First trimester pregnancy outcomes in a large IVF center from the Lombardy County (Italy) during the peak COVID-19 pandemic.

At the beginning of 2020, the Italian Lombardy region was hit by an "epidemic tsunami" which was, at that point in time, one of the worst pandemics ever. At that moment the effects of SARS-COV 2 were still unknown. To evaluate whether the pandemic has influenced ART (Assisted Reproduction Techniques) outcomes in an asymptomatic infertile population treated at one of the major COVID-19 epicentres during the weeks immediately preceding lockdown. All ART procedures performed during two time periods were compared: November 1st, 2018 to February 28th, 2019 (non-COVID-19 risk) and November 1st, 2019 to February 29th, 2020 (COVID-19 risk). In total 1749 fresh cycles (883 non-COVID-19 risk and 866 COVID-19 risk) and1166 embryos and 63 oocytes warming cycles (538 and 37 during non-COVID and 628 and 26 during COVID-19 risk, respectively) were analysed. Clinical pregnancies per cycle were not different: 370 (25.38%) in non-COVID versus 415 (27.30%) (p = 0.237) during COVID-19 risk. There were no differences in biochemical pregnancy rates 52 (3.57%) versus 38 (2.50%) (p = 0.089) nor in ectopic pregnancies 4 (1.08%) versus 3 (0.72%) (p = 0.594), spontaneous miscarriages 84 (22.70%) versus 103 (24.82%) p = 0.487, nor in intrauterine ongoing pregnancies 282 (76.22%) versus 309 (74.46%) p = 0.569. A multivariate analysis investigating differences in spontaneous miscarriage rate showed no differences between the two timeframes. Our results support no differences in asymptomatic infertile couples' ART outcomes between the pre COVID and COVID-19 periods in one of the earliest and most severe pandemic areas.

One step further: randomised single-centre trial comparing the direct and afterload techniques of embryo transfer.

STUDY QUESTION: What are the differences in ease of use between two different embryo transfer (ET) techniques: the preload direct approach and the afterload approach. SUMMARY ANSWER: The afterload technique seems to reduce the rate of difficult ETs. WHAT IS KNOWN ALREADY: Numerous published trials now document that the ET procedure has an impact on pregnancy and delivery rates after IVF. Difficult transfers should be avoided, as they reduce implantation and pregnancy rates. Preload direct ETs with soft catheters under ultrasound guidance is currently considered the best procedure. However, when using soft catheters, it is not known which technique is preferable or which one should be implemented to reduce the operator factor. STUDY DESIGN, SIZE, DURATION: This prospective randomised unblinded controlled clinical trial, included 352 ultrasound-guided ETs assigned to either direct ET or afterload ET, between September 2017 and March 2019. The sample size was calculated based on the historical rate of difficult ETs encountered between 2014 and 2015 with a direct ET procedure. PARTICIPANTS/MATERIALS, SETTING, METHODS: The inclusion criteria were women 18-38 years old, with BMI between 18 and 28, receiving a single-thawed blastocyst transfer. The exclusion criteria were use of testicular sperm and preimplantation genetic testing (PGT) cycles. The primary outcome was the rate of difficult or suboptimal transfers defined as: advancement of the outer sheath (specific for the direct transfer), multiple attempts, use of force, required manipulation, use of a stylet or tenaculum, dilatation, or use of a different catheter. The secondary outcome was clinical pregnancy rate. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 352 frozen ETs were randomised, with 176 patients in each group. The two arms were homogeneous for female and male age, female BMI, duration of infertility, secondary infertility, previous deliveries or miscarriages, myomas, previous surgery to the uterine cavity, cycle day at ovulation trigger, freeze all cycles, first transfers, indication for treatment, endometrial preparation protocol and duration, endometrial thickness, and blastocyst grade at vitrification. Across the entire population, 85 (24.1%) ETs were defined as difficult. The rate of difficult transfers was significantly higher in the direct ET group than in the afterload group: 68 (38.6%) versus 17 (9.7%), respectively (OR 0.17, 95% CI 0.09-0.30, P < 0.001). The mean percentage in the rate of difficult transfers per operator was 22.5% (SD ± 14.5%), of which 36.1% (SD ± 23.4%) were in the direct group compared with 8.6% (± 8.2%) in the afterload group (P < 0.001). The difficult transfer rate among operators varied from 0 to 43.8% (0-77.8% in the direct group and 0 to 25.0% in the afterload group). The clinical pregnancy rates (42.0% vs 48.3%, P = 0.239 in the direct and afterload groups, respectively) were not significantly different between the groups. LIMITATIONS, REASONS FOR CAUTION: There were 18 experienced operators who participated in the trial. Conclusions about the pregnancy rate should not be generalised, since the sample analysis was not performed on this outcome and, although clinically relevant, the difference was not significantly different. WIDER IMPLICATIONS OF THE FINDINGS: The rate of difficult transfers was significantly higher in the direct ET group compared with the afterload ET group, although a wide variation was observed among operators. Further studies regarding the association between transfer technique and ART outcomes are required. STUDY FUNDING/COMPETING INTEREST(S): No specific funding was sought and there are no competing interests. TRIAL REGISTRATION NUMBER: NCT03161119. TRIAL REGISTRATION DATE: 5 April 2017. DATE OF FIRST PATIENT'S ENROLMENT: 26 September 2017.

