Preimplantation Genetic Diagnosis Results Research Articles

The impact of pronuclear orientation to select chromosomally normal embryos

To find an effective method to select embryos from ICSI with better chromosomal status when preimplantation-genetic-diagnosis (PGD) is not applied. Several morpholological evaluations were done in same embryos. Embryos from ICSI were classified on day 1 according to pronuclear-nucleoli arrangement. On day 3, classification was done according to number, fragmentation, size and shape of cells. In some patients, embryos exhibiting good quality on day 3 (at least six regular blastomeres with cell fragmentation lower than 20%) were also submitted to PGD, irrespective to pronuclear-nucleoli morphology. Forty-two per cent of normally fertilized embryos showed pronuclear-nucleoli-good-morphology; from those, 86% were also classified as good quality on day 3. Good-quality embryos submitted to PGD have shown lower chromosomal abnormality rates when also classified as pronuclear-nucleoli-good-morphology. Pronuclear-nucleoli morphology seems to be correlated with PGD results. This criterion may prove useful for pre-select embryos with normal chromosomal package when PGD is not applied.

Preimplantation genetic diagnosis for β‐thalassaemia: the Sardinian experience

AbstractObjectivesTo report the experiences on preimplantation genetic diagnosis (PGD) in couples at risk for ß‐thalassaemia in Sardinia.Methods23 couples at risk for ß‐thalassaemia were included in the PGD programme with a total of 42 cycles performed. Among these, 11 couples were fertile, while the remaining 12 had associated fertility problems. In vitro Fertilization (IVF), PGD and prenatal genetic molecular confirmation protocols and results are reported.ResultsAll the patients followed the protocol of ovarian stimulation, oocyte retrieval, intracytoplasmic sperm injection (ICSI), embryo biopsy and genetic analysis. A total of 272 oocytes were fertilized in the regular way, and embryo biopsy was performed on 202 embryos. Out of these 202 embryos, 192 (95%) were successful. The genetic diagnosis was performed on 150 embryos (78.1%). Ninety‐eight were identified as unaffected and 75 were transferred in 31 cycles. In the infertile patient group, two biochemical pregnancies (11.1% per transfer), in the fertile patient group, four clinical pregnancies, two twin and two singleton pregnancies (30.8% per transfer), were obtained. The genetic molecular results were confirmed in all pregnancies by first‐trimester chorionic villus sampling (CVS).ConclusionOur study shows that PGD for ß‐thalassaemia is an available procedure for couples who wish to avoid termination of pregnancy, except in cases where the IVF cycle efficiency is very poor. Copyright © 2004 John Wiley & Sons, Ltd.

