Comparison of Nicolaides’ risk evaluation for down’s syndrome with a novel software: an analysis of 1,463 cases

Objective The individual risk assessment of fetal Down’s syndrome based on measurement of nuchal translucency (NT) according to Nicolaides, optionally complemented by the determination of PAPP-A and free beta HCG has progressively supplanted other search strategies for fetal aneuploidies. It could be shown that this diagnostic strategy equally detects other numeric aneuploidies at a comparable rate. A positive test result is also predictive for the presence of a fetal malformation. In this field, several computer programs are available for clinical use. The objective of our study was to re-evaluate the first consecutive 1463 NT-risk calculations determined by Nicolaides’ method and to compare the risk calculation to the JOY software (NT-risk calculation module, JOY Patient Database) introduced in 2002. Material and methods At the Department of Obstetrics and Gynecology, Hannover Medical School, 1463 consecutive complete data sets comprising first trimester screening performed between May 2, 2000 and June 26, 2003 and corresponding fetal outcome were analysed using risk assessment based on the Nicolaides method (PIA Fetal Database NT-Module) and compared with the risk evaluation as determined by the JOY software (JOY Patient Database NT module). A risk exceeding 1:300 was considered to indicate the need for further invasive testing. In a first step, only cytogenetically detectable chromosomal aberrations were analysed. Then, a second evaluation including fetal malformations was performed. Results Among the 1463 cases, 1445 (98.77%) fetuses revealed to be cytogenetically healthy. Both softwares showed identical detection rates at the genetic and somatic level:13 cases of Down-Syndrome (0.89%), 2 cases of trisomy 18 (0.14%), one case of triploidy, one Turner-Syndrome, one Klinefelter-Syndrome (0.07% each) were detected. A positive test result was found in 15 cases ending in a spontaneous abortion, intrauterine death or peripartum death (1.03%) and in 22 cases of fetal malformation (1.50%). At the level of genetic detection the test positive rate dropped from 92 (PIA) to 71 (JOY) (-22.8%). At the level of combined adverse outcome the test positive rate was reduced from 100 (PIA) to 76 (JOY) (-22.0%), thus yielding in a marked improvement of the characteristic test performance parameters. Conclusion The novel, recently developed JOY software package allowed reliable evaluation of the risk for aneuploidy with increased specificity whereas sensitivity was unchanged. Our data suggest an improvement of the screening for aneuploidy when using this novel software: With an identical detection rate, the number of unnecessary invasive measures may be reduced.     

A rare occurrence of trisomy 18 and trisomy 21 in a dizygotic twin pregnancy

Objective Case report of a rare combination of a trisomy 18 and 21 in a dizygotic twin pregnancy in a woman with a history of recurrent miscarriage, a neonatal death, no living offspring and Graves disease. Methods Case report and literature search. Results Only one other report in the literature of a combined trisomy 18 and 21 twin pregnancy was found. Conclusion The combination of a trisomy 18 and 21 in a dizygotic twin pregnancy is very rare. Despite the high frequency and clinical importance of aneuploidy, very little is known about the factors that may modulate meiotic non-disjunction.     

染色体微阵列分析技术在流产或死胎原因分析中的应用

目的探讨染色体微阵列分析技术(chromosomal microarray analysis,CMA)在流产或死胎原因分析中的应用价值及流产或死胎与染色体异常的关系。方法采用CMA技术对流产绒毛或死胎组织进行全基因组拷贝数变异(copy number variations,CNVs)检测。结果824例流产或死胎样本检测成功率100%,染色体异常381例(46.2%),其中数目异常312例(81.9%),结构异常66例(17.3%),单亲二倍体(uniparental disomy,UPD)3例(0.8%)。数目异常中占比最大的为非整倍体,共287例(92.0%),以16-三体和Turner综合征最为多见,分别为41例(13.1%)和63例(20.2%)。66例染色体结构异常中,26例(39.4%)发生拷贝数重复,20例(30.3%)发生拷贝数缺失,20例(30.3%)发生拷贝数重复伴缺失,其中33例检出临床致病的可能性大。结论CMA是诊断流产或死胎病因的一种可靠、稳定、高分辨的技术。胚胎染色体数目异常是引起临床流产的主要遗传因素,其中以非整倍体变异最为常见。     

Healthy live births after mosaic blastocyst transfers with the use of next-generation sequencing

ObjectiveTo determine whether transfer of high-mosaicism (≥50%) embryos can result in healthy newborns.Case reportTwo embryos resulting from controlled ovarian stimulation (COS) in Patient one, 41 years of age (y/o), underwent preimplantation genetic testing for aneuploidy (PGT-A), which demonstrated that one was mosaic (68%) and the other aneuploid; the mosaic embryo was transferred. Amniocentesis at 18 weeks of gestational age (GA) revealed a normal 46, XY karyotype. A phenotypically normal boy was delivered at 39 and 5/7 weeks of GA. For Patient two, 39 (y/o), nine embryos obtained after COS underwent PGT-A, indicating one euploid, four mosaic, and four aneuploid embryos. One euploid and one mosaic (50%) embryo were transferred, resulting in a twin pregnancy. Amniocentesis at 18 weeks of GA showed both fetuses had normal 46, XY karyotypes. Two phenotypically normal boys were delivered at 37 2/7 weeks of GA.ConclusionTransfer of high-mosaicism embryos selected using current techniques can result in healthy euploid newborns. Amniocentesis suggested that mosaic embryos can be self-corrected before 18 weeks of GA.     