Comprehensive protocol of traceability during IVF: the result of a multicentre failure mode and effect analysis.

STUDY QUESTION: Can traceability of gametes and embryos be ensured during IVF? SUMMARY ANSWER: The use of a simple and comprehensive traceability system that includes the most susceptible phases during the IVF process minimizes the risk of mismatches. WHAT IS KNOWN ALREADY: Mismatches in IVF are very rare but unfortunately possible with dramatic consequences for both patients and health care professionals. Traceability is thus a fundamental aspect of the treatment. A clear process of patient and cell identification involving witnessing protocols has to be in place in every unit. To identify potential failures in the traceability process and to develop strategies to mitigate the risk of mismatches, previously failure mode and effects analysis (FMEA) has been used effectively. The FMEA approach is however a subjective analysis, strictly related to specific protocols and thus the results are not always widely applicable. To reduce subjectivity and to obtain a widespread comprehensive protocol of traceability, a multicentre centrally coordinated FMEA was performed. STUDY DESIGN, SIZE, DURATION: Seven representative Italian centres (three public and four private) were selected. The study had a duration of 21 months (from April 2015 to December 2016) and was centrally coordinated by a team of experts: a risk analysis specialist, an expert embryologist and a specialist in human factor. Principal investigators of each centre were first instructed about proactive risk assessment and FMEA methodology. A multidisciplinary team to perform the FMEA analysis was then formed in each centre. After mapping the traceability process, each team identified the possible causes of mistakes in their protocol. A risk priority number (RPN) for each identified potential failure mode was calculated. The results of the FMEA analyses were centrally investigated and consistent corrective measures suggested. The teams performed new FMEA analyses after the recommended implementations. PARTICIPANTS/MATERIALS, SETTING, METHODS: In each centre, this study involved: the laboratory director, the Quality Control & Quality Assurance responsible, Embryologist(s), Gynaecologist(s), Nurse(s) and Administration. The FMEA analyses were performed according to the Joint Commission International. MAIN RESULTS AND THE ROLE OF CHANCE: The FMEA teams identified seven main process phases: oocyte collection, sperm collection, gamete processing, insemination, embryo culture, embryo transfer and gamete/embryo cryopreservation. A mean of 19.3 (SD ± 5.8) associated process steps and 41.9 (SD ± 12.4) possible failure modes were recognized per centre. A RPN ≥15 was calculated in a mean of 6.4 steps (range 2-12, SD ± 3.60). A total of 293 failure modes were centrally analysed 45 of which were considered at medium/high risk. After consistent corrective measures implementation and re-evaluation, a significant reduction in the RPNs in all centres (RPN <15 for all steps) was observed. A simple and comprehensive traceability system was designed as the result of the seven FMEA analyses. LIMITATIONS, REASONS FOR CAUTION: The validity of FMEA is in general questionable due to the subjectivity of the judgments. The design of this study has however minimized this risk by introducing external experts for the analysis of the FMEA results. Specific situations such as sperm/oocyte donation, import/export and pre-implantation genetic testing were not taken into consideration. Finally, this study is only limited to the analysis of failure modes that may lead to mismatches, other possible procedural mistakes are not accounted for. WIDER IMPLICATIONS OF THE FINDINGS: Every single IVF centre should have a clear and reliable protocol for identification of patients and traceability of cells during manipulation. The results of this study can support IVF groups in better recognizing critical steps in their protocols, understanding identification and witnessing process, and in turn enhancing safety by introducing validated corrective measures. STUDY FUNDING/COMPETING INTEREST(S): This study was designed by the Italian Society of Embryology Reproduction and Research (SIERR) and funded by the Italian National Transplant Centre (CNT) of the Italian National Institute of Health (ISS). The authors have no conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.

