Prenatal care in the new era: 2014 - 15 update on earlier and less invasive individualized risk assessment

Vrachnis Nikolaos1, Galazios Georgios2, Stefos Theodoros3

1 2nd Department of Obstetrics and Gynecology, National and Kapodistrian University of Athens, School of Medicine, Aretaieio hospital, Athens, Greece

2Department of Obstetrics and Gynecology, Democritus University of Thrace, Medical School, Alexandroupolis, Greece

3Department of Obstetrics and Gynecology, University of Ioannina, Medical School, Ioannina, Greece



Prenatal care is the most widely used preventive service in the western world due to the recognition of its fundamental role in improving pregnancy outcome. The preventive nature of antenatal care, inspired by the teachings of Hippocrates, was re - introduced by John William Ballantyne in Edinburgh at the end of the 19th century, who focused on congenital anomalies, and later, during the first three decades of the 1900s, by John Whitridge Williams at the Johns Hopkins Hospital in Baltimore who championed the potential benefits of prenatal care for the diminution of neonatal deaths due to prematurity.

In 1929, the United Kingdom Department of Health published the first official recommendations on antenatal care, these stressing the need for a high concentration of visits during the last trimester of pregnancy when the complications of pregnancy were considered to be more likely to occur. However, scientific advances during the last few decades have enabled detection, and thus potential prevention, of many pregnancy complications from much earlier, i.e. before the end of the first trimester of gestation.

The past 40 years have thus witnessed such important developments as: maternal age - based prenatal screening for chromosomal abnormalities in the 1970s, the nuchal translucency screening test in 2003 and, towards the end of that decade, the combined test with maternal serum β- hCG and PAPP - A in the first trimester (11 - 13 weeks). This last was in particular a major breakthrough, resulting in a marked improvement in screening results, with the detection rate rising from 30% to 90%. Finally, the past three years have seen the introduction of an extremely high - performance screening test, massively parallel sequencing of maternal plasma cell - free fetal DNA (cffDNA testing). In a series of clinical implementation and validation studies, the latter method proved capable of accurately detecting fetal autosomal aneuploidy in 99% of the cases of Down syndrome, 97% of trisomy 18 and about 92% of trisomy 13, at an overall false positive rate or unnecessary invasive testing rate of 0.4%. The outcome has been the successful completion of screening for aneuploidies by the end of the first trimester of pregnancy. The information obtained from the combined test can with great precision identify those women who should have an invasive diagnostic procedure in the second trimester (chorionic villus sampling or amniocentesis), those who will be classified as having a high risk of trisomy, namely higher than 1/300, which is the expected risk for amniocentesis or CVS - related miscarriage. Meanwhile, women classified as having an intermediate risk (1/1,000 to 1/300) could be offered cff DNA testing before a final decision on an invasive procedure is made.

Furthermore, thanks to the 11 - 13 weeks’ gestation ultrasound screening, it is possible to diagnose the majority of severe fetal structural abnormalities in order to achieve an early as possible diagnosis, complementing the ultrasound scan performed in the second trimester (20 - 22 weeks). The early identification of such disorders enables couples to make a timely informed choice as to whether they wish to continue with the pregnancy or have a termination.

Preeclampsia complicates approximately 10,000,000 pregnancies each year around the world. Up to 15% of the over half a million women who die annually from pregnancy related causes, as well as at least 15% of premature births, are associated with hypertensive disorders of pregnancy. Although in most countries there are currently no official guidelines, there is evidence that mathematical models which combine maternal characteristics, mean arterial pressure, uterine artery pulsatility index and biochemical tests including the measurement of PAPP - A and PlGF (placental growth factor) at 11 to 13 weeks could potentially identify about 90% and 60% of pregnancies that subsequently develop into early (before 34 weeks) or late (after 37 weeks) preeclampsia, respectively, at a false positive rate of 5%. Interestingly, research has reported that even simple information easily obtained from the woman’s history could be extremely helpful: for example, being overweight or obese is alone responsible for almost two thirds of total preeclampsia cases. First trimester screening combined with the results from the uterine artery Doppler at 20 - 24 weeks and a series of new biochemical tests, including PlGF and sFlt - 1 (soluble fms - like tyrosine kinase - 1), can identify virtually all women who will develop the disorder. However, while identification of high - risk women in the second trimester could potentially improve the pregnancy outcome by means of intensive maternal and fetal monitoring, first trimester screening is now considered more valuable because there is evidence that the rate of preeclampsia as well as the rate of intrauterine growth restriction linked to impaired placentation in these women can be reduced by more than 50% if aspirin therapy is initiated at less than 16 weeks of gestation.

Preterm birth is a major cause of neonatal mortality and morbidity, has important long - term consequences in the offspring’s later life and is associated with a devastating economic impact. It is estimated that globally 15,000,000 neonates are born before 37 completed weeks of gestation, with preterm birth rates increasing in most countries. Prematurity is directly linked to more than 1,000,000 neonatal deaths each year, more than one third of the total burden of neonatal deaths worldwide, while it is also the leading cause of child death in the high- and middle-income countries of the world. Preterm birth is also a major public health concern in Greece, which in 2010 registered the highest preterm birth rate among European countries. Spontaneous preterm birth is a syndrome of multifactorial and mostly unknown etiology. Patient - specific risk of preterm delivery provided by maternal characteristics and obstetric history can predict only about a third of prematurity cases before 34 weeks. Secondary prevention focuses on recurrent preterm birth, which is the most important risk factor. In the high - risk group of women, measurement of cervical length in the second trimester could increase the identification rate to almost 65% and these women should be closely monitored as pregnancy proceeds. Recommended options based on individualized risk are elective cerclage at 13 to 16 weeks of gestation for high - risk patients after the first trimester assessment, and progesterone therapy for those classified as high - risk in the second trimester.

