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verified independently, preferably in large multicentre studies. There are, however, several issues which will make an independent assessment difficult. These include the selection of candidate SNP loci, which is described in a patent not readily available to the research community. Additionally, how a quantitative procedure relying on numerous steps (preamplification, PCR, biotin-mediated capture, fluorescent labelling, electrophoresis, and digital imaging) can permit results involving 4 decimal places is unclear. Finally, this study relies on a former report wherein the authors suggested that the use of formaldehyde would increase the proportion of cellfree fetal DNA in maternal blood samples3—a feature which could not be reproduced in several independent studies.4,5 Hence this report in its current form could be too preliminary. The research activities of the Laboratory for Prenatal Medicine and Gynaecological Oncology at the Basel University Women’s Hospital Department of Research are funded by the EU (SH is the scientific director of the FP6 SAFE network) and the Swiss National Science Foundation. Additionally, a research agreement involving a patent transfer for the “non-invasive detection of fetal genetic traits” has been signed with Sequenom Inc, USA.
*Sinuhe Hahn, XiaoYan Zhong, Wolfgang Holzgreve
[email protected] Laboratory for Prenatal Medicine and Gynaecological Oncology, University Women’s Hospital Department of Research, University Hospital Basel, CH-4031 Basel, Switzerland 1
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Dhallan R, Guo X, Emche S, et al. A noninvasive test for prenatal diagnosis based on fetal DNA present in maternal blood: a preliminary study. Lancet 2007; 369: 474–81. Lo YM, Tsui NB, Chiu RW, et al. Plasma placental RNA allelic ratio permits noninvasive prenatal chromosomal aneuploidy detection. Nat Med 2007; 13: 218–23. Dhallan R, Au WC, Mattagajasingh S, et al. Methods to increase the percentage of free fetal DNA recovered from the maternal circulation. JAMA 2004; 291: 1114–19. Chinnapapagari SK, Holzgreve W, Lapaire O, Zimmermann B, Hahn S. Treatment of maternal blood samples with formaldehyde does not alter the proportion of circulatory fetal nucleic acids (DNA and mRNA) in maternal plasma. Clin Chem 2005; 51: 652–55. Chung GT, Chiu RW, Chan KC, Lau TK, Leung TN, Lo YM. Lack of dramatic enrichment of fetal DNA in maternal plasma by formaldehyde treatment. Clin Chem 2005; 51: 655–58.
Authors’ reply In researching methods to increase the proportion of fetal DNA recovered from maternal blood, and developing non-invasive methods for prenatal diagnosis, the main objective should be to contribute practical solutions to real-world problems. Blood samples must be routinely transported from laboratories or physicians’ offices to a testing facility and then processed, and there is great potential for maternal cell lysis because of mechanical shearing and degradation resulting from shipping, chemical processing, and centrifugation. Our formaldehyde method was developed to address this real-world problem. Addition of formaldehyde to maternal blood samples significantly increases the proportion of fetal DNA recovered from the plasma.1,2 Benachi and colleagues3 confirmed our results, reporting a range of 5·6–96% fetal DNA and a mean of 36·8% fetal DNA recovered from maternal plasma. We are perplexed by the apparent inability of others to practise this technique. Fetal genotypes were confirmed by genotyping parental genomic DNA. Paternal genotyping was used for reference purposes to reduce the number of single-nucleotide polymorphisms (SNPs) analysed in the plasma, and is not required for quantitative analysis of fetal DNA. We compared our results with amniocentesis or newborn reports from the clinical sites. Our aim is to develop a completely non-invasive test not reliant on analysis of invasively obtained samples. Presentation of methods as “non-invasive” when they clearly rely on analysis of invasively obtained tissues is misleading.4,5 Sinuhe Hahn and colleagues state that our approach “is inferior to that attained in a report in which cell-free fetal mRNA was used.” If a comparison is to be made with other methods, it should at least be done accurately. We sequentially analysed 60 samples, none of which were
excluded from our statistical analysis.2 In the paper cited in comparison as superior, Lo and colleagues collected and analysed 119 samples; however, 52 (44%) did not have a heterozygous phenotype at the locus of interest, and were removed from the dataset before statistical analysis. Lo and colleagues then reported successful identification of 55 of 57 normal samples and nine of ten trisomy 21 samples and estimated specificity and sensitivity based on 67 rather than the 119 samples collected, artificially improving their results. They fail to acknowledge that their methods were only diagnostic for 56% of the samples they collected. As discussed in our paper, it is evident that the number of SNPs visible in the plasma often, although not consistently, relates to the proportion of fetal DNA in the sample. An additional practical benefit of this method is its ability to be done in the first trimester, allowing for analysis of a follow-up sample should a low number of SNPs be identified in the initial sample. Furthermore, unlike methods based on quantification of one or few SNPs on one or few chromosomes,4,5 the approach we describe is unlimited in terms of the number of reference chromosomes and number of SNPs that could be added to the analysis. RD is the founder, chief executive officer, and chairman of the board of directors of Ravgen Inc, and a stockholder of that company. XG and SE are employed by, and have options to purchase stock in, Ravgen. MD is an unpaid member of Ravgen’s advisory board, and has options to purchase stock. PB is a member of the board of directors of Ravgen and has options to purchase stock. Ravgen Inc has been issued patents and has multiple patent applications pending for the methods described in this letter.