Preliminary evidence for high anti-PLAC1 antibody levels in infertile patients with repeated unexplained implantation failure.

OBJECTIVE: Placenta-specific1 (PLAC1) is a trophoblast-specific gene encoding for a protein that is highly expressed in human placenta, on the surface of the syncytiotrophoblast. PLAC1 was found to elicit spontaneous antibody responses in cancer patients. We aimed to determine the levels of anti-PLAC1 antibodies in infertile women with a history of unexplained repeated implantation failure after IVF cycles as compared to fertile women. STUDY DESIGN: An observational case-control clinical study. MAIN OUTCOME MEASURE(S): Two groups of patients were analysed in two different experimental settings: 21 infertile women and 81 control patients were enrolled in the first group, 16 infertile women and 67 fertile controls in the second group. Anti-PLAC1 antibody levels and ranking were analysed by ELISA test. RESULTS: In both groups of infertile patients enrolled, optical densities (OD) from ELISA test ranked significantly higher than those of controls (0.27 ± 0.2 vs. 0.13 ± 0.1 respectively; p = 0.0009 in the first group), (0.62 ± 0.38 vs. 0.39 ± 0.35 respectively; p = 0.0044 in the second experiment). In the first group about one case in four (29%) had OD levels above the 95th percentile (0.337) for healthy controls (p = 0.005). In the second experiment 4 out of 16 cases (25%) had OD levels above the 95th percentile (0.878) for healthy controls (p = 0.023). CONCLUSIONS: Anti-PLAC1 antibodies could represent a biomarker associated with infertility and with high probability of repeated implantation failure after ovarian stimulation and IVF-ET, greatly improving the diagnostic work up of infertile couples.

Five years (2004-2009) of a restrictive law-regulating ART in Italy significantly reduced delivery rate: analysis of 10,706 cycles.

STUDY QUESTION: Was the delivery rate of ART cycles negatively affected by the enactment of the Law 40/2004 by the Italian Parliament which imposed a long list of restrictions for ART procedures? SUMMARY ANSWER: This large and extensive comparative analysis of ART outcomes prior to and after the introduction of the Law 40 revealed a significant reduction in pregnancy and delivery rates per cycle, independent of age or other clinical variables, once the law went into effect. WHAT IS KNOWN ALREADY: Several studies have been published on the effect of Law 40/2004 on ART outcomes, some authors demonstrating a negative impact of the Law in relation to specific etiologies of infertility, other authors showing opposite conclusions. STUDY DESIGN, SIZE, DURATION: Retrospective clinical study of 3808 patients treated prior to the enactment of the Law, September 1996-March 2004 (Group I) and 6898 treated during the Law, March 2004-May 2009 (Group II). PARTICIPANTS/MATERIALS, SETTING, METHODS: A total of 10 706 ART cycles were analysed, 3808 performed before and 6898 after the application of the Law. An intention-to-treat statistical analysis was performed to detect pregnancy and delivery rates (pregnancies ≥ 24 weeks) per started cycle. A P value of <0.05 was considered statistically significant. We analysed different outcomes: differences in fertilization, pregnancy and delivery rate, multiple pregnancies and miscarriage rates between the two time periods. MAIN RESULTS AND THE ROLE OF CHANCE: The delivery rate for started cycle was 20% before and 16.0% after the introduction of the Law representing a 25% reduction (P < 0.001). The multivariate analysis, corrected by female age of >38 years, duration of infertility, basal FSH level and number of retrieved oocytes, showed a 16% lower delivery rate (odds ratio: 0.84; confidence interval: 0.75-0.94). This statistical approach removed the risk that the observed effects were due to chance and confirmed unequivocally that the Law was an independent factor responsible for the reduced likelihood of a successful outcome. LIMITATIONS, REASONS FOR CAUTION: This is a retrospective study. A prospective randomized study, with patients treated in the same time period and randomized to restrictions or not, would have minimized potential limitations due to differences in years of treatments. WIDER IMPLICATIONS OF THE FINDINGS: Our findings based on the analysis of such a large number of cycles proved clearly and unequivocally that imposing restrictions on the practice of ART penalized patients. These data represent a relevant clinical contribution for countries still debating the enactment of restrictive limitations of ART.

Italian Constitutional Court modifications of a restrictive assisted reproduction technology law significantly improve pregnancy rate.