Outcome of preimplantation genetic diagnosis in frozen-thawed embryos

Cryopreservation of embryos is widely applied in ART. And it is also useful for efficient preimplantation genetic diagnosis (PGD) in cases with supernumerary embryos or with collected embryos through several cycles in poor responders. The aim of this study is to evaluate the feasibility of FISH or PCR in the blastomeres biopsied from frozen-thawed embryos and the outcome of PGD, and to determine whether frozen- thawing process affect the embryo development after blastomere biopsy. Retrospective clinical study. In 86 patients, 319 PGD cycles were performed between Jan. 2000 and Dec. 2003. Patients were divided into 2 groups: thawed PGD group − PGD cycle was performed in frozen-thawed embryos (33 patients, 40 cycles, age = 34.2±4.7yrs. (mean±SD)), fresh PGD group − PGD cycle was performed in fresh embryos (patients=53, cycles=279, mean age = 31.8±4.3yrs.). In 20 cycles, supernumerary embryos were frozen and thawed, and in 19 cycles, embryos were collected through several cycles in poor responders and then thawed altogether. PGD was performed for translocation carriers or aneuploidy screening and some single gene diseases were included. Ovarian stimulation was done using GnRH agonist/FSH long protocol or clomifen citrate/GnRH antagonist protocol in poor responders. Oocytes were inseminated by ICSI and the fertilized embryos were cryopreserved at PN stage or cleaving embryo stage. In thawed embryo transfer cycles, estradiol was administrated from day 2 of menstrual cycle and progesterone injection was started from 1 day before 2PN thawing. The cryopreserved embryos were thawed and blastomeres were biopsied in survived embryos developed to 6–10 cell stage. PGD was performed by using FISH or PCR and the embryos were replaced 3 days after PN thawing. The survival rate of the thawed embryos, the rate of successful diagnosis for FISH or PCR, embryo development and pregnancy rate were compared between thawed and fresh PGD cycles. The survival rate of thawed embryos was 71.2±29.8% (mean±SD). The FISH or PCR analysis was successful in 88.1±22.0 % in thawed PGD group and 95.2±10.3% in fresh PGD group (P>0.05). The proportion of transferable embryos in thawed PGD group was similar to fresh PGD group (21.6±19.7% vs. 27.4±18.8%; P>0.05). The rate of embryos developed cleavage after biopsy was also similar in both groups (62.0±26.5% vs. 66.4±26.1%; P>0.05). The clinical pregnancy rate per transfer was 11.4 % (4/35) in thawed PGD group and 21.3% (57/267) in fresh PGD group. Although it was not statistically significant, the pregnancy rate tends to be lower in thawed PGD group. Our data suggest that blastomere biopsy and analysis with FISH or PCR is feasible in frozen-thawed embryos if the embryos survive well. The embryo development was not affected by biopsy in thawed embryos, but the pregnancy rate was lower in thawed PGD cycles. Therefore PGD in thawed embryos could be applied usefully in cycles with supernumerary embryos or collected cyropreserved embryos, and optimization of freezing-thawing condition might be important.

Predictive value of preimplantation genetic diagnosis of aneuploidy and translocations

To ascertain if the results of a Preimplantation Genetic Diagnosis (PGD) cycle are predictive for the next. This is important for those patients that fail to conceive in the first cycle due to high rates of chromosomally abnormal embryos. Retrospective study. 152 patients from our database of 2500 cycles from 71 IVF centers were selected following these criteria: a) patients with have two or more cycles in less than two years, more than two years past between the first and second cycle could increase the proportion of abnormal embryos due to age; b) patients which do not involve polar body biopsy considering that post-zygotic abnormalities will not be observed; c) patients which do not involve PGD for single gene defects because the number of repeated cycles was too low for a meaningful analysis; d) patients with more than four embryos analyzed in their cycles, because small cohorts of embryos are difficult to compare. PGD was performed for numerical abnormalities or for translocations following protocols previously published by our center. For numerical abnormalities the PGD procedure consisted of the FISH analysis of eight chromosome pairs (XY, 13, 15, 16, 18, 21, 22 plus either 1, 7, 14 or 17). Abnormalities detectable with this system are monosomies, trisomies, polyploidy, haploidy, chaotic embryos and a fraction of mosaic embryos. For PGD of translocations, usually one or two telomeric probes for the chromosomes involved in the translocation were analyzed with two or one centromeric probes for the same chromosomes, respectively.We defined that the results of a PGD cycle are predictable of the next if there was a difference of 0–20% points in the rate of normal embryos from one cycle to the next. For example, if in one cycle a patient had 30% normal embryos and in the next it had 30% to 50% normal embryos, we would consider for that the PGD results for the first cycle were “predictable” of the second cycle. The results are reported in Table 1 according to the PGD indication. In addition to Table 1, it was found that when all embryos were abnormal in the first cycle, the chance of finding at least one normal one in the second cycle was 95% for PGD of aneuploidy and 66% for PGD of translocations. The results of a PGD cycle of translocation are 90% predictable for the next. This is valuable data for patients with this condition to decide if they want to attempt a new PGD cycle. We have reported in the past that if 70% or more embryos are abnormal for the translocation, it is really difficult to conceive since in addition there is a baseline of 50% abnormalities for other chromosomes. Thus a patient with 70% or more abnormalities in the first cycle may decide to go to oocyte or sperm donation. Instead, cycles of PGD of aneuploidy show less predictability. Most probably translocation cycles are more predictable because they are genetic in origin, while numerical may have more environmental factors affecting the rate of chromosome abnormalities in these patients.

