Prenatal Screening: A Tool to Predict, Prevent, and Prepare

Abstract

There has been a considerable reduction in maternal mortality from 6 to 9/1000 live births and infant mortality from 100/1000 live births in the 1900s to less than 0.1/1000 live births and 7/1000 live births, respectively, in the 2000s. This is mostly due to nutritional improvement and obstetric and fetal medicine advancements. However, in the current era, prevention of mortality is not the only goal but also the prevention of morbidity. Thus comes the importance of prenatal screening, which would help us to predict and prevent maternal-fetal complications and in non-preventable conditions to prepare ourselves for optimal care of the mother and fetus. Prenatal screening is thus a test to detect potential health disorders in pregnant mothers or the fetus and to identify a subset who may need additional testing to determine the presence or absence of disease. It is done to categorize mothers into high-risk and low-risk pregnancies to prevent maternal complications, screen the fetus for aneuploidies, anomalies, and growth abnormalities, and decide on any indicated interventions and the time and mode of safe delivery so that an optimal perinatal outcome is achieved. Prenatal screening not only caters to identify fetal complications but also attempts to identify maternal complications early.

1. Introduction

In 1929, the Ministry of Health in the UK set forth guidelines advocating for pregnant women to be first seen in the antenatal clinics at 16 weeks, followed by 24 and 28 weeks visits, fortnightly till 36 weeks, and weekly until delivery [1, 2] (Figure 1a). However, in 2011, Prof. Nicolaides inverted this pyramid of prenatal care by introducing a new model where a comprehensive assessment of the mother and the fetus is done at 11–13 weeks. According to this “inverted pyramid model”, combining the data from maternal characteristics and history along with biophysical and biochemical tests performed on the mother can define the patient-specific risk for a variety of pregnancy complications, namely aneuploidies, preeclampsia, preterm delivery, gestational diabetes, fetal growth restriction, and macrosomia [5]. (Figure 1b). In 2017, Ljubic proposed an “extended inverted pyramid of care” based on the concept that the roots of these disorders are dysfunctional placentation and thus must be sought in the earlier period of pregnancy and in the deeper, subcellular level [6]. This means that the changes that lead to insufficient implantation should be sought in the preimplantation period, in the relation between the embryo and endometrium. Prepregnancy approaches such as optimizing maternal comorbidities, adequate weight management, blood pressure and glycemic control, smoking cessation, and spacing pregnancies may improve the placentation leading to an optimal pregnancy outcome [7]. Year 2016 saw the emergence of another model put forward by Moshe and Nicolaides called the “Three-floor model” where the care is extended to prepregnancy and postnatal periods [4] (Figure 1c). This model was proposed based on the concept that the health effects of women and their offspring are mediated by epigenetic and genetic pathways contributing to the increased risk of developing non-communicable diseases (NCD) that are passed onto the next generations, which is a vicious cycle. Thus, this model of care helps in the assessment of NCDs in the prepregnancy period (first floor) and optimizing the disease state followed by the inverted pyramid of care (second floor) starting at 11–14 weeks till delivery, to the postnatal period (third floor) where appropriate management can minimize the long term harmful effects in both mother and her offspring. Hence, this model of prenatal screening, which starts from the prepregnancy period and continues through pregnancy into the postnatal period, would be an ideal screening model, and this would reduce the harmful effects of the epigenetic/genetic factors on the fetus and the mother, thereby reducing the long term development of NCDs.

Figure 1.

(a) Pyramid of care in 1929, (b) inverted pyramid of prenatal care (adapted from [3]), (c) three floor model which includes prepregnancy and postpregnancy care (adapted from [4]).

In this chapter we shall discuss the different screening methods which can be applied to these three floors of prenatal care:

1. First floor—Prepregnancy period

2. Second floor—Inverted pyramid of care from 11 weeks till delivery

3. Third floor—Postnatal period

2. Prepregnancy period

Prepregnancy care aims to identify the women with NCDs and treat them and optimize their disease state. This is done by the family physician or primary obstetrician by gathering patient information either through personal interviews or from electronic records regarding medical, pregnancy, and family history, drug intake, and smoking. A physical examination is done to calculate the BMI and BP and investigations like HbA1C and total cholesterol are conducted. Seven cardiovascular health (CVH) metrics proposed by the American heart association (AHA) are assessed which include four health behaviors (weight, physical activity, smoking, and diet) and three health risk factors (blood pressure, fasting blood glucose, and total cholesterol). Based on these metrics patients are stratified into different risk categories, namely—ideal [8, 9, 10, 11, 12], intermediate [4, 7, 13, 14, 15], and poor (0–4) categories [13]. Two points are awarded for ideal, one point for intermediate, and zero points for poor, ranging from 0 to 14 [13].