Gestational diabetes is a silent epidemic closely linked to the concurrent growing prevalence of obesity worldwide. It has been estimated that almost half of gestational diabetes cases are attributable to the increased body mass index (BMI of 25 kg/m2 or above) at the beginning of pregnancy. Unfortunately, the current prognostic models for gestational diabetes recommended for the first trimester attain relatively low detection rates. These models, combining certain maternal characteristics and history information (including pre - pregnancy body mass index, maternal age, personal or family history of diabetes and birth - weight in previous pregnancies) with certain biochemical parameters (such as adiponectin and sex hormone binding globulin), are potentially able to identify up to 75% of the subsequent cases of gestational diabetes - however, with a high false positive rate of more than 20%. This is explained by the existence of other factors that play an important role in the pathogenesis of the disorder and which occur later in pregnancy, particularly gestational weight gain, which has been shown to be an important independent risk factor for the disorder. Currently, the identification of high - risk pregnancies by the oral glucose tolerance test at the end of the second trimester and the subsequent treatment has been shown to be effective in reducing many important adverse pregnancy outcomes associated with gestational diabetes, including macrosomia, shoulder dystocia and gestational hypertension. Nevertheless, the results are disappointing for the long - term metabolic outcomes of offspring, which are typically associated with fetal pancreatic hyperplasia that occur during the second trimester. The objective thus appears to be the integration of the abovementioned model with a modified oral glucose tolerance test performed 10 weeks earlier than the current one in order to identify the high-risk group at the beginning of the second trimester and, through interventions, to promptly prevent occurrence of pancreatic hyperplasia in the fetus. Of note, with regard to the pharmacotherapy of gestational diabetes, there is a trend towards replacing the traditional insulin therapy with oral hypoglycemic agents. Already, over the past decade in the United States, glyburide has replaced insulin as the more common medication for gestational diabetes, especially among younger women.

Similar algorithmic approaches are also being developed for the early detection of women at risk for other pregnancy complications such as stillbirth and intrauterine growth restriction. Although currently there is no clinically useful first - trimester test to predict stillbirth and intrauterine growth restriction, risk assessment based on the uterine artery pulsatility index and maternal serum PAPP - A levels appears to be a good predictor of these disorders in cases where they are related to placental dysfunction disorders.

The philosophy of the new era in prenatal care translates as utilization of appropriate risk - as sessment algorithms from universal screening in the first trimester, followed by a final evaluation of women of intermediate risk in the second trimester. Application of the above will lead to the highest yield of high - risk pregnancies, these women subsequently being offered interventional diagnostic procedures and close monitoring with cost and side - effects minimization.


Prenatal care in the new era 2014 15 update on earlier and less invasive individualized risk assessment iphonef


The current approach to prenatal care places emphasis on early detection of high-risk pregnancies and less use of invasive diagnostic procedures for low-risk pregnancies. The recently issued Hellenic Society of Obstetrics and Gynecology (HSOG) Guideline “Prenatal care: surveillance of low-risk pregnancy” is published in Greek, but is also available at and can be downloaded as an application for tablets and smartphones. Users, both health professionals and the lay public, can access evidence-based information and find answers to their clinical questions just one click away on their mobile devices.

Vaginal birth after cesarean section in Greece and the contribution of the midwives

Nousia Konstantina1, Michalopoulos George2, Grigoriadis Charalampos3, Stournaras Stamatis1, Brezeanou Christina3, Vrachnis Nikolaos3, Farmakides George1

1 6th Department of Obstetrics and Gynecology, Elena Venizelou hospital, Athens, Greece

2 Lito Maternity hospital, Athens, Greece

3 2nd Department of Obstetrics and Gynecology, Aretaieio hospital, University of Athens Medical School, Athens, Greece

Correspondence: Farmakides George, 6th Department of Obstetrics and Gynecology, Elena Venizelou hospital, 2 Elena Venizelou Square, GR-11521 Athens, Greece

E - mail: This email address is being protected from spambots. You need JavaScript enabled to view it. 




Introduction: The aim of this study was to examine the success rate and safety of vaginal delivery after cesarean section, as well as the value of midwives’ contribution in these cases.

Material and Methods: This was a retrospective clinical study, including women diagnosed with singleton uncomplicated pregnancies and history of one or more previous cesarean sections, who underwent trial of labor, supported by midwives during the antepartum and intrapartum period.

Results: In total, 79% of this study group achieved vaginal birth after cesarean section. No major complications, such as uterine rupture or massive obstetric hemorrhage, were observed during the trial of labor in any case. There was no need for blood transfusion, emergency cesarean section or obstetric hysterectomy in this study group.

Conclusion: It seems that pregnant women who were appropriately informed and prepared from a midwife during the antepartum period achieved higher rates of vaginal birth and satisfaction from the trial. All pregnant women should be informed about the higher risk of complications after repeated cesarean sections in comparison with the potential risks of vaginal delivery after cesarean section and a trial of labor should be offered if they agree with the process in the absence of other obstetrical indications for cesarean section.

Key words: VBAC; TOLAC; cesarean section; midwife; Greece


During the last decades the incidence of cesarean section (CS) in Greece has dramatically increased. Unpublished data report cesarean section rates over 60% in several obstetric care departments. From 1994 to 2000 the incidence of primary cesarean section raised from 6% to 19% in selective University obstetrical departments1.  

Nowadays, the most common indication for elective CS is the obstetrical history of previous CS, leading to elevated total incidence of CS. However, the history of CS could not be considered as an indication for CS in all future pregnancies. Vaginal birth after cesarean section (VBAC) could be characterized as a safe, alternative option, in the absence of obstetrical contra - indications, that leads to decreased percentages of CS and offers satisfaction to the mother.

In addition, increased CS rates are reported in the United States of America as well, probably because of the decrease in VBAC rates during the same period2. At 1990’s a peak in VBAC incidence was noticed; however, during the last decades the percentage of pregnant women who underwent a trial of labor after CS (TOLAC) was lower.