Over a decade of experience with preimplantation genetic diagnosis: A multicenter report

Objective To review a 12-year experience of the world's three largest preimplantation genetic diagnosis (PGD) centers. Design Multicenter analysis of the clinical outcome of PGD. Setting In vitro fertilization programs at the Reproductive Genetics Institute, Chicago, Illinois; Saint Barnabas Medical Center, West Orange, New Jersey; and SISMER, Bologna, Italy. Patient(s) Poor-prognosis IVF patients, patients carrying balanced chromosomal translocations, and couples at risk for producing children with Mendelian disorders. Intervention(s) In vitro fertilization, intracytoplasmic sperm injection, polar body removal, blastomere biopsy, and ET. Main outcome measure(s) DNA or chromosomal analysis of biopsied polar bodies or blastomeres, implantation and clinical pregnancy rates, and live-born pregnancy outcome. Result(s) A total of 754 babies have been born as a result of 4,748 PGD attempts, which shows the expanded application and the practical relevance of PGD for single-gene disorders, chromosomal aneuploidies and translocations, late-onset diseases with genetic predisposition, and nondisease testing in couples at need for human leukocyte antigens-matched offspring for treatment of affected siblings. Conclusion(s) Preimplantation genetic diagnosis is evolving to become a clinical option for couples at risk for producing offspring with Mendelian diseases, has a positive numerical impact in standard assisted reproduction practices through aneuploidy testing, and reduces by at least fourfold the spontaneous abortion rate in couples carrying translocations.

Efficacy and clinical outcome of preimplantation genetic diagnosis using FISH for couples of reciprocal and Robertsonian translocations: the Korean experience.

To evaluate the efficacy and clinical outcome of preimplantation genetic diagnosis (PGD) using fluorescence in situ hybridization (FISH) for couples with chromosomal translocations. PGD using FISH was performed in 59 cycles of 43 couples with reciprocal translocations, and 11 cycles of 6 couples with Robertsonian translocations. The diagnostic and clinical data were reviewed in a series of 70 treatment cycles of 49 couples from January 2001 to June 2002 at Samsung Cheil Hospital, Korea. A total of 1408 oocytes were retrieved, and 938 (81.7%) out of 1148 matured oocytes were fertilized by intracytoplasmic sperm injection (ICSI). Single blastomere biopsy and FISH analysis were successfully carried out in 99.3% (890/896) and 94.4% (840/890), respectively. Among 193 normal or balanced embryos, 169 embryos were transferred in 64 cycles (91.4% per started cycle). Twenty clinical pregnancies including two ectopic pregnancies and three spontaneous miscarriages (28.6% per started cycle, 31.3% per transfer cycle, 40.8% per couple) were established. Of the three spontaneous miscarriages, one was karyotyped as normal, one had an unbalanced arrangement and one was tetraploid. One case of preterm twin delivery occurred and 16 healthy babies were delivered in 12 single and 2 twin pregnancies. The clinical outcome was successful in 28.6% (14/49) of the treated couples with translocations after PGD. The spontaneous abortion rate was significantly reduced from 95.8% (69/72) to 16.7% (3/18) in these couples.