Women who score ideal risk are reassured and advised to plan their families. Those women who score intermediate risk should be referred to either dieticians or physical trainers to optimize their pregnancy issues at hand. The poor score women are referred to a maternal-fetal medicine (MFM) specialist to optimize comorbidities like anemia/hypertension/diabetes, evaluation of the end organs in chronic morbidities, conversion to pregnancy-safe medications, screen for infections, and immunization of varicella/rubella and hepatitis B. Carriers of inherited genetic disorders should be offered counseling and workup by geneticist including index child workup, genetic evaluation for carrier status, and preimplantation genetic diagnosis (PIGD). Periconceptional folic acid should be advised as and where indicated.

3. Inverted pyramid of care

In the 11–14 weeks period, a comprehensive evaluation of the mother is done based on the demographics, medical/obstetric, and family history along with biophysical markers like mean arterial pressure (MAP), biochemical markers like human chorionic gonadotrophin (HCG), pregnancy-associated placental plasma protein A (PAPP-A) and placental growth factor (PLGF), and ultrasound (USG) parameters like nuchal translucency (NT) and uterine artery Doppler pulsatility index (UTPI) thus quantifying the woman’s risk for developing any chromosomal aberrations, preeclampsia, spontaneous preterm birth, fetal growth restriction, and gestational diabetes. Thus, they are stratified as low-risk and high-risk pregnancies. Low-risk mothers enter the routine care regimen and high-risk mothers enter the specialist care regimen.

The inverted pyramid of care thus includes:

1. Aneuploidy screening

2. Preeclampsia/SGA screening

3. Screening for preterm labor

4. Screening for diabetes

5. Screening for anomalies

3.1 Aneuploidy screening

Aneuploidy screen has come a long way since its inception in the 1970s. Early detection of Down syndrome is the main objective of prenatal aneuploidy screening since this syndrome is the most common autosomal trisomy among live births which is compatible with life. Trisomy 21 affects 1 per 500 pregnancies with a live birth prevalence of 1 per 740 while trisomy 18 occurs in 1 per 2000 pregnancies and 1 per 6600 live borns, and trisomy 13 is identified in 1 per 5000 pregnancies and 1 per 12,000 live borns [14, 15]. As detection of aneuploidies is also observed in younger age groups, screening is universal in the current era and all pregnant women should be offered screening for aneuploidies. There are two different types of aneuploidy screening:

1. Conventional screening

2. Cell-free DNA-based screening

Conventional screening is the established method of screening using NT performed by USG along with biochemical screening in the first trimester and only biochemical screening in the second trimester. It is further divided into three types:

1. First trimester combined screening

2. Second trimester biochemical screening

3. Combinations of first and second trimester screening: integrated and sequential screens

Cell-free DNA screening which was implemented in 2011, identifies circulating DNA fragments that are primarily placental in origin, from apoptotic trophoblasts [8, 9] and is considered to be the best available screening test with a good positive predictive value, and a very low false positive rate especially when applied appropriately.

3.1.1 Conventional screening

3.1.1.1 First trimester combined screening (FTS)

Components of FTS are as follows:

1. Pretest counseling

2. USG evaluation

3. Biochemical screening/Risk assessment

4. Posttest counseling

3.1.1.1.1 Pretest counseling

Every pregnant woman is counseled regarding the options of undergoing a screening test, diagnostic test, or no test at all, their detection rates and it is completely her choice to proceed with any testing. She is counseled that the purpose of the screening test is to provide information and if the test results come positive, it does not mean that the fetus is affected (false positive) and there is the option of diagnostic testing to confirm the same. Decisions cannot solely be taken based on screening tests. Similarly, it does not mean the fetus is unaffected if the test results come negative (false negative). The benefits of diagnosis, early intervention if affected and the costs of the screening and diagnostic tests are also explained. Additional evaluation and counseling are suggested if a patient has had a previous fetus or neonate with autosomal trisomy, Robertsonian translocation, or other chromosomal abnormality.