BACKGROUND: In May 2009, the Italian Constitutional Court banned most of the limitations of a restrictive law regulating assisted reproduction technology on the grounds that it limited a couple's right to have access to the best possible medical treatment and reduce any possible higher risk of complications. The aim of the study was to compare our results in fresh cycles before and after this change. MATERIALS AND METHODS: We analysed retrospectively 3274 IVF cycles: 2248 before and 1026 after the law was modified. RESULTS: There was no significant difference between the two groups in terms of age, basal FSH levels, years of infertility, the number of previous cycles or the number of oocytes retrieved but the number of oocytes used (2.7 ± 0.6 versus 4.6 ± 1.8; P = <0.001), the number of embryos obtained (2.0 ± 0.9 versus 3.3 ± 1.8; P = <0.001) and transferred (2.2 ± 0.7 versus 2.3 ± 0.7; P = <0.001) were all higher after the removal of the previous restrictions, as was the pregnancy rate per started cycle (23.49% versus 20.42%; P = 0.047). Before modification of the law, the pregnancies were single in 74.11% of the cases (versus 71.43% afterwards), twins in 23.44% (versus 26.89%; P = 0.318) and triplets in 2.46% (versus 1.68%; P = 0.594). CONCLUSIONS: Our preliminary results after the removal of the previous legal restrictions show a higher pregnancy rate per started cycle (3.7% represents a 15% difference) and a positive (albeit non-significant) trend towards a reduction in the number of multiple pregnancies.

Oocyte cryopreservation.

Over the last few years, there has been renewed interest and scientific debate concerning human oocyte cryopreservation. The aim of this study was to analyse the clinical data coming from our long experience of slow-freezing oocytes. Between 2001 and 2007, 1280 thawing cycles were carried out using oocytes previously frozen by means of a slow 1,2 propaniedol+sucrose protocol. A total of 7585 oocytes were thawed, of which 4409 survived and 3622 were microinjected; 144 clinical pregnancies were obtained. The number of thawing cycles increased from 19 in 2001 to 268 in 2007, and the number of thawed oocytes from 197 to 1652. Although the survival rate was significantly lower in the period 2002-2005 than in the period 2006-2007, pregnancy and implantation rates steadily improved from respectively 6.7% and 2.4% in 2001 to 15% and 8.2% in 2007. Our data demonstrate a clinically important improvement in oocyte crypreservation over the years in a Centres with proved experience, and can be offered as a standard of care not only before cancer treatment but also for couples refusing embryo crypreservation or in countries with very restrictive limitations on embryo or zygote freezing.

Cancer risk in male factor-infertility.

Severe forms of male-factor infertility are associated with an increased risk of testicular cancer and scrotal ultrasonography is widely used for diagnosis. In this study, 2172 male members of infertile couples referred to our Reproductive Medicine Unit were submitted to scrotal ultrasonography and 835 selected patients had been followed during a 2-year period. Eight out of nine neoplastic nodules found at the initial examination were unpalpable and discovered by ultrasonography. Ten tumoral lesions were found in 370 testicular biopsies performed for diagnostic purposes or to extract spermatozoa; and eight additional neoplastic lesions were discovered during the 2-year follow-up of 835 patients. The cumulative rate of neoplastic disease was 3.2%. Thirteen cases (1.5%) were malignant (12 germ cell tumours and one non-Hodgkin lymphoma of testicular origin); the remaining 14 were benign forms (Leydig cell tumours and hyperplasias, Sertoli cell nodules, adenomatoid tumours). Testicular volume (cut-off: 12ml) resulted weakly correlated with germ cell cancer (p=n.s., odds ratio 2.01) while low total sperm count (<40x10(6)) (p=0.002, odds ratio 8.4), previous cryptorchidism (p=0.04, odds ratio 7.5) and hypergonadotrophic hypogonadism (p=0.04, odds ratio 7.9) were associated with an increased risk. But a stronger correlation with germ cell cancer was found in the patients with some utrasonographic anomalies, i.e. testicular microlithiasis (p=0.0015, odds ratio 37.1) or larger calcifications not fitting the description of testicular microlithiasis (p<0.0001, odds ratio 69.5). Our findings indicate that scrotum ultrasonography should always be advised in subfertile men with <40x10(6) spermatozoa/ejaculate or hypergonadotrophic hypogonadism or previous cryptorchidism, and that particular care should be taken in the presence of testicular microlithiasis or testicular calcifications. These men should be aware of the existence of higher risk of testicular cancer and trained in testicular self-examination.

