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Prenatal Diagnosis (PND) forms an important part of primary preventive management for families having a child affected with primary immunodeficiency. Although individually sparse, collectively this group of genetic disorders represents a significant burden of disease. This paper discusses the prenatal services available for affected families at various centers across the country and the challenges and ethical considerations associated with genetic counseling. Mutation detection in the index case and analysis of chorionic villous sampling or amniocentesis remain the preferred procedures for PND and phenotypic analysis of cordocentesis sample is reserved for families with well-characterized index case seeking PND in the latter part of the second trimester of pregnancy. A total of 112 families were provided PND services in the last decade and the presence of an affected fetus was confirmed in 32 families. Post-test genetic counseling enabled the affected families to make an informed decision about the current pregnancy.

Objective: The study aim was to determine the hemoglobin genotypic and allelic distributions in fetal population, and to quantitatively evaluate the effect of heterozygous maternal DNA contamination of homozygous fetal DNA in chorionic villi, on fetal hemoglobin genotypes.Materials and Methods: A descriptive, cross sectional study of amplification refractory mutation system‑polymerase chain reaction (ARMS‑PCR) analysis of fetal hemoglobin genotype using DNA from chorionic villi and maternal venous blood. The primary fetal hemoglobin genotypes were obtained in the first phase and the secondary hemoglobin genotypes after contaminating homozygous primary genotypes (HbAHbA and HbSHbS) with varying proportions of heterozygous maternal DNA (HbAHbS). Data analysis was done with Micosoft Excel 2010 statistical package and Chi‑square (goodness‑of‑fit).Results: There was no statistically significant deviation in the hemoglobin genotypic and allelic counts between the observed and the expected counts in the fetal population based on Mendelian expectation. Contaminating homozygous fetal DNA with >11.1% (0.5 μl) of heterozygous maternal DNA produced significant change in fetal hemoglobin genotype results. Homozygous hemoglobin genotypes HbAHbA were affected more than HbSHbS.Conclusion: Study established Mendelian distribution in the fetal population and the levels of heterozygous maternal contamination of homozygous fetal DNA that resulted in significant risk of misdiagnosis.Key words: ARMS‑PCR; fetal; maternal contamination; sickle cell anemia.

Objective Celocentesis, which involves aspiration of celomic fluid at 7–9 weeks' gestation, can potentially provide early prenatal diagnosis of single-gene disorders. The main barrier to wide acceptability of this technique is contamination of the sample by maternal cells. This problem can be overcome through selection of embryo-fetal erythroid precursors, which are found in celomatic fluid. Method Embryo-fetal erythroid precursors were selected by an anti-CD71 MicroBeads method or by direct micromanipulator pickup of the cells selected on the basis of their morphology. Results In our series of 302 singleton pregnancies at high risk for hemoglobinopathies, Celocentesis provided a sample of celomic fluid in all cases. In 100 (33.1%) samples, maternal contamination was absent or very low ( 60%, and selection of embryo-fetal cells was achieved by micromanipulation. In all 302 cases, there was concordance between DNA obtained from celomic fluid samples and fetal or newborn DNA. Conclusions Celocentesis can be a reliable procedure for earlier prenatal diagnosis of fetal monogenic diseases. © 2016 John Wiley & Sons, Ltd.

Background: We performed counselling for prenatal diagnosis (PD) of haemoglobinopathies in 372 couples. Thirty-four out of 372 (9.1%) did not undergo PD: six due to spontaneous abortion; nine because it was too difficult to make a decision if PD was positive; 18 because counselling excluded the carrier status of one or both parents; and one because parental mutations were mild. Methods: Eleven out of 338 (3.3%) couples underwent PD because they had a thalassaemic child; 106 (31.4%) were found to be at high risk during pre-conceptional screening; 221 (65.4%) because of familiarity. Of 523 PDs in 486 (92.9%), including six dichorionic twin pregnancies, PD was performed on DNA from chorionic villi (CV), and in 37 from amniocytes (7.1%). In 1/523 cases, PD was not completed because DNA from CV was not sufficient; in two cases single tandem repeat analysis revealed maternal contamination of foetal DNA; in 7/522 (1.3%) cases PD revealed non-paternity. In 435/522 (83.3%) cases, PD was performed using reverse dot-blot and ARMS; 34/522 (6.5%) required sequencing. In 53/522 (10.2%) cases it was necessary to test globin loci for large rearrangements. Results: One hundred and twenty out of 522 (23.0%) PDs revealed an affected foetus. In all but two cases the couple interrupted pregnancy. In the six twin pregnancies PD revealed a normal and a carrier foetus (two cases), carrier status in both foetuses (two cases) and a carrier and an affected foetus (two cases). In these latter cases the couple planned selective interruption. Conclusions: Our PD procedure is successful and reliable, and is useful in high-risk areas characterised by molecular heterogeneity.