PGD results in embryos with odd versus even cell number on day 3 after retrieval

Background: In vitro fertililization success rates have increased with improvements in laboratory techniques. Preimplantation genetic diagnosis (PGD) has improved embryo selection by allowing the option to eliminate embryos with chromosomal abnormalities. Most embryos have an even number of cells as would be expected from normal cell division. We hypothesized that an odd number of cells might confer a higher probability of chromosomal abnormalities. Purpose: The purpose of this study was to determine 1) the frequency of embryos with an odd-number of cells and 2) to evaluate whether odd-number cell embryos are associated with more PGD abnormalities. Materials and Methods: Embryos from patients that underwent PGD at ART (Assisted Reproductive Technologies) from January through June 2003 were analyzed retrospectively. A total of 626 embryos were obtained from 92 women (mean age 38). Cell number and embryo-grade were documented at the time of the blastomere biopsy for PGD (day 3 after oocyte retrieval). PGD analysis was done for the following chromosomes: 13, 18, 21, X and Y. Results from PGD analysis were reported as normal female, normal male, abnormal female, abnormal male, and inconclusive. The embryo cell-number was compared with the results of PGD. Statistical analysis was performed using the χ2 test. Results: The results of PGD were inconclusive for chromosomal abnormalities, and therefore excluded in a total of 77 of 626 embryos (12%). Of the remaining embryos that were analyzed, 99 (19%) were odd-numbered (3,5,7, or 9 cells) and 450 (81%) were even-numbered (2,4,6,8,10,12, or 14 cells). No significant difference was noted in those embryos having chromosomal abnormalities between the odd-number cell and even-number cell groups (49% and 43% respectively). Similarly, no significant difference was noted between the groups for embryos without detectable chromosomal abnormalities or labeled as normal (50.5% vs. 57 %). Tabled 1PGD Chromosomal AnalysisEmbryo Cell NumberOddEvenNormal50 (50%)257 (57%)Abnormal49 (50%)193 (43%)Total99 (100%)450 (100%)Inconclusive embryos excluded (total 77).no significant difference. Open table in a new tab Inconclusive embryos excluded (total 77). no significant difference. Conclusion: In this study, a minority of embryos had an odd-number of cells (19%). The presence of an odd-number of cells was not helpful in predicting an abnormality in the five chromosomes tested using PGD. While not statistically significant, a higher percentage of embryos with even-cell number were noted to be normal by PGD.

PGD results in embryos with odd versus even cell number on day 3 after retrieval

Background: In vitro fertililization success rates have increased with improvements in laboratory techniques. Preimplantation genetic diagnosis (PGD) has improved embryo selection by allowing the option to eliminate embryos with chromosomal abnormalities. Most embryos have an even number of cells as would be expected from normal cell division. We hypothesized that an odd number of cells might confer a higher probability of chromosomal abnormalities. Purpose: The purpose of this study was to determine 1) the frequency of embryos with an odd-number of cells and 2) to evaluate whether odd-number cell embryos are associated with more PGD abnormalities. Materials and Methods: Embryos from patients that underwent PGD at ART (Assisted Reproductive Technologies) from January through June 2003 were analyzed retrospectively. A total of 626 embryos were obtained from 92 women (mean age 38). Cell number and embryo-grade were documented at the time of the blastomere biopsy for PGD (day 3 after oocyte retrieval). PGD analysis was done for the following chromosomes: 13, 18, 21, X and Y. Results from PGD analysis were reported as normal female, normal male, abnormal female, abnormal male, and inconclusive. The embryo cell-number was compared with the results of PGD. Statistical analysis was performed using the χ2 test. Results: The results of PGD were inconclusive for chromosomal abnormalities, and therefore excluded in a total of 77 of 626 embryos (12%). Of the remaining embryos that were analyzed, 99 (19%) were odd-numbered (3,5,7, or 9 cells) and 450 (81%) were even-numbered (2,4,6,8,10,12, or 14 cells). No significant difference was noted in those embryos having chromosomal abnormalities between the odd-number cell and even-number cell groups (49% and 43% respectively). Similarly, no significant difference was noted between the groups for embryos without detectable chromosomal abnormalities or labeled as normal (50.5% vs. 57 %). Tabled 1PGD Chromosomal AnalysisEmbryo Cell NumberOddEvenNormal50 (50%)257 (57%)Abnormal49 (50%)193 (43%)Total99 (100%)450 (100%)Inconclusive embryos excluded (total 77).no significant difference. Open table in a new tab Inconclusive embryos excluded (total 77). no significant difference. Conclusion: In this study, a minority of embryos had an odd-number of cells (19%). The presence of an odd-number of cells was not helpful in predicting an abnormality in the five chromosomes tested using PGD. While not statistically significant, a higher percentage of embryos with even-cell number were noted to be normal by PGD.