3.1.1.1.2 USG evaluation

There has been a paradigm shift in utilizing USG for the detection of aneuploidy markers to early identification of structural and genetic abnormalities. The salient applications of USG in the first trimester are:

1. To calculate the GA

2. To identify multiple gestations and their chorionicity

3. To identify aneuploidy markers

4. To identify major structural malformations

3.1.1.1.2.1 Calculation of GA

Dating is of paramount importance before aneuploidy testing because each screening test is valid only within a specific gestational age window, 11–14 weeks for first trimester screening and 15–21 weeks for second trimester screening. Moreover, when risk assessment is done each component of a screening test should be adjusted for gestational age when calculating multiples of the median, and false positive rates are reduced when gestational age is assessed by USG [10]. Crown-rump length (CRL) is the length of the embryo or fetus from the top of its head to the bottom of the torso (Figure 2a). Popularly called the Robinson’s CRL curve, it is the most accurate estimation of gestational age in early pregnancy, owing to the little biological variability at that time. Thus, CRL measurement has become the universal pregnancy dating tool to avoid the last menstrual date recall error [11, 12]. If the GA is >14 weeks then head circumference (HC) is used for dating the pregnancy (Figure 2b).

Figure 2.

Dating parameters. (a) Midsagittal plane of a fetus showing measurement of -rump length (CRL)-dating parameter <14 weeks GA, (b) measurement of head circumference (HC)-dating parameter >14 weeks GA.

3.1.1.1.2.2 Diagnose multiple pregnancies and determination of chorionicity

Overall, twin pregnancies are at higher risk than singleton pregnancies for aneuploidy. This is mostly due to advanced maternal age in twin pregnancies. Determination of chorionicity of a twin pregnancy is of paramount importance, and the first trimester assessment has a better accuracy rate of 96–100% versus approximately 80% in the second trimester [16, 17, 18, 19]. Chorionicity, rather than zygosity, has a major impact on the outcome of twin pregnancies mainly because of specific complications secondary to placental anastomoses, such as twin-to-twin transfusion syndrome (TTTS), selective fetal growth restriction (sFGR), twin anemia polycythemia sequence (TAPS), twin reversed arterial perfusion (TRAP) [20, 21]. The presence of the lambda sign (due to the interposed chorionic tissue) (Figure 3a) is suggestive of DCDA twins and the presence of the T sign is suggestive of MCDA twins (Figure 3b).

Figure 3.

Assessment of chorionicity. (a) Lambda sign-DCDA twins, (b) T sign-MCDA twins.

3.1.1.1.2.3 Identification of aneuploidy markers

Nuchal translucency (NT) is a subcutaneous collection of fluid between the skin and soft tissue overlying the fetal spine at the back of the neck in the sagittal plane. It is the most important marker used in the first trimester for aneuploidy risk calculation. This was demonstrated by Nicolaides in the early 1990s and was found to be strongly associated with fetal aneuploidy [22, 23]. Increased NT is associated with trisomy 21/13/18, Turner syndrome and other chromosomal defects, fetal structural malformations, and genetic syndromes. Though NT tends to resolve, it can evolve into increased nuchal fold thickness or cystic hygromas with or without hydrops. Figure 4a and b is representative of normal and increased nuchal translucency, respectively.

Figure 4.

Measurement of nuchal translucency (NT). (a) Sagittal section of fetus showing normal nuchal translucency—1.8 mm, (b) sagittal section of fetus showing increased nuchal translucency—4.1 mm.