Cryopreservation of supernumerary oocytes in IVF/ICSI cycles.

BACKGROUND: The aim of the present study is to investigate cryopreservation of oocytes in patients refusing embryo cryopreservation for ethical reasons, patients from whom no sperm could be retrieved and patients with enough oocytes to yield a number of fresh and cryopreserved embryos to transfer. METHODS: A total of 2900 oocytes out of 6216 retrieved were cryopreserved in 286 patients undergoing 303 cycles. The reasons for cryopreservation were because no sperm was found in 16 cycles, for ethical or personal reasons in 80, and in 207 only supernumerary oocytes were frozen. In 159 cycles, the oocytes were thawed and the surviving metaphase II oocytes microinjected. RESULTS: A total of 1087 oocytes were thawed, 760 (69.9%) survived and 687 were microinjected. We obtained 368 (53.5%) normally cleaved embryos, 331 were transferred and 37 were cryopreserved. One hundred and forty-five transfers (range 1-3 embryos/patient) were performed and 18 (12.4%) pregnancies were obtained. Twelve patients delivered 13 healthy children, and six first trimester abortions were observed (33.3%). CONCLUSION: Although a low implantation rate was observed and a higher abortion rate than in fresh cycles, our results show that in sibling oocytes, the process of cryopreservation apparently does not affect the fertilization and cleavage rate. In this group of patients, producing a large number of mature gametes, oocyte cryopreservation gives the couple extra chances to achieve a pregnancy within a single retrieval and is a good effort towards reducing the number of embryos cryopreserved and enhancing our experience in this new technology.

Outcome of assisted reproductive technologies after different embryo transfer strategies.

The outcomes of 1028 assisted reproductive technology cycles were studied retrospectively, considering two different periods of embryo transfer. In the first period, 262 cycles in women < 36 years old were studied, in which three embryos were transferred, followed by 157 cycles in women > or = 36 years, in which four embryos were transferred. In the second period, 332 cycles were evaluated in women < 36 years and 277 cycles in women > or = 36 years old, reducing the number of embryos transferred to two and three respectively. Embryos were only scored morphologically, and the best embryos were chosen for transfer. In the first period, in women < 36 years old, a clinical pregnancy rate of 55.7% was achieved, compared with 42.5% in the second period (P < 0.01). In women > or = 36 years old, the first period of embryo transfer showed a clinical pregnancy rate of 39.5%, compared with 28.5% in the second period (P < 0.01). The number of twin pregnancies in the three groups of patients in whom one to four embryos were transferred was not significantly different (24.2, 28.4, 24.8%). It is concluded that even with the biases induced by a retrospective study, the reduction in the number of embryos transferred, from three to two in women < 36 years of age, and from four to three in women > or = 36 years of age, without any selection other than pre-transfer morphological score, adversely affects the outcome of treatment, without a significant reduction in twin gestation rate. Other strategies are to be implemented in gametes and embryo selection, and patients must be aware that, even with a reduction in pregnancy rate, the goal is to achieve a high cumulative pregnancy rate, reducing the complications induced by multiple pregnancies.

Implantation failure in assisted reproduction technology and a critical approach to treatment.

In this article, we review the literature and our personal experience regarding the many factors that appear to influence implantation rate. Oocyte quality, as determined by patient age and aneuploidies, probably plays a major role in RIF. However, a panoply of other factors have been brought under investigation, quite often with contradictory results and additional intriguing questions to be studied. Infections of the vagina, cervix, and endometrium, the role of mucus aspiration and washing of the cervix on transfer, the role of catheter guidance for a correct transfer and potion of embryos, the effect of mock transfer, and the role of hysteroscopy and its timing before transfer procedures are analyzed both as a review of the literature and as opinions and data from our experience. Many of these factors are interlaced and from the apparently simple issue of trauma, to infections and immune modulation of hatching and implantation, a biological continuum can easily be identified. The impact of abnormalities of the immune system and of homeostasis abnormalities is also covered in a brief overview of reported works and our experience. These latter areas probably constitute the common biological background of all other external factors that, however, the skilled must equip themselves for improving implantation success.

Testicular sperm extraction in azoospermic men submitted to bilateral orchidopexy.

