.

Primary flagellar abnormality is associated with an increased rate of spermatozoa aneuploidy.

The frequencies of aneuploid and diploid spermatozoa were determined in 3 patients presenting a complete asthenozoospermia due to a primary and specific flagellar anomaly: patients 1 and 2 presented a "stump tail syndrome," more than 50% of spermatozoa with a short flagella, patient 3 had a Kartagener syndrome including situs inversus, sinusitis, and bronchiectassis. No pregnancy was obtained after 3 intracytoplasmic sperm injection (ICSI) attempts in patients 1 and 2. A 3-color fluorescence in situ hybridization analysis was performed on their spermatozoa using centromeric probes for chromosomes X, Y, and 18 and compared with those of 8 fertile males. The frequency of disomic 18 and hyperhaploid XY spermatozoa was not significantly increased in the 3 patients when compared with controls. However, the 3 patients showed elevated frequencies of XX, YY, and diploid spermatozoa. These data add to growing evidence that systematic sperm anomalies of flagella increase the rate of spermatozoa aneuploidy and may also reduce the chances of pregnancy after intracytoplasmic sperm injection.

Comparison of six density gradient media for selection of cryopreserved donor spermatozoa.

The aim of our study was to evaluate the efficiency of 4 density gradient media for motile cryopreserved spermatozoa selection to Percoll (Kabi Pharmacia, Uppsala, Sweden) and to Puresperm (J.C.D. International Laboratory, L'Aigle, France). Puresperm was the new medium chosen in our laboratory in 1996 as the substitute for Percoll. The solutions tested were 3 colloidal silane-coated silica particle media (Isolate, SpermGrad-100, Sil-Select Plus) and iodixanol (Optiprep). Semen parameters analyzed after selection were concentration, motility, and morphology. Semen parameters after Puresperm gradient had similar values compared to Percoll. Optiprep was less efficient with a poor concentration. Isolate had a comparatively better concentration, but the capacity of selection was not satisfactory. SpermGrad-100 and Sil-Select Plus were less effective than Puresperm. In conclusion, Puresperm could be considered a better alternative to Percoll for cryopreserved spermatozoa migration.

Fluorescence in situ hybridisation (FISH) analysis of chromosome segregation and interchromosomal effect in spermatozoa of a reciprocal translocation t(9,10)(q11;p11.1) carrier.

A couple was referred for exploration of repetitive abortions. The man was found to be a carrier of a balanced reciprocal translocation t(9;10)(q11;p11.1). The meiotic segregation of chromosomes 9 and 10 was analysed in 5,157 spermatozoa from this translocation carrier and in 15,255 spermatozoa from three control donors using three-colour fluorescence in situ hybridisation (FISH). The theoretical viability of the different segregation patterns was performed using the computer system HC Forum developed by the Department of Cytogenetics at the Grenoble University Medical School, La Tronche, France. A normal or balanced constitution was found in 56.25% of the analysed spermatozoa. The tertiary 3:1 segregation mode was the most frequently observed (14.37%). The frequencies of adjacent-1, adjacent-2 and 3:1 interchange modes were 12.85, 9.38 and 7.14% respectively. The cumulative frequency of non-viable imbalance was estimated at 20.91% according to the theorical viability of the different segregation patterns. Spermatozoa aneuploidy frequency was also evaluated for chromosomes X, Y and 18, and there was no evidence of interchromosomal effect in spermatozoa from the translocation carrier. FISH analysis of spermatozoa in combination with the viability theorical estimation of the different segregation patterns could be considered a useful tool for genetic counselling in carriers of reciprocal translocation.