Predictability of preimplantation genetic diagnosis of aneuploidy and translocations on prospective attempts

The aim of this study was to determine if the outcomes of aneuploidy and translocation testing by preimplantation genetic diagnosis (PGD) at the 8-cell stage have a predictive value for new genetic diagnosis cycles. In total, 83 cycles (39 patients) undergoing PGD of translocations and 378 cycles (176 patients) of aneuploidy were included. Predictability, defined as having similar rate (±20%) of euploid embryos in the first and successive cycles, was found in 66% of patients undergoing aneuploidy testing. Predictability was found significantly more often in patients undergoing PGD of translocations (90%, P = 0.006). In addition, patients with 0, <30 or ≥30% euploid embryos in the first cycle were compared and groups 0 and <30% had significantly fewer euploid embryos in the second cycle (22–26%) than those of the group with ≥30% (37%) ( P < 0.05). Patients who did not become pregnant after the first attempt were stimulated more aggressively than those becoming pregnant, producing significantly more embryos in the second than in the first cycle ( P < 0.001). Therefore, correlation between euploidy rate and pregnancy rate could not be assessed objectively between cycles. In conclusion, the PGD results of a first cycle can predict the results of the second cycle, but this is likely to be of more value when the condition investigated is translocation rather than aneuploidy. The chance of pregnancy is usually related to the number of euploid embryos.

Blastocyst formation is not an indication of genetically normal embryos

Objective: Blastocyst formation is an important marker of embryo development. While embryo transfer at blastocyst stage improves implantation rate, development to the blastocyst stage may not indicate that the embryo is genetically normal. Preimplantation genetic diagnosis (PGD) can be used to screen for numerical chromosomal abnormalities and for single gene defects embryos at the 6–8 cell stage. This study examines the relationship between blastocyst formation and PGD results.

Is reduced blastocyst development associated with chromosomal abnormality after PGD?

Objective: Preimplantation genetic diagnosis (PGD) procedures are commonly used to check for specific genetic diseases and chromosomal abnormalities. This study was undertaken to determine if preembryo morphology on day 3 and rate of blastocyst development on day 5 are associated with PGD results.

The minisequencing method: an alternative strategy for preimplantation genetic diagnosis of single gene disorders.

We have applied a new method of genetic analysis, called 'minisequencing', to preimplantation genetic diagnosis (PGD) of monogenic disorders from single cells. This method involves computer-assisted mutation analysis, which allows exact base identity determination and computer-assisted visualization of the specific mutation(s), and thus facilitates data interpretation and management. Sequencing of the entire PCR product is unnecessary, yet the same qualitative characteristics of sequence analysis are maintained. The main benefit of the minisequencing strategy is the use of a mutation analysis protocol based on a common procedure, irrespective of the mutations involved. To evaluate the reliability of this method for subsequent application to PGD, we analysed PCR products from 887 blastomeres including 55 PGD cases of different genetic diseases, such as cystic fibrosis, beta-thalassaemia, sickle cell anaemia, haemophilia A, retinoblastoma, and spinal muscular atrophy. Minisequencing was found to be a useful technique in PGD analysis, due to its elevated sensitivity, automation, and easy data interpretation. The method was also efficient, providing interpretable results in 96.5% (856/887) of the blastomeres tested. Fifteen clinical pregnancies resulted from these PGD cases; conventional prenatal diagnosis confirmed all the PGD results, and 10 healthy babies have already been born. Its applicability to PGD could be helpful, particularly in cases in which the mutation(s) involved are difficult to assess by restriction analysis or other commonly used methods.