The aim of this work was to compare the performances of the Projector Operator (PO) and the independent component analysis (ICA) based method for fetal signal extraction from fMCG. Both methods were applied on data recorded with the SARA system installed at the UAMS, Little Rock (Arkansas-USA), and with the ATB system installed at ITAB-Chieti (Italy). Twenty datasets recorded with both systems were analysed. The detection rates, signal-to-noise ratios, fetal signal amplitude reconstructions and residual maternal contamination of the reconstructed fetal signals were calculated, and the PO and ICA-based approach provided fetal signals with similar features, although ICA seemed to be more effective when noisy data were processed. © 2007 Elsevier B.V. All rights reserved.

Made available in DSpace on 2018-12-11T17:04:36Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-07-01. Added 1 bitstream(s) on 2019-10-09T18:30:32Z : No. of bitstreams: 1 S0102-86502016000700490.pdf: 254673 bytes, checksum: 267b4dc1f9bbf72e8143aa945cb66847 (MD5) PURPOSE: To evaluate the contamination index of metals and pesticides in pregnant women, and to relate this to perinatal outcomes. METHODS: Descriptive, retrospective, exploratory study, developed from existing secondary data analyses at Level III maternity center. A total of 40 mothers with their newborns (NB), living in a rural area in Botucatu-Brazil and surrounding region. Blood samples from mothers and newborn were collected to determine the total contamination index for metals and pesticides. The concentrations of each metal and each pesticide were determined in blood samples of mothers and their newborns by Rudge’s results. After obtaining these concentrations, the total contamination index in mother and NB was calculated, along with its correlation with clinical parameters of NB. RESULTS: There was no correlation (p> 0.05) between maternal contamination index with NB clinical parameters, and NB contamination index versus NB clinical parameters. CONCLUSION: The maternal contamination index of metals and pesticides was not related to perinatal outcomes, but it could be used as baseline parameter in future toxicological studies, regarding to long-term toxic characteristics as persistent organic pollutants, its long half-lives, bioacumulative, and expected to impose serious health effects on humans. Laboratory of Experimental Research on Gynecology and Obstetrics and Gynecology Obstetrics and Mastology Graduate Course Botucatu Medical School Universidade Estadual Paulista (UNESP) Laboratory of General Physiology and Reproductive Toxicology Institute of Biological and Health Sciences University Center of Araguaia Universidade Federal do Mato Grosso (UFMT) Laboratory of Experimental Research on Gynecology and Obstetrics Botucatu Medical School UNESP Laboratory of Experimental Research on Gynecology and Gynecology Obstetrics and Mastology Graduate Course Botucatu Medical School UNESP Gynecology Obstetrics and Mastology Graduate Course Botucatu Medical School UNESP Department of Morphology Institute of Biosciences UNESP Department of Community Medicine Faculty of Health Sciences The Arctic University of Norway Laboratory of Experimental Research on Gynecology and Obstetrics and Gynecology Obstetrics and Mastology Graduate Course Botucatu Medical School Universidade Estadual Paulista (UNESP) Laboratory of Experimental Research on Gynecology and Obstetrics Botucatu Medical School UNESP Laboratory of Experimental Research on Gynecology and Gynecology Obstetrics and Mastology Graduate Course Botucatu Medical School UNESP Gynecology Obstetrics and Mastology Graduate Course Botucatu Medical School UNESP Department of Morphology Institute of Biosciences UNESP

Purpose: About 50% of spontaneous abortions are caused by fetal chromosome abnormalities. Identification of these abnormalities helps to estimate recurrence risks in future pregnancies. However, due to culture failures or maternal contamination often no fetal karyotype can be obtained. Array comparative genomic hybridization can overcome some of these limitations. Methods: In this study, we analyzed 103 miscarriages by both T-banding and 1-Mb array comparative genomic hybridization. Results: We found an overall abnormality rate of 35% (34 of 96). In a comparison of 70 samples that were successfully analyzed by both techniques, 54 (77%) had identical karyotypes (42 normal, 12 abnormal) and 16 (23%) cases showed discrepancies. Most of these differences were due to maternal contamination during cell culture, which resulted erroneously in a normal female karyotype. Conclusion: These results demonstrate the improved diagnostic yield of array comparative genomic hybridization as compared with conventional karyotyping. Therefore, we implemented this technique in the diagnostic workup of miscarriages.