NT must be accurately imaged and measured in a reproducible way following the standards put forth by the Fetal Medicine Foundation and Perinatal Quality Foundation for aneuploidy detection to be accurate. The optimal gestational age for measurement of fetal NT is 11–13 + 6 weeks when the fetal CRL is between 45 and 84 mm. Measurement is done in the sagittal plane with the neck in a neutral position, and the image is magnified so that the screen is filled with the fetal head, neck, and upper thorax. The calipers are placed on the inner borders of the widest aspect of the nuchal space, perpendicular to the long axis of the fetus, with the horizontal crossbar within the space [24]. Though there is no clarity in the definition of increased NT beyond the cutoff of 3.5 mm, NT is said to be increased when it is >99th centile for the CRL or > 1.9 MOM for the measured CRL [25]. The causes of increased NT [23, 26] are cardiac defects and dysfunction, venous congestion in the head and neck, the altered composition of the extracellular matrix, failure of lymphatic drainage, fetal anemia, fetal hypoproteinemia, and fetal infection [26].

3.1.1.1.2.4 Other aneuploidy markers

3.1.1.1.2.4.1 Heart rate

In normal pregnancy, the fetal heart rate (FHR) increases from about 100 bpm at 5 weeks of gestation to 170 bpm at 10 weeks and then decreases to 155 bpm by 14 weeks. Between 11 and 14 weeks, trisomy 13 and Turner syndrome are associated with tachycardia, whereas bradycardia is noted in trisomy 18 and triploidy. Inclusion of FHR is important in differentiating trisomy 18 and 13, which in other respects show common features like increased fetal NT and decreased maternal serum free B hCG and PAPP-A [26].

3.1.1.1.2.4.2 Nasal bone

Nasal bone (NB) assessment is done between 11 and 13 + 6 weeks when the CRL is between 45 and 84 mm. In the midsagittal view of the fetal profile, NB is imaged as three distinct lines. The top line represents the skin and the bottom one, which is thicker and more echogenic than the overlying skin, represents the NB. A third line, almost in continuity with the skin, but at a higher level, represents the tip of the nose. At 11–13 + 6 weeks the NB is absent in 1–3% of euploid fetuses, 60% of fetuses with trisomy 21, 50% of fetuses with trisomy18, and 40% of fetuses with trisomy 13. Assessment of the NB improves the performance of combined screening, increasing the detection rate from 90% to 93% and decreasing the false positive rate from 5% to 3% [26]. Figure 5a and b shows the ossified and unossified nasal bone, respectively.

Figure 5.

Assessment of NB. (a) NB—seen as the bottom line, (b) unossified nasal bone—absence of bottom line.

3.1.1.1.2.4.3 Tricuspid regurgitation (TR)

TR assessment is done between 11 and 13 + 6 weeks when the CRL measures 45–84 mm. No regurgitation should be noted across the fetal tricuspid valve during systole. Regurgitation is significant if it occurs in more than half of systole with a velocity of more than 60 cm/s and is noted in about 1% of euploid fetuses, 55% of fetuses with trisomy 21, and 30% in fetuses with trisomy 18 and trisomy 13. Figure 6a and b shows normal tricuspid flow pattern and tricuspid regurgitation, respectively. Assessment of the tricuspid flow improves the detection rate from 90% to 95% and decreases the false positive rate from 3% to 2.5%. If there is tricuspid regurgitation, it is pertinent that a detailed cardiac evaluation is done to diagnose or exclude major cardiac defects [26].

Figure 6.

Assessment of tricuspid flow. (a) Axial section of the fetal thorax with spectral wave Doppler imaging showing normal flow pattern across the tricuspid valve, (b) axial section of the fetal thorax with spectral wave Doppler flow showing tricuspid regurgitation.

3.1.1.1.2.4.4 Ductus venosus flow

The ductus venosus is a short trumpet-shaped vessel that shunts oxygenated blood preferentially to the fetal heart. It has a triphasic pattern and the forward ‘a wave corresponds to the atrial systole. Reversed a wave in ductus venosus is associated with an increased risk for chromosomal abnormalities, cardiac defects, and fetal death. At 11–13 + 6 weeks, abnormal ductal flow is observed in 5% of chromosomally normal fetuses and about 80% of fetuses with trisomy 21. Assessment of ductus venosus improves the performance of combined screening, increasing the detection rate from 90% to 95% and decreasing the false positive rate from 3% to 2.5%. Though in 80% of cases with isolated reversed a wave the pregnancy outcome is normal, a detailed cardiac evaluation should be done to rule out cardiac defects in fetuses with an a-wave reversal in ductus venosus (Figure 7a and b) [26]’.

Figure 7.