Predictive value of sperm fluorescence in situ hybridization analysis on the outcome of preimplantation genetic diagnosis for translocations

Objective To determine whether the proportion of abnormal sperm is predictive of the proportion of abnormal embryos from couples in which the males are translocation carriers. Design Controlled clinical study. Setting Private in vitro fertilization (IVF) center. Patient(s) Eleven cases of reciprocal translocation male carriers. Intervention(s) Blood sample and sperm sample collection from each male partner. Embryo biopsy of the embryos produced in each cycle. Main outcome measure(s) Fluorescence in situ hybridization on lymphocyte slides to characterize each translocation case, then fluorescent in situ hybridization (FISH) with specific probes for each of the sperm samples. Preimplantation genetic diagnosis of the translocations in the 11 cases. Result(s) A correlation was found between the percentage of abnormal gametes and the percentage of abnormal embryos, and a predictive equation is proposed for this relationship: A = −55 + (1.9 × B), where A is the percentage of abnormal embryos and B the percentage of abnormal sperm. Conclusion(s) The predictive value of the sperm analysis was established. Patients with 65% or less chromosomally abnormal sperm have a good chance at conceiving; patients with higher rates would need to produce 10 or more good quality embryos to have reasonable chances of conceiving.

Results of preimplantation genetic diagnosis in patients with Klinefelter's syndrome

With the application of preimplantation genetic diagnosis (PGD), a possible genetic contribution of spermatozoa obtained from 47,XXY non-mosaic Klinefelter patients on preimplantation embryos was analysed in eight couples. Interpretable fluorescence in-situ hybridization results were obtained for 28 out of 33 embryos biopsied (84.8%) and 23 blastomeres were analysed for chromosomes 13, 18, 21, X and Y. Nine out of 23 embryos were diagnosed as abnormal (39.1%). Five out of nine contained sex chromosome abnormalities (55.5%). Two were diagnosed as 47,XXY and three were found to have monosomy X. Besides sex chromosomal abnormalities, other abnormalities detected were haploidy, triploidy, monosomy 13, monosomy 18 and trisomy 13. Five blastomeres were analysed for sex chromosomes only and all of them were found to be normal. Overall, the rate of sex chromosome abnormality in biopsied embryos was found to be 17.8% (5/28). Moreover, among 33 embryos biopsied, five of the eight zygotes, which were classified as a poor prognosis group according to pronuclear morphology scoring, showed an impaired growth profile after biopsy and were found to be chromosomally abnormal. Elimination of abnormal embyos and transfer of normal ones resulted in four pregnancies in eight cycles (50%). Two pregnancies, one singleton and one twins resulted in healthy births. Two pregnancies, one singleton and one twins are continuing beyond the second trimester. These results show that there is in fact elevated chromosomal abnormality for both sex chromosomes and autosomes in embryos developed from Klinefelter males. Furthermore together with PGD, embryo scoring according to pronuclear morphology can give additional benefit for selecting chromosomally abnormal embryos. Therefore, PGD should be recommended in cases with Klinefelter's syndrome and this information should be discussed with the couple when genetic counselling is given.

ESHRE Preimplantation Genetic Diagnosis Consortium: data collection III (May 2001).