The purpose of this study was to investigate the efficacy and safety of vaginal birth after cesarean section in low risk populations, as well as the value of midwifery contribution during the antepartum and intrapartum period.


Material and Methods

This was a clinical study organized under the cooperation of obstetricians and midwives from two private (Lito and Gaia hospital) and one public (Elena Venizelou hospital) obstetrical departments in Athens, Greece. All cases of TOLAC that took place in three departments (Lito, Gaia and Elena Venizelou hospital) between January 2013 and October 2014, were retrospectively analyzed. Written informed consent was provided in all cases.

During the antepartum period, midwives gave responsible and detailed information about the process of VBAC to the women. Additionally, midwives tried to offer emotional support to the pregnant women and to teach them position exercises and the breathing technique during labor, in order to prepare them for a successful TOLAC.

During the intrapartum period monitoring of the labor was performed. Pain management was achieved by encourangement of the midwives and included ambulation, showers, emotional support and changing positions of the mother. Ambulation and breast stimulation were used according to the individual’s needs. The obstetricians were responsible for the decision of labor augmentation, oxytocin administration or epidural anesthesia. In the presence of signs suspicious for fetal distress during the trial of labor, the decision for CS or continuation trial was responsibility of the obstetricians.

The primary outcome of this study was to estimate the total success rate of TOLAC. Moreover, the incidence of major reported complications during VBAC, such as uterine rupture or massive obstetrical hemorrhage, was investigated as well in this study group.



During the study period, 66 cases of TOLAC were examined. All were Greek pregnant women, with high level of education, aged between 27 - 40 years old (mean age 33.8 years), under the care of the midwifes and the obstetricians during the antepartum and intrapartum period. Three pregnant women who were included in this study group had an obstetric history of two previous CS. All were singleton, uncomplicated pregnancies with cephalic presentation.

The most common indication for CS at previous gestation was the failed induction of labor or the absence of labor progress (46%). Moreover, in 14% of cases fetal distress during the intrapartum period was the reason for the previous CS.

In total, the success rate of TOLAC in this study group was 79%, as 52 out of the 66 study group women achieved vaginal delivery. Labor started spontaneously in 89% of cases, while induction by breast stimulation and rupture of the membranes took place in 11% of cases. All infants had excellent Apgar scores at the first and fifth minute after labor, with a mean birth - weight of 3,410 gr (range: 2,500 - 4,300gr).

The majority of cases in which trial of labor was not successful were led to CS because of non progress of labor (11/14, 79%). Suspicious signs for fetal distress during the intrapartum monitoring led to CS in three cases.

No major complications such as uterine rupture or massive obstetrical hemorrhage were reported. There was no need for blood transfusion or obstetrical hysterectomy in any case. The mean hospital stay was 2.3 days (range: 1 - 5 days).



In our study study, 66 pregnant women with singleton, uncomplicated pregnancies after previous CS were led to TOLAC. All women were informed and appropriately prepared from midwives during pregnancy and a high success rate - comparable to other published studies - is reported (79%)3.

In agreement with other, previously published studies, it seems that women who receive detailed information and are prepared for VBAC by midwives throughout pregnancy achieve vaginal birth in higher percentages than pregnant women without appropriate preparation4.

It is true that the population of women who wish to undergo TOLAC has special needs during the antepartum and intrapartum period. An holistic approach that a certified nurse - midwife can offer, is very useful.

Moreover, the risk of complications after repeated cesarean sections, such as placenta previa or placenta accreta in future pregnancies associated with an elevated risk for massive hemorrhage and obstetrical hysterectomy, is high. The American College of Obstetricians and Gynecologists (ACOG) promotes the value of trial of labor after low transverse CS in the absence of obstetrical contra - indications, in an attempt to decrease the incidence of CS and associated complications5.

Furthermore, the fear of uterine rupture in case of spontaneous labor after CS leads a number of obstetricians to perform elective CS before the achievement of complete neonatal lung maturity; this practice is associated with an increase of infants’ admission to neonatal intensive care units. The ACOG recommends that the uncomplicated elective cs should not be performed prior to 39 weeks of gestation.

However, the abovementioned risk of uterine rupture during TOLAC remains low, being estimated at 6.2 cases per 1,000 vaginal births after CS6. In addition, the incidence of obstetric hysterectomy due to TOLAC is approximately 0.5 per 1,000 vaginal births after CS6. Although the incidence is low, all candidates should receive detailed information and give informed consent prior to the trial of labor after CS, as these changes are not acceptable by all women.

Nevertheless, TOLAC should be undertaken in organized obstetric units, under continuous cardiotocography, with high standards of obstetric, anesthesiology, surgical, neonatal and hematology care, capable to support emergency cesarean section or other complications.

After appropriate counseling the ultimate decision to undergo TOLAC or repeat CS should be made by the pregnant woman in consultation with the team of health providers. The midwife can contribute in order to increase the success rate of the trial7,8. The collaboration between obstetricians and midwives is crucial for the high quality care of the woman, according to the ACOG statement of policy8.

In conclusion, the midwife can be of a great help to educate, give psychological - emotional support and lead the pregnant woman to achieve a successful TOLAC.