Isolation of fetal cells from maternal circulation is the subject of intense research to eliminate the need for currently used invasive prenatal diagnosis procedures. Fetal cells can be isolated using magnetic-activated cell sorting or fluorescence-activated cell sorting, however no technique to specifically isolate and use fetal cells for genetic diagnosis has reached routine clinical practice. This paper demonstrates the use of a micromachined device to separate fetal cells from maternal circulation based on differences in size and deformation characteristics. Nucleated fetal red blood cells range in diameter from 9 to 12 microm can deform and pass through a channel as small as 2.5 microm wide and 5 microm deep. Although the white blood cells range in diameter from 10 to 20 microm, they cannot deform and are retained by the 2.5 microm wide and 5 microm deep channels under our experimental conditions. Fetal cells were isolated from cord blood and DNA analysis confirmed their fetal origin with ruled out maternal contamination.

Objective To evaluate the service for prenatal diagnosis of β-thalassemia in Pakistan. Methods All prenatal diagnoses (PNDs) for β-thalassemia since the introduction of the service in 1994 were studied. PND was done by the Amplification Refractory Mutation System (ARMS), or linkage analysis, when required. The reported errors in PND were investigated for clerical mistakes, technical problems with PCR, maternal contamination and nonpaternity. Results In the 12 years 2174 PNDs were done for β-thalassemia at the country's main referral center. The use of PND has increased from 26 in 1994 to 381 in 2006. Over 97% of the couples who requested PND already had an affected child. In over 97% of the cases PND was done by direct mutation analysis. The reported rate of misdiagnosis was 0.37%. The causes of misdiagnoses included one clerical mistake, three false positive PCR results, and two maternal contaminations in the chorionic villus sampling (CVS). Conclusion PND for β-thalassemia is technically feasible by direct mutation analysis in most cases in Pakistan. The procedure is quick and cost effective. Strict quality assurance can achieve an acceptably low error rate. Copyright © 2007 John Wiley & Sons, Ltd.

Beta-thalassaemia constitutes a major health burden on the limited health resources of India and prenatal diagnosis is seen as an important preventive measure to reduce the burden of the disease. Prenatal diagnosis has been offered to 99 women in 112 pregnancies by fetal DNA analysis, using a PCR-based Amplification Refractory Mutation System (ARMS) for the common and uncommon Indian mutations. Restriction fragment length polymorphism (RFLP) for the beta-globin gene was used when the mutation remained unidentified in one of the parents or to complement the ARMS result. In 53 cases the fetus tested had beta-thalassaemia trait (betaTT) (47.3%), 22 were normal (19.6%) and 31 had thalassaemia major (27.6%). In five cases (4.5%), a definitive report could not be given due to maternal contamination. In one case (0.9%), there was a misdiagnosis. Pitfalls encountered in the diagnosis were maternal contamination and occasional non-amplification of the primers. Having established a regional centre for the prenatal diagnosis for thalassaemia, the screening programmes will be enlarged both to identify carriers and prevent the birth of further homozygous children, even during the first pregnancy.

ApoB3' hyper variable region is one of the highly polymorphic genetic marker and reveals a high degree of allelic variation in different populations therefore; it can be a useful marker for different clinical tests in which individual differences at DNA level form the basis of detection. In the present study we compared Apo B3 HVR with other 28 STR markers at allele frequency level, heterozygosity, polymorphism information content (PIC) and power of exclusion. Our results indicated a high degree of heterozygosity, PIC and power of exclusion for Apo B3 HVR. These criteria lead us to investigate this marker for different purposes like detection of maternal contamination in chorionic villus samples and chimerism studies after the engraftment of bone marrow in bone marrow transplantation patients. The utility of this marker has been discussed in comparison of other markers.

Summary Newborn screening for sickle cell disease commenced in 1992 in Sao Paulo State and by the end of 2000, the programme covered 78 institutions in 36 municipalities with the screening of 281 884 babies. Initially based on liquid cord blood samples, these are being replaced by dried filter paper capillary samples to ease handling and avoid diagnostic confusion from maternal contamination. The prevalence of sickle cell trait (2.0%) and HbC trait (0.6%) increased significantly between 1996 and 2000, apparently because of improved detection rather than the later introduction of institutions serving populations with higher trait frequencies. There were 29 babies with homozygous sickle cell SS disease and 26 with sickle cell-haemoglobin C (SC) disease, the latter significantly exceeding expectation and possibly attributable to a nonrandom selection of partners. Sickle cell-β thalassaemia syndromes were proportionately more common than in Jamaica, and it is possible that this results from interaction with other Brazilian populations carrying higher β thalassaemia gene frequencies. The frequency of abnormal haemoglobins in this population is lower than in Jamaica, but clinically significant sickle cell disease occurred once in every 5527 births, comparable with the frequencies of other significant inborn errors of metabolism.