Assessment of ductus venosus (DV) flow. (a) Axial section of the fetal abdomen with spectral wave Doppler flow showing normal ductus venosus flow with normal ‘a’ wave, (b) axial section of the fetal abdomen with spectral wave Doppler flow showing abnormal ductus venosus flow with reversed ‘a’ wave.

3.1.1.1.2.5 Identification of structural anomalies

There has been a paradigm shift from the identification of fetal aneuploidies in the 1980s to the early identification of structural/genetic abnormalities due to innovation in USG imaging technology. The commonly identified anomalies are given below: Acrania, alobar holoprosencephaly, body stalk anomaly, omphalocele, megacystis, and early hydrops. Figure 8a–f depicts these commonly diagnosed first trimester anomalies.

Figure 8.

Commonly identified anomalies in the first trimester. (a) Acrania, (b) alobar holoprosencephaly, (c) body stalk anomaly, (d) omphalocele, (e) megacystis, (f) early hydrops.

3.1.1.1.3 Biochemical screening/risk assessment.

Every pregnant woman has a background or apriori risk to bear a fetus with aneuploidy. This is based on her age and history of aneuploidy. The risk for trisomies increases with maternal age, but Turner syndrome and triploidy do not change with maternal age. The patient-specific risk is calculated by multiplying the apriori risk with a composite likelihood ratio, which is obtained from the screening tests performed during the pregnancy in the first and second trimesters. Each time a test is carried out the apriori risk is multiplied by the likelihood ratio of the test to calculate a new risk, which subsequently becomes the apriori risk for the next test.

Aneuploid pregnancies are associated with altered maternal serum concentrations of various fetoplacental products, namely HCG and PAPPA. First trimester combined screening integrates nuchal translucency with maternal serum HCG and PAPPA. HCG assay can be either intact HCG or free beta unit of HCG depending on the local laboratory guidelines and clinical practice, albeit the two are considered comparable [23]. Individual analytes are converted to multiples of the median (MOM) adjusting for maternal age, maternal weight, smoking status, ethnicity, and gestational age. The pattern of increase or decrease in analyte levels affects the risk for trisomies 21, 18, and 13 and at a predetermined value the test is deemed positive or abnormal. For trisomy 21, this is often presented as a first trimester risk of 1:250 and for trisomy 18 and trisomy 13, a cut-off of 1:150 is used (Table 1).

Maternal	Fetal
Risk stratification into high/low risk if not done already	Determine the age of the fetus
Identify women who need additional care	Assess chorionicity in twins and higher-order multiples
Screening for preeclampsia	Screening for aneuploidies/anomalies
Screening for preterm labor	Screening for small for gestational age (SGA)

Table 1.

Indications of 11–14 weeks assessment.

In euploid pregnancies, the average maternal serum HCG is 1.0 MOM and PAPP-A is 1.0 MOM. In trisomy 21 pregnancies, maternal serum HCG is increased by twofold and PAPP-A is reduced to half compared to normal pregnancies. In trisomies 18 and 13, maternal serum HCG and PAPP-A are decreased. In cases of sex chromosomal anomalies, maternal serum HCG is normal and PAPP-A is low. In paternally derived triploidy, maternal serum free HCG is greatly increased, whereas PAPP-A is mildly decreased. Maternally derived triploidy is associated with markedly decreased maternal serum HCG and PAPP-A [26, 27]. Table 2 depicts the analyte levels in various fetal trisomies in the first trimester and screening by a combination of fetal NT and maternal serum PAPP-A and HCG can identify about 90% of all these chromosomal abnormalities for a false positive rate of 5% [26, 27]. Table 3 depicts the detection rates and false positive rates of various combinations of screening tools and markers used in the first trimester [26].

Analytes	Trisomy 21	Trisomy 18/T13	Paternally derived triploidy (Type I)	Maternally derived triploidy (Type 2)
NT	Increased	Increased	Increased	Normal
HCG	Increased	Decreased	Marked increase	Marked decrease
PAPPA	Decreased	Decreased	Mild decrease	Marked decrease

Table 2.

Multiple marker levels associated with trisomies in the first trimester.