Conflict of interest

All authors declare no conflict of interest



  1. Tampakoudis P, Assimakopoulos E, Grimbizis G, et al. Cesarean section rates and indications in Greece: data from a 24 - year period in a teaching hospital. Clin Exp Obstet Gynecol 2004;31:289-92. PubMed
  2. Martin JA, Hamilton BE, Ventura SJ, Osterman MJ, Mathews TJ. Births: final data for 2011. Natl Vital Stat Rep 2013;62:1-69. PubMed
  3. Harrington LC, Miller DA, McClain CJ, Paul RH. Vaginal birth after cesarean in a hospital - based birth center staffed by certified nurse - midwives. J Nurse Midwifery 1997;42:304-7. PubMed
  4. Avery MD, Carr CA, Burkhardt P. Vaginal birth after cesarian section: a pilot study of outcomes in women receiving midwifery care. J Midwifery Womens Health 2004;49:113-7. PubMed
  5. American College of Obstetricians and Gynecologists. ACOG Practice Bulletin no115: Vaginal birth after previous cesarian delivery. Obstet Gynecol 2010;116:450-63. PubMed
  6. The Royal Australian and New Zealand College of Obstetricians and Gynecologists. Planned vaginal birth after caesarean section. College Statement. C - Obs 38, October 2013.
  7. King T. Can a vaginal birth after cesarean delivery be a normal labor and birth? Lessons from midwifery applied to trial of labor after a previous cesarean delivery. Clin Perinatol 2011;38:247-63. PubMed
  8. ACOG College Statement of policy. Joint Statement of practice relations between Obstetrician - Gynecologists and certified nurse - midwives/certified midwives. July 2014

Dynamic spectral imaging colposcopy

Sofoudis Chrisostomos, Salakos Nikolaos

2nd Department of Obstetrics and Gynecology, University of Athens, Aretaieio hospital, Athens, Greece

Correspondence: Sofoudis Chrisostomos, 209 Hippokratous St, GR -11472, Athens, Greece. E - mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

Keywords: DySIS; colposcopy; high grade intraepithelial neoplasia; low grade intraepithelial neoplasia



Cervical cancer is the most common cause of death from gynecological cancer in developing countries, with a mortality rate ten times higher than that in developed countries. It represents the third most common cancer in women after breast and colorectal cancer, with 528,000 estimated cases in 2012, 84% of these in less developed countries1,2.

In terms of cervical cancer prevention the role of Pap smear remains unquestionable3. The Pap smear reflects a quick, easy, and relatively inexpensive method, which is used for screening all socioeconomic - level populations. Cervical cancer incidence rates have decreased dramatically since the implementation of the Pap smear screening. Nevertheless, the American Cancer Society estimates for 2013 predicted more than 12,000 new cases of cervical cancer in the United States4.

Colposcopy represents an alternative method concerning cervical cancer prevention. It refers to all abnormal cytological reports and abnormal cervical epithelial lesions. The final diagnosis is being established through histological confirmation of specimen collected from suspicious areas identified during colposcopy. During colposcopy, through the application of acetic acid, abnormal cervical epithelial lesions turn into white. This condition refers not only to intraepithelial neoplasias but also to several physiologic and benign conditions. The ultimate scope remains the detection of these lesions and the establishment of a suitable treatment plan5

Due to low sensitivity (55 - 65%) and specificity (70 - 90%) of colposcopy, an improved procedure must be conducted6. The Dynamic Spectral Imaging System (DySIS, by DySIS Medical Ltd, Livingston, UK) focuses on improving the performance of standard colposcopy by offering an objective way to quantify and map the acetowhitening areas. The DySIS cervical mapping offers a better optical evaluation and decreases biopsy sampling errors7. When used in combination with all other colposcopic indicators, it can improve the ability of the colposcopist, concerning not only the detection of intraepithelial lesions but also benign conditions such as metaplasia, inflammation and cervical infection8. In previous clinical studies, the use of DySIS managed to increase the low sensitivity and specificity of conventional colposcopy by introducing the cervical mapping9.

The DySIS colposcope has digital imaging equipment capable of sufficient visualization of the cervix, using a variety of color filters. It is able to detect and map acetowhite changes that may be linked to cervical epithelial lesions and at the same time consider all the parameters such as intensity and acetowhite persistence10. Color cervical mapping indicates with different colors the areas of suspicion based on their acetowhitening (Figure 1). Due to digital analysis, the colposcopist is able to correctly characterize abnormal areas, detect the difference between intraepithelial lesions and benign conditions, and select the biopsy locations accurately11.

In conclusion, DySIS appears to be a promising new technique for the evaluation of benign, low and high grade intraepithelial lesions. The increased sensitivity and specificity, in comparison with conventional colposcopy, provides assiduous cervical evaluation and mapping.


colp1 colp2 colp3
a b c

Figure. High grade intraepithelial neoplasia

a) reference image, b) after acetic acid application, c) Dynamic Spectral Imaging color-coded map



 Conflict of interest

All authors declare no conflict of interest




  1. World Health Organization. International Agency for Research on Cancer. Estimated cancer incidence, mortality and prevalence worldwide in 2012.
  2. Aggarwal P, Patra S, Gandhi G, Zutshi V. Can visual inspection with acetic acid under magnification substitute colposcopy in detecting cervical intraepithelial neoplasia in low-resource settings? Arch Gynecol Obstet 2011;284:397-403. PubMed
  3. NHS Cervical Screening Programme. Annual Review 2008.
  4. Cervical cancer overview. American Cancer Society 2013. Cancer / OverviewGuide/cervical-cancer-overview-key -statistics.
  5. Bekkers RL, van de Nieuwenhof HP, Neesham DE, Hendricks JH, Tan J, Quinn MA. Does experience in colposcopy improve indetification of high grade abnormalities? Eur J Obset Gynecol Reprod Biol 2008;141:75 - 8. PubMed
  6. Soutter WP, Diakomanolis E, Lyons D, et al. Dynamic spectral imaging: improving colposcopy. Clin Cancer Res 2009;15:1814 - 20. PubMed
  7. Jeronimo J, Sciffman M. Colposcopy at a crossroads. Am J Obstet Gynecol 2006;195:349 - 53. PubMed
  8. Alvarez RD, Wright TC. Increased detection of high grade intraepithelial neoplasia utilizing an optical detection system as an adjunct to colposcopy. Gynecol Oncol 2007;106:23 - 8. PubMed
  9. Desantis T, Chaktoura N, Twiggs, et al. Spectroscopic imaging as a triage test for cervical disease: a prospective multicenter clinical trial. J Low Genit Tract Dis 2007:11:18 - 24. PubMed
  10. Balas C. A novel optical imaging method for the early detection , quantative grading and mapping of cancerous and precancerous lesions of cervix. IEEE Trans Biomed Eng 2001;48:96 - 104. PubMed
  11. Craine BL, Craine ER, O’Toole CJ, Ji Q. Digital imaging colposcopy: corrected area measurements using shape - from - shading. IEEE Trans Med Imaging 1998;17:1003 - 10. PubMed

Non-invasive prenatal diagnosis and cell free DNA analysis

Kalinderi Kallirhoe, Fidani Liana

Department of Medical Biology and Genetics, Medical School, Aristotle University of Thessaloniki, Thessaloniki, Greece

Correspondence: Fidani Liana, Department of Medical Biology and Genetics, Medical School, Aristotle University of Thessaloniki, GR 54124, Thessaloniki, Greece

E-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.