BACKGROUND: This study was carried out to evaluate whether bilateral orchidopexy represents a poor or good prognostic factor in azoospermic men undergoing testicular sperm extraction (TESE). METHODS: One hundred and seven presumed non-obstructive azoospermia (NOA) patients, according to conventional clinical parameters (volume of testis, FSH, clinical history) were submitted to testicular biopsy with TESE. Thirty men (28%) had a history of bilateral orchidopexy for cryptorchidism. RESULTS: Normal spermatogenesis or mild hypospermatogenesis was diagnosed in 12/30 ex-cryptorchid patients and in 7/77 presumed NOA patients (P = 0.0004). Conversely, pure Sertoli cell-only syndrome or complete maturation arrest was found in 10/30 ex-cryptorchid patients and in 48/77 presumed NOA patients (P = 0.0094). In 53/107 patients (49.5%), TESE allowed a positive sperm retrieval. At least one spermatozoon was observed in 22/30 ( approximately 73%) ex-cryptorchid patients and in 31/77 ( approximately 40%) presumed NOA patients (P = 0.0026). A large number of spermatozoa (equivalent to an obstructive pathology) were retrieved in 13/30 ex-cryptorchid and in 10/77 presumed NOA patients (P = 0.001). A history of bilateral orchidopexy in presumed NOA patients correlates positively for the chance of retrieving testicular spermatozoa (odds ratio 3.8; 95% confidence interval 1.41-10.21; P = 0.008). CONCLUSIONS: Although bilateral cryptorchidism is usually considered a testicular secretive dysfunction, TESE permits retrieval of a large number of spermatozoa in almost 40% of cases. Our data suggest the existence of congenital or acquired obstructive anomalies of the seminal ducts in azoospermic orchidopexed men.

The impact of embryo transfer on implantation--a review.

Embryo transfer has received little clinical attention and has been, until recently, the most inefficient step in in-vitro fertilization (IVF). In this article, the authors review the literature and their personal experience regarding the process of intrauterine transfer of embryos, which remains the object of much discussion. Factors which appear to influence implantation rates are: contamination of the catheter tip with cervical bacteria, stimulation of uterine contractions during the procedure, the type of catheter, ultrasound guidance during the transfer, and the position of the embryos in the uterine cavity. Easy and atraumatic transfer is essential for successful implantation and the embryos need to be placed in the middle of the cavity, away from the fundus. Knowing, beforehand, the position and length of the uterus can provide better results and may reduce the rate of ectopic pregnancies. Evidence from randomized studies has supported this claim. Despite the number of available studies controlling certain variables, most authors, even using the same catheter, ultrasound guidance and/or a trial transfer use different protocols or similar instruments in different ways. Standardization of the transcervical intrauterine transfer of embryos in a large randomized study is needed before definitive conclusions can be drawn. The goal of improved implantation and pregnancy rates deserve these efforts.

Aspermia and chronic testicular pain after imperforate anus correction. Cryopreservation of sperm cells extracted from whole orchiectomized testis: case report.

This paper describes an unusual association of aspermia and untreatable, chronic testicular pain in a young man who underwent 14 surgical interventions for an imperforate anus. Physical examination and ultrasonography revealed left epididymal and vas enlargement, normal-sized testes, tubular ectasia of the left rete testis and a small intraprostatic paramedian left cyst. Retrograde ejaculation and urogenital infections were excluded, and the FSH and karyotype results were normal. The patient gave his consent to an exploratory intervention with possible radical left orchiectomy. The patency of the left distal seminal duct was unexpectedly normal, and no sperm were found in the epididymis or vas deferens despite their obstructive appearance. Sperm were only found in a 'testicular touch' preparation. The removed testis was immediately opened and most of the testicular lobules were removed, thus allowing the extraction of 25 x 10(6) sperm, which were cryopreserved in 35 straws. An 8-month follow-up examination documented the complete absence of pain and, during the next few months, it is planned to use the thawed sperm for ICSI. Radical orchiectomy plus the cryopreservation of sperm extracted from the whole testis must be considered in the case of the co-existence of chronic unilateral testicular pain and aspermia.

Insemination of HIV-negative women with processed semen of HIV-positive partners.

Many HIV-discordant couples want to have children so much that they are willing to abandon condom-protected sexual intercourse irrespective of the risks. Previous testing in our laboratory showed that gradient centrifugation followed by a swim-up procedure effectively removed HIV-1-infected cells from the semen of HIV-seropositive men. 85 HIV-discordant couples were screened for fertility; 29 women were found suitable for a timed insemination course with the processed semen of their HIV-seropositive partner. None of the inseminated women seroconverted, and 17 pregnancies were achieved in 15 women. All 10 babies born to these mothers remain HIV seronegative. The findings may help in the counselling of such couples and also give them hope of having healthy babies.