Screening tool	Detection rate (%)	False positive rate (%)
Age	30	5
NT alone	60	5
Combined (NT + biochemistry)	85–90	5
NT + NB	90	5
NT + NB + biochemistry	95	3
NT + NB + DVflow + TR + biochemistry	95–96	2–3

Table 3.

Detection and false positive rates of different combinations of screening tools and markers used in the first trimester. Modified from [26].

As the biochemical analytes (HCG and PAPPA) are increased twofold in twin pregnancies, the performance of combined screening is 15% lower when compared to singleton pregnancies [28].

3.1.1.1.4 Posttest counseling

Posttest counseling is mandatory for any screening program. When a positive or negative screening test result is obtained, the patient should be counseled regarding the adjusted likelihood of carrying a fetus with the aneuploidies evaluated and a diagnostic test is offered. The possibility for the fetus to be affected by genetic disorders which are not evaluated by the screening or diagnostic test should also be reviewed. In the event of a prenatal diagnosis of fetal aneuploidy, the patient and family should be counseled appropriately so that she can make informed decisions regarding further pregnancy management.

3.1.1.2 Second trimester screening

The triple screen which includes the analytes alpha feto protein (AFP), HCG, and serum estriol has a poor detection rate of 60% and was used in the 1980s. The quadruple test which was formulated by the addition of dimeric inhibin A to the triple screen in the 1990s replaced it and is the only current second trimester multiple marker screening tests widely used. A quadruple screen is performed between 15 and 21 weeks when the Biparietal diameter (BPD) is between 34 and 52 mm, the measurement varying between labs. The pattern of change in analytes is depicted in Table 4 [29].

Analytes	Trisomy 21	Trisomy 18
AFP	Decreased	Decreased
Estriol	Decreased	Decreased
HCG	Increased	Decreased
Inhibin	Increased	Not applicable

Table 4.

Multiple marker levels associated with trisomies in the second trimester.

Since the early 2000s, the quadruple test detection rate is 81–83% at a 5% false positive rate in two large prospective trials- the Serum, Urine, and Ultrasound Screening Study (SURUSS) [24] and the First and Second Trimester Evaluation of Risk (FASTER) [10] trials.

As the second trimester quadruple marker screening offers no advantage over first trimester screening, it is used only if first trimester screening is unavailable in certain settings or if the antenatal woman books too late to receive first trimester screening.

3.1.1.3 Combination of first and second trimester screening

This is based on the concept that aneuploidy detection will be significantly increased if first trimester biochemistry and nuchal translucency screening is combined with a second trimester quadruple marker test. However, these two tests should not be done as independent tests as it increases the false positive rate thus making counseling difficult. There are two different methods of screening—integrated test and sequential screening, which are further subdivided into stepwise and contingent sequential screening.

Integrated screening involves testing the first trimester serum analytes (HCG and PAPPA) and NT at 11–14 weeks followed by quadruple screening between 15 and 21 weeks of GA and a single risk is calculated. This integrated approach has a 94% sensitivity in detecting T21 and 93% detection of T18 [30] and the result was abnormal in 93% of cases with trisomy 13, in 91% with triploidy, and 80% with monosomy. Serum integrated screening is done when NT is unavailable and only the biochemistry is taken into consideration. This has a detection rate of 85–88% for T21 at a 5% false positive rate [24].

In sequential screening, first trimester screening with nuchal translucency and serum analytes is performed and the patient is informed about the results based on the understanding that, if the risk exceeds a predetermined cut-off (≥1 in 30), she will be offered diagnostic testing. There are two types of sequential screening. With stepwise sequential screening, women at high risk in the first trimester are offered diagnostic testing and the rest go on to complete the quadruple marker screening in the second trimester, after which the women who are screen positive are offered diagnostic testing and the rest are reassured and no further testing is indicated. Based on data from the FASTER trial, stepwise sequential screening using a first trimester risk cut-off of 1:30 and an overall cut-off of 1:270 yielded a 92% trisomy 21 detection rate at a 5% false positive rate [24]. In contingent sequential screening, after women at high risk are offered a diagnostic test, the remaining women are divided into two groups. The lowest risk women (<1:1500) are reassured and receive no further screening, whereas those at intermediate risk (1 in 270 to 1 in 1500) are followed up with quadruple marker screening. Based on data from the FASTER trial, the trisomy 21 detection rate was 91% with the contingent screening at a 5% false positive rate [24].