The ESHRE PGD Consortium was formed in 1997 to undertake a long-term study of the efficacy and clinical outcome of preimplantation genetic diagnosis (PGD). Here, the third report of the ESHRE PGD Consortium is presented, collating data received from 25 centres on referrals, cycles, pregnancies and babies born after PGD. The second report, published in December 2000, reported on 886 referrals, a total of 1318 started cycles (of which 465 for aneuploidy screening, 386 for fluorescence in-situ hybridization (FISH) and 385 for PCR going beyond oocyte retrieval), 163 pregnancies and 162 children born. This year, 675 referrals from 12 centres were added giving a total of 1561 referrals, 370 regular PGD cycles, 334 PGD-aneuploidy screening (PGD-AS) cycles and 78 cycles for social sexing from 24 centres and 215 pregnancies and 117 babies from 12 centres. Because more in-depth information was asked for the cycles, this year's data will be shown separately as well as cumulatively. One striking feature of this year's data collection is the appearance of the first data for gender screening on preimplantation embryos for social reasons. The ethical concerns regarding social sexing will be discussed, as well as the forthcoming changes in timing of data collection. When the data collection was discussed at the last meeting of the members of the ESHRE PGD Consortium in Lausanne, Switzerland (June, 2001), it appeared that the current system of data collection, although yielding results very quickly, showed fundamental flaws. The ESHRE PGD Consortium Steering Committee intends to remedy to these problems, on the one hand by introducing a new type and timing of data collection, and on the other hand by re-analysing and correcting the data which have already been sent in during the past 4 years.

Preimplantation genetic diagnosis for cancer predisposition

Preimplantation genetic diagnosis (PGD) has recently been offered for couples with an inherited predisposition for late onset disorders. This paper presents the results of PGD for a group of couples at risk for producing children with cancer predisposition. Using a standard IVF procedure, oocytes or embryos were tested for different mutations predisposing to cancer, preselecting and transferring only mutation-free embryos back to the patients. The procedure was performed for patients with predisposition to familial adenomatous polyposis coli (FAP), Von Hippel-Lindau syndrome (VHL), retinoblastoma, Li-Fraumeni syndrome, determined by p53 tumour suppressor gene mutations, neurofibromatosis types I and II and familial posterior fossa brain tumour (hSNF5). Overall, 20 PGD cycles were performed for 10 couples, resulting in preselection and transfer of 40 mutation-free embryos, which resulted in five unaffected clinical pregnancies and four healthy children born by the present time. Despite the controversy of PGD use for late onset disorders, the data demonstrate the usefulness of this approach as the only acceptable option for at-risk couples to avoid the birth of children with an inherited predisposition to cancer, and to have a healthy child.

ESHRE preimplantation genetic diagnosis (PGD) consortium: data collection II (May 2000).

In 1997, the ESHRE PGD Consortium was formed as part of the ESHRE Special Interest Group on Reproductive Genetics, in order to undertake a long-term study of the efficacy and clinical outcome of preimplantation genetic diagnosis (PGD). In December 1999, the first PGD Consortium report was published discussing referrals of 323 couples, 392 PGD cycles and 82 pregnancies and 79 children born. In the second round of data collection, contributing centres were asked to send in data from their PGD activities before January 1997, as well as from 1st October 1998 until 1st May 2000, in order to have as complete as possible an overview of PGD practices in these centres. A further 563 referrals were sent in as well as 926 PGD cycles, and data on 89 pregnancies (including seven pregnancies ongoing from the previous group) and 83 children were collected. This has led to a considerable amount of cumulative data being acquired: over a period of 7 years (the oldest PGD cycle reported dates from 1994), referral data on 886 couples, cycle data on 1318 PGD cycles and data on 163 pregnancies and 162 babies were collected. In all, these data are encouraging: they show first, that the practice of PGD is becoming more and more established, and an increasing number of different applications is emerging; and second, that collecting these data is worthwhile, as they will be a valuable source of information for all those involved, e.g. in counselling patients and interacting with governmental bodies.

Neonatal outcome of preimplantation genetic diagnosis by polar body removal: the first 109 infants.