karyotype is a well established technique in the study of spontaneous miscarriages but is associated with selective overgrowth of maternal cells and other culture artefact (spp) such as tetraploidy, which could mask the true karyotype of the conceptus.328 cases of pregnancy losses were studied by karyotype and Multiplex Ligation Dependent Probe Amplification technique. Touch FISH performed in non-cultured cells was used to evaluate the ploidy complement and sex discrepancies using centromeric probes for chromosomes X, Y and 18.touch FISH confirmed 13 cases of maternal contamination, identified a triploidy and a monosomy X. True tetraploidy was confirmed in 7/14 cases studied.touch FISH protocol is extremely accurate in the distinction of genuine mosaicism from tissue culture artifacts namely in cases with suspicion of tetraploidy and can be used to confirm maternal cell contamination in cases with sex discrepancy between karyotype and MLPA.

Several aspects of the biology and life cycle of Toxoplasma gondii are important limiting factors for the evaluation of drugs for the treatment of congenital toxoplasmosis: (i) T. gondii is an obligate intracellular parasite which multiplies within a parasitophorous vacuole or a cyst; (ii) two distinct parasitic stages i.e. tachyzoites and bradyzoites are present during the course of infection; (iii) parasites can invade different organs, mainly brain, striated muscles, lungs, eyes and placenta in case of maternal contamination.

I read with interest the paper by Ho et al. (1), which described a rapid prenatal diagnostic test for simultaneous detection of Hb Barts hydrops fetalis and exclusion of maternal contamination. The main background to their study is that PCR-based techniques are highly sensitive and are also prone to false-positive results in prenatal diagnosis due to amplification of contaminating maternal DNA that may be present in the fetal samples. Current established PCR-based diagnostic tests require a separate test to exclude maternal contamination. Hence, the authors developed a novel quantitative fluorescent PCR (QF-PCR) assay that allowed diagnosis and exclusion of maternal contamination in one step. Their approach seemed excellent, with a sensitivity of 100% and specificity of 100%. However, I believe that the presented data did not justify …

Aim To compare the results of low resolution typing by sequence specific primers and sequence specific Oligonucleotide probes on Luminex platform. Methods Prior to switching over to Luminex we performed initially low resolution HLA typing for thirty samples for ABDR antigens. We further validated HLA typing results on Luminex platform with two-third volumes of reagents. Kits from CPC Diagnostics (USA) and Innotrain (Germany) were used for SSOP and SSP typings respectively. We subsequently typed another twenty samples by both methods. Any addtional discreapancies encountered on typing over next two months shall also be presented. Results The results were concordant for all antigens at the A locus. One antigen was not identified at the B locus on typing by Luminex. Three typing results were discordant for DRB1 locus. There were no ambiguities in Class I typing on Luminex platform whereas six samples gave ambiguous results at B locus by SSP. We got confounding results by on Luminex platform for a sample of cord blood with maternal chimerism and also for DR typing of three samples. Conclusions SSOP typing gave fewer ambiguities for class I typing. For class II typing specially in cases of cord blood with maternal contamination we prefer SSP which gave clear results.

Background0 : Human chimerism is rare and usually uncovered through investigations of ambiguous genitalia or blood grouping or prenatal diagnosis. Most of the publications on placental chimerism are mainly case reports. There is no systematic search with sensitive techniques for placental chimerism in human. Aim0 : This study was aimed to asses placental chimerism through two sensitive molecular techniques i.e., interphase fluorescent in situ hybridization and quantitative fluorescent PCR. Material and methods0 : Placental chimerism was analyzed using X & Y dual color fluorescent in-situ hybridization onto 154 placentae from natural conceptions, obtained at termination of pregnancy between 7 to 16 weeks of gestation. Results0 : Three cases of placental sex chromosome chimerism were observed (1.95%). Exclusion of maternal contamination and diagnosis was confirmed later by quantitative fluorescent PCR. Conclusion0 : This finding indicates that placental chimerism in early human pregnancy is not rare.

This paper reports on 110 chromosome analyses of prenatal diagnosis after chorion biopsy. Normal findings were recorded in 105 cases (51 female, 54 male), and 5 chromosomic abnormalities were found, including triploidy, rarely to be observed in the second trimester, and a trisomy 8. Both direct preparation and long-term cultures were employed for chromosome diagnosis. With long-term cultures, three mosaic findings were recorded that were not confirmed in subsequent examinations of amniotic cells. The observation of certain cells with female karyotype in a long-term culture with otherwise definitely male karyotype illustrates the necessity of polymorphic comparison in order to exclude maternal contamination in the analysis of long-term cultures.