3.1.2 Cell-free–based DNA screening

Noninvasive prenatal screening (NIPS) or cell-free–based DNA screening was introduced into the armamentarium of aneuploidy screening in 2011. This test identifies circulating DNA fragments that are primarily placental in origin, from apoptotic trophoblasts. Assaying of cell-free DNA is done in three ways for aneuploidy screening: whole-genome sequencing (massive parallel shotgun sequencing-MPSS); chromosome selective sequencing (targeted); and single nucleotide polymorphism analysis (SNP). It can be performed after 9–10 weeks of gestation, and the turnaround time of the results is within 7–10 days. The detection rate is 99% for trisomy 21, 96% for trisomy 18, and approximately 90% for trisomy 13 and monosomy X [8, 9]. According to a meta-analysis of 37 studies of cell-free DNA screening in high-risk pregnancies, the pooled sensitivity to detect trisomy 21 was 99.2% (95% confidence interval 98.5–99.6%), and the specificity was 99.9%, and the false positive rate was only 0.1% [29, 31]. Detection rates of trisomies 18 and 13 are 96% and 91%, respectively, each with a specificity of 99.9%. For detection of monosomy X (Turner syndrome), the sensitivity of cell-free DNA was approximately 90% with a specificity of 99.8% [29, 31, 32, 33, 34, 35]. Table 5 depicts the different characteristics, the detection rates, the false positive rates, and the positive predictive values of the different screening tests to detect T21 [29].

Screening test for T21	Sensitivity %	False positive rate %	Positive predictive value %
First trimester screening (NT + biochemistry)	80–84	5	3–4
Triple screen	69	5	2
Quadruple screening	80–82	5	3
Serum integrated test	85–88	4.9	5
Fully integrated test	94–96	5	5
Stepwise sequential	92	5.1	5
Contingent sequential	91	4.5	5
Cell-free DNA	99	0.1	48–98 (for ages 20–45 respectively)

Table 5.

Characteristics and performance of different screening tests [29].

The high PPV of cell-free DNA screening is dependent on maternal age at delivery which means in younger women, a positive screening test result is more likely to be falsely positive regardless of the aneuploidy. For a woman in her early 20s, the PPV may be close to 50% for fetal trisomy 21, but this percentage is considerably higher in older women, which is clinically relevant while counseling before cell-free DNA screening considering the expensive nature of the test [29].

As the placenta and the fetus do not share the same chromosomal content, false positives can occur especially when there is confined placental mosaicism (CPM) and a vanishing co-twin with an identifiable fetal pole. Hence cell-free DNA screening is not recommended in such conditions [36, 37]. Moreover, as this screening examines the maternal DNA, rare cases of maternal mosaicism and malignancy have also been identified [38, 39] by the presence of more than one aneuploidies in the test. Another disadvantage of this screening method is the ‘No Call’ result which is seen in 4–5% of screened pregnancies. This is due to a reduced fetal fraction of less than 4% which is seen in lower gestational age, obese women, small placentation, and aneuploidies [40, 41, 42]. If a no-call result is reported the patient should be counseled by a geneticist in detail regarding the possible cause and is offered a repeat test or invasive amniocentesis keeping in mind the high chance of no-call in the repeat test which is as high as 40%.

Because of its high detection and low false positive rate, cell-free DNA screening may be offered as either a primary screen or secondary screening test to women who test positive on a traditional screening test before proceeding with a diagnostic test. If an abnormal traditional screening result is followed by a normal cell-free DNA screen, the risk for a chromosomal abnormality is approximately 2% [43]. However, the time required for cell-free DNA screening (7–10 days) may delay aneuploidy diagnosis to the point that pregnancy termination may no longer be an option for those who choose it. Because of the above-mentioned limitations and the reduced cost-effectiveness in low-risk pregnancies, traditional screening tests are still considered the choice of first-line screening for low-risk pregnancies [8]. However, cell-free DNA screening is recommended as a screening option in advanced maternal age (maternal age > 35 years at delivery), high/intermediate-risk in traditional screening, presence of an ultrasonographic soft marker, prior pregnancy with h/o trisomy, or known carrier of a balanced Robertsonian translocation involving chromosomes 21, 13, and 14 [9]. Currently, cell-free DNA screening detects specific chromosomal abnormalities, namely trisomy 21, 18, and 13; 45, X; and 47 XXX, XXY, and XYY [44]. It should be noted that prenatal diagnosis is recommended whenever an aneuploidy screening test is abnormal and pregnancy termination should not be based on the results of any screening test. A comparison of traditional and cell-free DNA screening is given in Table 6. Cell-free DNA is not offered if the first trimester scan reveals any structural abnormalities.