*Ravinder Dhallan, Xin Guo, Sarah Emche, Marian Damewood, Philip Bayliss
[email protected] Ravgen Inc, Columbia, MD 21045, USA (RD, XG, SE); Department of Obstetrics and Gynecology, York Hospital/WellSpan Health, York, PA, USA (MD); and Department of Maternal Fetal Medicine, Lancaster General Women and Babies Hospital, Lancaster, PA, USA (PB)
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Dhallan R, Au WC, Mattagajasingh S, et al. Methods to increase the percentage of free fetal DNA recovered from the maternal circulation. JAMA 2004; 291: 1114–19. Dhallan R, Guo X, Emche S, et al. A noninvasive test for prenatal diagnosis based on fetal DNA present in maternal blood: a preliminary study. Lancet 2007; 369: 474–81. Benachi A, Yamgnane A, Olivi M, Dumez Y, Gautier E, Costa JM. Impact of formaldehyde on the in vitro proportion of fetal DNA in maternal plasma and serum. Clin Chem 2005; 51: 242–44. Lo YM, Tsui NB, Chiu RW, et al. Plasma placental RNA allelic ratio permits noninvasive prenatal chromosomal aneuploidy detection. Nat Med 2007; 13: 218–23. Tong YK, Ding C, Chiu RW, et al. Noninvasive prenatal detection of fetal trisomy 18 by epigenetic allelic ratio analysis in maternal plasma: theoretical and empirical considerations. Clin Chem 2006; 52: 2194–202.
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Acceptance of abortion by doctors and medical students in Cameroon Restrictions on safe abortion remain a danger to women’s health in industrialised countries,1 but strike hardest in developing countries. Legal bans on abortion exist in virtually all African countries, where they contribute to 4·2 million unsafe abortions yearly, and to 12% of maternal deaths.2 Conscientious objection by health professionals can hinder safe abortion even where policies are liberal.3
We anonymously surveyed attitudes of medical students and physicians towards abortion in Cameroon—a sub-Saharan country where a population of 16 million is served by 1500 physicians practising mostly in public hospitals without universal insurance coverage. Our sample consisted of physicians attending the 2002 Cameroon National Medical Conference (n=300), and medical students at the Yaounde Medical School (n=400). The methods are reported elsewhere.4 101 preclinical medical students (51%), 95 clinical medical students (48%), and 110 physicians (37%) responded. Respondents were mostly Christian (91%, 93%, and 88%, respectively) and male (64%, 65%, 81%). The mean age of physicians was 35 years, and 43% were general practitioners and 14% gynaecologists or residents in gynaecology. Despite discomfort with voluntary abortion, most accepted abortion for medical reasons (figure). Acceptance of both voluntary and medical abortion increased with the level of medical education. Acceptance of voluntary abortion for economic reasons was low (12%, 8%, 13%). Opponents to voluntary abortion mostly cited religious reasons (85%, 74%, 53%).
We declare that we have no conflict of interest.
*Ambroise Wonkam, Samia A Hurst
[email protected]
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Department of Morphology and Pathology, Faculty of Medicine and Biomedical Sciences, University of Yaoundé I, Yaoundé, Cameroon (AW); *Department of Genetic Medicine and Development, Geneva University Hospitals, Rue Michel-Servet 1, CH 1211, Geneva 4, Switzerland (AW); and Institute for Biomedical Ethics, Geneva University Medical School, Switzerland (SAH)
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Maternal health was cited as a benefit both by proponents of voluntary abortion (71%, 74%, 79%) and medical abortion (57%, 59%, 94%). In Cameroon, carrying out abortion can lead to criminal charges. Physicians’ unexpectedly supportive views could spring from experience with complications of unsafe abortions. Liberalisation led to a drop in mortality due to abortion by 90% in South Africa.5 Maternal deaths are opening debates on liberalising abortion in Africa,2 and this should be fostered as an important public health goal. Women’s health in most developing countries is badly neglected. These bans, however, actively contribute to maternal harm. Abortion opponents sometimes believe that fetus’s interests trump any hardship for women. Mandating gestation, however, is ineffective. In Cameroon, experience seemed to increase awareness of this. Simply allowing clinicians to follow their conscience on this issue would improve women’s health. In the meantime, better access to information, contraception, and post-abortion care is urgent in African countries.
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Figure: Attitudes of Cameroonian medical students and physicians towards voluntary and medical abortion
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Wright AA, Katz IT. Roe versus reality— abortion and women’s health. N Engl J Med 2006; 355: 1–9. Moszynski P. Conference warns of epidemic of unsafe abortions in Africa. BMJ 2006; 332: 874. van Bogaert LJ. The limits of conscientious objection to abortion in the developing world. Developing World Bioeth 2002; 2: 131–43. Wonkam A, Njamnshi AK, Angwafo FF 3rd. Knowledge and attitudes concerning medical genetics amongst physicians and medical students in Cameroon (sub-Saharan Africa). Genet Med 2006; 8: 331–38. Jewkes R, Rees H, Dickson K, Brown H, Levin J. The impact of age on the epidemiology of incomplete abortions in South Africa after legislative change. Br J Obstet Gynaecol 2005; 112: 355–59.
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