Prenatal diagnosis is a very important part in the obstetric care of a pregnant woman. The use of invasive methods of prenatal screening provides great accuracy in the prenatal diagnosis of genetic and chromosomal abnormalities of the fetus; however it carries a small risk of fetal miscarriage. For this reason, the development of non-invasive methods of prenatal diagnosis is an area of intense research in which recently great progress has been made.

Keywords: prenatal diagnosis; cell free DNA




The term prenatal diagnosis refers to all the diagnostic tests and methods that achieve early intrauterine diagnosis of fetal anomalies or genetic disorders, providing the opportunity to the parents to terminate a pregnancy in case of a congenital abnormality. The main indications for prenatal diagnosis are increased maternal age (> 35 years), a previous pregnancy with fetal chromosomal abnormality, the presence of a balanced translocation or aneuploidy in a parent, increased risk of monogenic disease due to a positive family history, reproductive problems such as repeated pregnancy losses and exposure to potentially teratogenic factors. Prenatal diagnostic procedures can be invasive and non - invasive. The main invasive prenatal methods currently used are amniocentesis, carried out at 16 - 18 weeks of gestation and chorionic villus sampling, carried out at 8 - 11 weeks of gestation1. The small but finite risk of miscarriage after invasive prenatal procedures led to intensive efforts to improve methods of non - invasive prenatal diagnosis. In this review, the main methods of prenatal diagnosis are described, focusing on new advances in the field of non - invasive prenatal diagnosis.


Ultrasonography and maternal serum screening

Ultrasound scanning is one of the main methods of non - invasive prenatal diagnosis, as it is a safe and painless method for both the fetus and the mother. Ultrasound examination is performed during the first trimester of pregnancy, at 6 - 14 weeks of gestation and aims to confirm the existence of a pregnancy, to exclude the probability of an ectopic or molar pregnancy and to detect fetal viability. In the 11 - 14 week, fetal ultrasound scanning is used to measure nuchal translucency (NT). NT is the thickness of the fluid accumulated in the cervical area, between the fetal spine and skin. It normally has a size of 1 - 2 mm, while in cases of chromosomal abnormalities mainly trisomy 21 (Down syndrome) or severe heart disease, its size is > 3 mm. Specifically in Down syndrome, the NT measurement in combination with the maternal age and maternal serum levels of beta chorionic gonadotropin (β - hCG) and PAPP - A protein may lead to the detection of up to 92% of pregnancies with Down syndrome2. This percentage increases with the addition of other ultrasound markers, foremost of which is the absence / hypoplasia of fetal nasal bone3.

During the second trimester, between 20 - 24 weeks of gestation, another ultrasound (level II) scan is performed by a specialized obstetrician - gynecologist. With this, a detailed examination of all the organs of the fetus is done. If anomalies incompatible with life are diagnosed and the fetus has not exceeded the viability limit of 24 weeks, the possibility of interrupting the pregnancy is discussed. Some additional ultrasound markers, such as short femur and short humerus, the increased nuchal fold, dilatation of the renal pelvis, the echogenic bowel, echogenic cardiac foci and cysts of choroid plexus are used to further evaluate the possibility of chromosomal abnormalities. During the second trimester of pregnancy, between 16 - 20 weeks of gestation, the triple test (alfa test) can also be performed in order to check for chromosomal abnormalities, mainly for Down syndrome. Provided the precise determination of the gestational age, by measuring the biparietal diameter of the fetus, measurements of maternal serum levels of beta - hCG (β - HCG), alpha - fetoprotein (AFP) and estriol (E3) are made3. These measurements are recorded in a computer program and the probability of a chromosomal abnormality or an open lesion of the central nervous system is determined. With the triple test up to 67% of pregnancies with Down syndrome and a significant proportion of trisomy 18 and 13 can be diagnosed4. In case of a lesion of the central nervous system, the sensitivity of the method is 90%. The addition of inhibin - A measurement can improve the diagnostic sensitivity by approximately 7%. After the 24th week of gestation, uterine artery Doppler is also performed by measuring blood flow velocity. In case of increased resistance in both uterine arteries there is an increased risk of preeclampsia, placental abruption or intrauterine growth restriction (IUGR)3.

In the third trimester, with an ultrasound after 28 weeks of gestation, an estimation of fetal development is done based on measurements such as biparietal diameter, head and abdominal circumferences and fetal femur length3. If symmetrical IUGR is assessed, congenital or chromosomal abnormalities or intrauterine infections are suspected. In asymmetrical IUGR, there is increased probability of placental dysfunction and preeclampsia. In the third trimester of pregnancy the fetal biophysical profile is monitored by the movements of the fetal body and limbs, the fetal breathing, the fetal heart rate as well as the amount of amniotic fluid. Towards the end of pregnancy, ultrasound scanning is useful to check the position of the fetus and of the placenta and to measure the amount of amniotic fluid.

Apart from ultrasound and biochemical markers, non - invasive methods of prenatal diagnosis include the use of magnetic resonance imaging and analysis of fetal cells in maternal circulation, however these methods have serious drawbacks and its use is very limited, thus they will not be discussed in this review. The development of proteomics and the search for new biomarkers especially for aneuploidies such as trisomy 21 could help a lot in the field of non - invasive prenatal diagnosis, however so far no clear and valid results have come out5.