氯化汞对雄性果蝇生殖细胞非整倍体的影响

应用黑腹果蝇Ｘ或Ｙ染色体丢失试验研究了氯化汞对雄性果蝇生殖细胞染色体的损伤。结果表明，染毒浓度为０．１％及０．２％的氯化汞组染色体丢失率均为０．０４％，与阴性对照组的染色体丢失率用Ｋａｓｔｅｎｂａｕｍ－Ｂｏｗｍａｎ表检验，差异无显著性，即氯化汞对雄性果蝇生殖细胞的染色体无损伤作用，此结果可能由于所用雄蝇品系不够敏感所致。     

Inducing the loss of conditionally dispensable chromosomes in Nectria haematococca during vegetative growth

A procedure for inducing and detecting the loss of conditionally dispensable (CD) chromosomes in filamentous fungi during vegetative growth was developed using Nectria haematococca mating population VI as a model. CD chromosomes in two different isolates of N. haematococca were tagged via integrative transformation with a gene conferring resistance to hygromycin B. In each case the transformation vector included chromosome-specific DNA in order to direct its homologous recombination with the desired chromosome. Chromosome loss was induced by exposing tagged isolates to inhibitory concentrations of benomyl either for protracted periods of time on solid medium or for short periods of time in liquid medium. After exposure to benomyl, isolates that lost the tagged chromosome were identified by their loss of resistance to hygromycin B. Electrophoretic karyotyping was used to verify that isolates which failed to grow on hygromycin B lacked an intact CD chromosome. Ten other chemicals known to interfere with mitotic events or cell development in other organisms did not induce CD chromosome loss in N. haematococca. Received: 2 September / 8 December 1997     

Interindividual Variations in the Disomy Frequencies of Human Spermatozoa and their Correlation with Nuclear Maturity as Evaluated by Aniline Blue Staining

Objective: To evaluate the importance of interindividual variations in the disomy frequencies of human sperm and their possible correlation with the principal semen parameters.

Design: Prospective randomized analysis of sperm nuclei by fluorescence in situ hybridization and analysis of semen parameters.

Setting: University-based laboratory for reproductive biology.

Patient(s): Fifty-seven human ejaculates selected at random from a population of men undergoing semen analysis.

Intervention(s): Semen specimens were analyzed, and sperm samples were prepared for fluorescence in situ hybridization.

Main Outcome Measure(s): Semen parameters, including necrozoospermia, global motility, sperm concentration, multiple abnormalities index, and teratozoospermia were evaluated, aniline blue staining was completed, and disomy frequencies for chromosomes 8, 15, 18, X, and Y were determined using fluorescence in situ hybridization.

Result(s): Noticeable differences in disomy frequencies between individuals were observed, and these frequencies were correlated with the degree of nuclear maturity.

Conclusion(s): We hypothesize that the positive correlation can be explained by an abnormality of chromosomal segregation at the time of meiosis that would cause disturbances during the transition of nucleoprotein or by one or several premeiotic abnormalities of chromatin that would perturb both the meiotic process and the construction of definitive proteins.     

New protocol based on massive parallel sequencing for aneuploidy screening of preimplantation human embryos

Novel next-generation sequencing procedures have rapidly emerged into the preimplantation genetic screening framework. This work presents the design and validation of a new low-coverage whole-genome sequencing assay for aneuploidy detection in single blastomeres and trophectodermal samples from preimplantation embryos. The validation ensures analytical sensitivity, specificity, robustness, precision, limit of detection, resolution, and reproducibility. Specific parameters to measure the performance are defined, and the results are compared with a standardized array-based method to stablish the concordance. From the single cell genomics point of view, the main novelties are the length of reads of the libraries (150 nucleotides) together with a paired-end strategy and the design of an original algorithm and copy number viewer. A total of 129 samples were included in six experimental runs using a MiSeq Illumina platform. Samples included: single amniocytes, single blastomeres (cleavage-stage embryos), trophectoderm samples (blastocyst), and diluted DNA. Sensitivity and specificity were calculated per chromosome yielding 96% and 99%, respectively. The percentage of concordant samples was 98.2% and all of the aneuploid samples were confirmed. In conclusion, the validation yields highly reliable and reproducible results, representing an accurate and cost-effective strategy for the routine detection of aneuploidy in human embryos.