Our center developed the technique of preimplantation genetic diagnosis (PGD) by sequential polar body removal (PBR) for the diagnosis of Mendelian disorders and aneuploidies. This study examines the obstetric and neonatal outcome of the first 109 live births after PGD by PBR. To determine if there were any observable effects of PGD by PBR on perinatal morbidity and mortality, birth defects, and growth parameters. Data on perinatal outcome were gathered for the first 109 infants by parental reporting and confirmed by telephone interview and chart review when indicated. In infants >6 months old, a follow-up telephone interview was performed establishing the developmental milestones attained by the child. A research center conducting an institutional review board-approved research protocol in PGD. All patients who had PGD by PBR who had clinical pregnancies. Gestational age, mode of delivery, perinatal mortality, birth weight, birth length, the presence of birth defects, and developmental milestones. There was no significant decrease in birth length or weight, or the frequency of small for gestational age infants. No specific pattern of birth defects was observed. Thus far, there are no observable detrimental effects of PGD by PBR on children born after the procedure.

Clinical Use of a Pronuclear Stage Score Following Intracytoplasmic Sperm Injection (ICSI) Compared with Chromosomal Status

Objectives: Selection of viable embryos for transfer currently plays a significant role in the effort to increase in vitro fertilization (IVF) programs and embryos transfer pregnancy rates. Recently, studies have published new classification based for pronuclear (PN) stage. This score included the morphological appearance of pronuclei and nucleoli. Our purpose in this study was evaluated the role of PN morphology and compared embryo morphology preimplantation genetic diagnosis (PGD) from day + 3 embryos. Design: Retrospective study. Materials and Methods: A total of 21 cycles (74 embryos) from intracytoplasmic sperm injection (ICSI) procedure was evaluated in this study. For each embryo, the following data were recorded during sequential observations. PN morphology 16–20 hours after injection, embryo morphology about 68 hours after ICSI (day + 3) and PGD on day +3. Two patterns of pronuclear morphology were distinguished, according to position of the PN: aligned or close pronuclei and separated or unequal pronuclei. Nucleoli morphology was also performed based on: nucleoli aligned; nucleoli aligning and scattered nucleoli. Cumulative embryo score (CES) was performed in each embryo referring to PN and nucleoli morphology simultaneously. An embryo was classified as having good quality (GE) if more than 4 cells were present with less than 20% fragmentation at 68 hours after ICSI (day + 3). Probes with chromosomes X, Y, 18, 13 and 21 were numerically analyzed through the biopsy of 1 or two blastomeres. Results: The diagnosis was obtained in 70 embryos. Close or aligned PN showed to have a higher number of embryos with normal results to PGD when compared with embryos that had separated or unequal PN (56% vs. 45.5%, p<0.05). When nucleoli were aligned, best PGD results were noted when compared with aligning or scattered nucleoli (83.3%, 33.3% and 52.2% of normal embryos, respectively, p<0.05). Nucleoli aligning seemed to exhibit lower rates (33.3%) of numerical normality for these chromosomes when we compared with others embryos. Aligned PN and nucleoli provide a better score to the embryo with better PGD results for numerical normality (90.9%, p<0.05). CES and embryo morphology was also compared and 88% of GE demonstrated normal results to PGD. However, relationship between embryo morphology and PGD results was established only in embryos with aligned PN and nucleoli (88% GE vs. 12% no GE; p<0.005). Conclusions: Nucleoli and PN morphology can be a predictive factor to select embryos with normal chromosomal status and represents an efficient criteria for transfer citogenetically normal embryos in ICSI cycles.

Preimplantation Genetic Diagnosis Failures: What Happened?

Objectives: For preimplantation genetic diagnosis (PGD) where only one or two cells (no more than 25% of the blastomeres) are available, fluorescent in situ hybridization (FISH) can be used, for sexing embryos at risk of passing on X-linked disorders for which there is, as yet, no specific molecular diagnosis. In addition, FISH is useful for couples who are subfertile owing to chromosomal disorders, recurrent miscarriage or maternal age. Since first report the use of FISH for PGD several improvements to the technique have been made. However, cell embryos can be lost during the PGD procedure for several factors. The purpose of this study was to assess the PGD results and probable failure causes.