One of the most important recent achievements in the field of noninvasive prenatal diagnosis is the discovery of cell free fetal DNA in the maternal circulation6. The existence of cell free DNA in the maternal circulation makes it possible that in the near future genetic tests for prenatal diagnosis could be done with a blood test in the pregnant woman.


Cell - free DNA analysis

According to the discovery of Lo et al in 1997, about 10% of the DNA in the maternal circulation is of fetal origin, while the remaining 90% is maternal DNA6 - 8. Fetal DNA consists of small fragments of <313bp which can be detected in the maternal circulation from the 5th week9 and disappear two hours after birth10. The main source of cell free fetal DNA appears to be the apoptosis of trophoblast cells. Although the presence of a large amount of maternal DNA complicates the identification and analysis of fetal DNA, nevertheless today the analysis of cell free fetal DNA has already started to be used in clinical practice in the context of non - invasive prenatal diagnosis for the determination of sex, Rhesus genotype and recently for Down syndrome. The diagnosis of monogenic diseases and other aneuploidies may also be performed in clinical practice very soon.


Fetal sex determination

The first application of cell free fetal DNA was to determine fetal sex. More specifically, the detection of DNA sequences of the Y chromosome (SRY, DYS14 genes) in the maternal plasma indicates that the pregnant woman carries a male fetus. This knowledge can be used to prevent sex - linked diseases such as hemophilia and muscular dystrophy11. Conversely, in pregnancies where there are no detectable DNA sequences of the Y chromosome it is assumed that the fetus is female and in such a case the pregnant women does not need to undertake an invasive prenatal testing. The accuracy of the determination of fetal sex by analysis of cell free fetal DNA has been found to be > 95%, that is why its use in clinical practice was the first to be implemented and has led to a 50% reduction of invasive prenatal diagnostic methods in pregnancies with a high risk of sex-linked diseases12,13. Although, the free fetal DNA, can be detected almost by the fifth week of gestation, the validity of the results is 100% from the seventh week of gestation and afterwards14.

Timely and accurate knowledge of the sex of the unborn fetus is particularly useful in cases of congenital hyperplasia of the adrenal cortex as well. In this condition there is a shortage of the 21 hydroxylase, enzyme resulting in hyperandrogenism which in females results to the masculinization of the external genital organs15. To prevent this, corticosteroids can be administered16. Thus, the detection of fetal sex at 7 weeks of gestation, much earlier than by ultrasound scanning, is particularly useful in these circumstances.


Rhesus genotyping

It is well known that Rhesus D (RhD) (+)) fetuses carried by a RhD (-) mother are at risk of hemolytic disease after prior sensitization. Therefore anti - RhD globulin is prophylactically administered in RhD (-) pregnant women, blocking the process of sensitization. In the last 40 years, the administration of anti - RhD globulin has reduced the rates of death from hemolytic disease from 1/2,200 to 1/21,000 live births. Nowadays, prophylactic anti - RhD globulin is recommended in all RhD (-) pregnant17. Following this strategy, 38% of RhD (-) pregnant women that carry a RhD (-) fetus receive anti - RhD globulin without having any need of it18. Therefore it is of particular clinical importance to know the fetal RhD genotype before birth. The presence of RhD sequences in the plasma of an RhD negative pregnant woman indicates that the unborn fetus is RhD (+)12,19 - 22. By using cell free fetal DNA the accuracy of the detection of the fetal Rh genotype is 94.8%23. With the aid of cell free fetal DNA and the early identification of fetal Rh genotype, RhD (-) pregnant women with a RhD (-) fetus can avoid unnecessary administration of anti - RhD globulin. However, the benefit compared to the cost of cell free DNA prenatal diagnosis of fetal RhD genotype remains a matter of debate24,25.


Single gene disorders

Prenatal diagnosis is particularly useful for common monogenic diseases such as the thalassemias and cystic fibrosis, but also for rarer monogenic diseases in families with a positive family history. The non - invasive prenatal diagnosis using cell free fetal DNA initially focused on identifying autosomal dominant diseases of paternal origin26. In such cases, the detection or not of paternal mutations in the maternal plasma, can lead to the confirmation or exclusion of the inheritance by the fetus of an autosomal dominant disease transmitted from the father. In the case of an autosomal recessive disease or an autosomal dominant disease of maternal origin, the large amount of maternal DNA in the plasma (maternal DNA background), makes it difficult to determine the possibility of inheritance of a maternal mutation to the fetus. If the couple carries different mutations for an autosomal recessive disease and the fetus has inherited a normal allele from the father it will not express the disease, regardless of whether it has inherited a mutation from the mother. Greater difficulties are faced if both the father and mother carry the same mutation for an autosomal recessive disease. In such cases new methods have begun to be implemented such as the relative mutation dosage (RMD) method using digital PCR27-29. In this method, the amounts of the mutant and normal alleles at a particular locus are compared in the maternal plasma. This methodology is used mainly in cases where the mother is heterozygous for a known mutation. For a pregnant woman heterozygous for a mutation who carries a fetus heterozygous for this particular mutation, equal amounts of mutant and normal sequences are expected in the maternal plasma. If the fetus is homozygous for the mutation, more mutant compared to non - mutant sequences are expected to be detected in the maternal plasma, whereas if the fetus is homozygous for the normal sequences a larger proportion of non - mutated relative to mutated sequences are expected to be found30. In conclusion, the RMD method can diagnose autosomal dominant diseases inherited from the mother, whereas when this method is combined with techniques for the detection of mutations of paternal origin, autosomal recessive diseases can also be found, enabling the non invasive prenatal detection of all monogenic diseases simply by a DNA analysis test in the maternal plasma30.



Chromosomal aneuploidies, such as trisomy 21, are the most common reasons why couples proceed to prenatal diagnosis. The use of non - invasive prenatal diagnostic techniques in such cases would be very useful eliminating the small but finite risk of miscarriage. However, the use of cell free fetal DNA in the diagnosis of chromosomal aneuploidies presents certain difficulties such as the existence of a high rate of maternal DNA in the maternal plasma, which complicates the analysis of fetal DNA, as well as the fact that the fetal DNA is free and not within cells.

A strategy used to overcome these difficulties is to analyze nucleic acids specific to the fetus, such as mRNA expressed exclusively in the placenta or placental epigenetic markers specific for the studied chromosome. A considerable number of such markers have been used, including the placenta - specific 4 (PLAC4) and chromosome 21 open reading frame 105 (c21orf105) mRNA for the chromosome 2131,32, serpin peptidase inhibitor, clade B (ovalbumin), member 2 (SERPINB2) mRNA for chromosome 18 and hypermethylated holocarboxylase synthetase (HLCS) for chromosome 2133,34. These markers are specific for the fetus, as they contain fetal - specific single nucleotide polymorphisms (SNP) alleles, whereas they do not contain maternal - specific SNP alleles. In order to extract information about the chromosome dosage of free nucleic acid molecules specific for the fetus, two methods are used. The first method is the allelic ratio approach such as the RNA - SNP approach that determines the amount of heterozygous alleles in specific nucleic acid molecules. Therefore, if the RNA - SNP method for PLAC4 mRNA is applied in the maternal plasma for an euploid fetus, the amount of the PLAC4 mRNA alleles will be almost equal, while if the fetus has a trisomy 21 one of the alleles will be overrepresented in the maternal plasma31,32. Similar studies after analysis of SERPINB2 mRNA have been done for trisomy 18. The methodology of allelic ratio is applied to fetal epigenetic markers as well (epigenetic allelic ratio approach, EAR). More specifically, in the EAR method placental - specific epigenetic markers are used such as hypomethylated SERPINB2 on chromosome 18 and mRNA expressed in the placenta such as PLAC4 mRNA, and the ratio for specific alleles on the target gene can be determined. In general, the ratio of 1:1 for both alleles is expected in a euploid fetus, while ratios of 1:2 or 2:1 are expected for a trisomic fetus33. The main drawback of this method is that it can only be applied to heterozygous fetuses, thus multiple markers are needed to be tested in order to enhance the population coverage of the method.

The second method used is the EGG method (epigenetic - genetic approach)34. In this method, a fetal - specific epigenetic marker from the potentially aneuploid chromosome, e.g. HLCS for chromosome 21, is used as well as a fetal - specific genetic marker from a non - aneuploid chromosome, e.g. the Y chromosome for male fetuses or a SNP - allele of paternal origin34. HLCS gene located on chromosome 21 is hypermethylated in the placental tissue, from which the cell free fetal DNA comes from, while it is hypomethylated in maternal blood cells from where the majority of the maternal DNA originates. By using specific PCR which can identify the hypermethylated HLCS, fetal DNA is amplified and then is compared to a fetal - specific genetic marker. In cases of trisomy 21, the epigenetic / genetic ratio will be increased due to the extra chromosomal dosage derived from fetus35. The advantage of this method is that it does not require the placental epigenetic marker to contain a polymorphism.

Besides quantifying fetal - specific DNA sequences in the maternal plasma, an alternative approach is to directly detect fetal aneuploidy by determining the total (maternal + fetal) amount of the studied aneuploid chromosome in the maternal plasma and comparing it with other chromosomes in the maternal plasma36. For this purpose, specific quantitative methods such as digital and RMD PCR methodology are used. According to this strategy, if a pregnant woman carries a fetus with trisomy 21, an additional dose of chromosome 21 derived from the fetus will be present which means a 50% increase of the copies of the fetal chromosome 21. For example, in a sample from the plasma of a pregnant woman who carries a fetus with trisomy 21 and in which the fetal DNA is 12%, a 6% (half of 12%) increase is expected in the total number of DNA copies of chromosome 21 in this sample35. Thus, non - invasive prenatal diagnosis of trisomy 21 could be done by comparing the number of copies of a genetic region on chromosome 21 with the number of copies of another reference chromosome. For an euploid fetus, the dose of chromosome 21 is expected to be equal to the reference chromosome and for a fetus with trisomy 21 a 50% increase of fetal DNA is expected35.

Finally, a new method for the quantitative detection of DNA sequences from an aneuploid chromosome in maternal plasma is the massively parallel sequencing (MPS)37. With this method, sequencing of many small nucleic acid molecules in a sample is done and thereafter the chromosomal location of these molecules is determined. For example, a pregnant woman who carries a fetus with trisomy 21 will increase the percentage of specific gene fragments (sequences) of chromosome 21 in comparison to the data obtained from pregnancies carrying euploid fetuses. The use of MPS has been considered for the non - invasive prenatal diagnosis of trisomy 21 and is expected to be applied to other aneuploidies as well38,39. Currently, the major limiting factor for MPS is the high cost of this methodology. Nevertheless, considering that the cost of sequencing is expected to be lowered in the near future, the MPS methodology could become more affordable and could be widely applied in routine clinical practice in the context of non invasive prenatal diagnosis.



The development of non - invasive methods of prenatal diagnosis is a field of intense research in which great progress has already been achieved. The discovery of cell free fetal DNA in the maternal blood 17 years ago, has brought substantial changes and new opportunities in the context of antenatal screening. Currently, the analysis of cell free fetal DNA is used for determining fetal sex, Rhesus genotype and for identifying mutations of paternal origin. Sex determination has resulted in 50% reduction of invasive prenatal screening methods, while the identification of Rhesus genotype has limited the use of anti - D globulin to Rhesus D (-) pregnant women who carry Rhesus D (+) fetuses. Although the detection of aneuploidies and of all monogenic diseases by analysis of cell free fetal DNA has been described, a lot of research is carried out to find the best technical approach and methodology, in order to put these applications in routine clinical practice. Advances in technology as well as in molecular techniques are expected to lead to the implementation of new reliable and affordable methods of non - invasive prenatal diagnosis with the view that in the near future, prenatal testing could be performed with just a blood draw and a low cost DNA test.


 Conflict of interest

All authors declare no conflict of interest




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Malignant melanoma and pregnancy: A case report

Thanasas Ioannis

Department of Obstetrics and Gynecology, Trikala General hospital, Trikala, Greece

Corresponding author: Thanasas Ioannis, Settlement of Agia Triada 3, GR-42100, Trikala, Greece. E – mail: This email address is being protected from spambots. You need JavaScript enabled to view it. 




Malignant melanoma in pregnancy is rare but its incidence is rapidly increasing due to the growing trend for child-bearing in advanced age. A case of malignant melanoma diagnosed in the 25th gestational week and treated with local surgical resection is presented. The patient had an uneventful remaining pregnancy, delivered by caesarian section at 37 weeks and has been disease free for 1 year postpartum. Early diagnosis and prompt treatment can lead to a favorable outcome.

Key words: malignant melanoma; pregnancy; treatment.



Cancer during pregnancy is not uncommon, estimated to occur in approximately one case in 1,000 pregnancies1,2. Cancer cases associated with pregnancy have increased in recent years, primarily due to the increasing age of childbearing. Furthermore, more cancer cases are identified due to more diagnostic procedures in pregnancy. The malignancies most frequently observed in pregnancy are breast cancer and malignancies of the female genital tract, followed by melanoma.3

Malignant melanoma develops from melanocytes, cells of neural origin which migrate from the neural crest to skin sites and produce melanin. In the majority of cases (more than 90%), melanoma appears on the skin and constitutes the most severe form of skin cancer4, 5.The incidence of the disease varies across the geographical distribution of the population, with a higher annual incidence in Australia and New Zealand (40 to 60 cases per 100,000) and a lower frequency of 10 to 15 cases per 100,000 population per year in Europe and the United States6. The actual incidence of melanoma in pregnancy is unknown, but several authors have estimated it to be between 4 to 8 cases per 100,000 pregnancies, accounting for almost 8% of all malignant neoplasms that occur during pregnancy and 1% of all diagnosed melanoma cases7-14. A significantly increased risk has been reported for primigravida women especially those of advanced age at first birth15,16.


Case report

A primigravida woman at 25 weeks of gestation presented to the outpatient clinic of our department due to a painful cutaneous lesion localized in the right thigh. The lesion had been noticed by the patient since one week. Clinical examination revealed the presence of ulceration on a preexisting nevus, complicated by slight bleeding and itching. Due to the clinical suspicion of malignancy, the nevus was surgically excised and the histopathological examination confirmed the diagnosis of malignant melanoma of the skin, stage I. The patient was referred to a specialized center, where she underwent further surgical therapy without dissection of local lymph nodes or the application of other complementary treatment. The patient had a premature rupture of membranes in 37 weeks of gestation and gave birth to a healthy neonate with a birth-weight of 2,950 gr by cesarean section due to breech presentation. A year later, the patient remained disease-free.



Malignant melanoma during pregnancy is a rare life-threatening entity that requires multidisciplinary approaches in organized centers with the collaboration of obstetrician-gynecologists, surgeons, oncologists, radiotherapists and neonatologists. Currently, there is no consensus on the treatment of pregnant women with melanoma, however therapeutic approach should include saving mother’s lives by adequate treatment of curable malignancies and efforts to protect the fetus and the newborn from the harmful effects of cancer therapy.17

The“gold standard” in therapeutic approach of melanoma in pregnancy is surgical treatment. It can be performed safely in all trimesters of pregnancy, although it is preferable to be avoided in the first trimester, in order to minimize the risk of fetal loss.The type of surgery therapy depends on the stage of the disease.Wide surgical excision of the lesion, biopsy and examination of the sentinel node, is recommended, and if the sentinel lymph node is positive, a complete lymphadenectomy of recruited lymphatic station is indicated17. The main complications of surgical therapy of melanoma in a pregnant woman include increased risk of preterm delivery and intrauterine growth restriction as well as augmented perinatal morbidity and mortality18,19.

Radiotherapy as an adjuvant therapy to surgery should be postponed for the period after childbirth in order to avoid the exposure of fetus to high doses of radiation20,21.Chemotherapy must be displaced for the second or third trimester of pregnancy, since its use in the first trimester was associated with a significantly increased risk of spontaneous miscarriages and congenital abnormalities in the offspring. Dacarbazine in combination with other chemotherapeutic agents including bleomycin, vincristine and lomustine have been shown to be the most effective in the treatment of metastatic melanoma22,23. Dacarbazine has been assigned to pregnancy category C by the FDA. Although there are no control data in human pregnancy, a number of case reports has shown that the use of dacarbazine in the second trimester of pregnancy does not cause birth defects in the newborn24. The administration of chemotherapeutic drugs to pregnant women in second or third trimester of gestation has been associated with an increased risk of preterm birth, preeclampsia, intrauterine growth restriction and transient leukopenia in the neonate25,26. Finally, the use of targeted therapies, such as the use of the BRAF-inhibitor vemurafenib have a promising role in advanced disease27.

In contrast with earlier views, the termination of pregnancy in the case of a diagnosis during the first trimester, is now considered not to affect the prognosis of melanoma during pregnancy. Nevertheless,the couple should be informed about the therapeutic treatment required and the arising risks. Selective abortion in the first trimester is indicated only for women who are at the final stage of the disease28.



Malignant melanoma in pregnancy is a life-threatening disease with increasing incidence in pregnancy due to advanced maternal age. In the case presented, the patient suffered from a localized melanoma, outwent surgical treatment alone and the pregnancy outcome was excellent. Obstetricians should be alerted to the need for careful examination of pigmented lesions during pregnancy. Management depends on the stage of the disease, the time of diagnosis and patient preferences.


Conflict of interest

The author declares no conflict of interest




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