Traditional screening	Cell-free DNA screening
First-line screening in low-risk pregnancies	Can be considered as first-line in high-risk pregnancies; A secondary screening if the traditional screen is positive
Accurate GA is essential	Done any time after 10 weeks GA
85–94% sensitivity	99% sensitivity
5% FPR	0.1% FPR
Soft markers are used to modify the risk	Soft markers are not used to modify the risk
Screen positive results may include other chromosomal abnormalities not detected with cell-free DNA	When used as a secondary screening test, does not diagnose other aneuploidies other than T21, 18, 13, 45X, 47 XXX, XXY, and XYY

Table 6.

Comparison of traditional and cell-free DNA screening [29].

Soft marker	Description	Likelihood ratio
Ventriculomegaly	Lateral ventricular atrial measurement >10 mm	3.81
Thickened nuchal fold	Distance between the outer edge of the occipital bone to outer skin in transcerebellar diameter if >6 mm	3.79
Unossified nasal bone	Unossified nasal bone in the profile view of the fetal face	6.58
ARSA	The right subclavian artery arises directly from the aortic arch instead of originating from the brachiocephalic artery	3.94
Echogenic bowel	Fetal small bowel as echogenic as bone	1.65
ECF LV	Echogenic tissue in one or both ventricles of the heart seen on a standard 4 chamber view	0.95
Pelviectasis	Renal pelvis measuring >4 mm	1.08
Short femur	Measurement <5th percentile for gestational age	0.61
Short humerus	Measurement <5th percentile for gestational age	0.78

Table 7.

Various second trimester soft markers and their likelihood ratios.

3.1.2.1 Role of USG in the second trimester (to rule out anomalies and evaluate for soft markers)

We have already elaborately learned about the role of USG in the first trimester. Targeted imaging of fetus for anomalies (TIFFA) is a level 2 USG done at 18–24 weeks depending on the local protocols. There is a role for targeted scan after a positive aneuploidy testing as the presence of an abnormality or multiple soft markers increases the risk of aneuploidy by 50–60% [45]. It is also noted that 25–30% of fetuses with Down syndrome and almost all fetuses with T18/13 will have major abnormalities [46, 47]. Soft markers are normal USG variants with no/trivial clinical sequelae, are transient and resolve with advancing gestation or after birth, and are noted in 10% of euploid pregnancies. The most commonly noted second trimester soft markers are unossified/hypoplastic NB, ventriculomegaly, increased nuchal skinfold thickness, aberrant right subclavian artery, echogenic intracardiac focus, echogenic bowel, pelviectasis, and short femur or humerus length. When a marker has been identified, the posttest odds for trisomy 21 are derived by multiplying the pretest odds (obtained by first/second trimester screening) by the positive LR for each detected marker. The images of various second trimester soft markers are depicted in Figure 9a–h. Metaanalysis by Agathakleous et al. in 2013 suggested that when the targeted anomaly scan reveals no abnormalities and soft markers then the aneuploidy risk is reduced by 7.7-fold. Table 7 shows the various markers and their likelihood ratios [48].

Figure 9.

Commonly noted second trimester USG soft markers. (a) Profile view of the fetal face showing unossified nasal bone (UNB), (b) transcerebellar plane showing increased nuchal fold thickness (NFT), (c) fetal transventricular plane showing ventriculomegaly, (d) axial section of fetal thorax showing aberrant right subclavian artery (ARSA), (e) axial section of the fetal abdomen showing bilateral renal pelviectasis, (f) axial section of fetal thorax showing echogenic cardiac focus in the left ventricle, (g) sagittal view of abdomen showing fetal bowel as echogenic as surrounding bone, (h) short femur length corresponding to 17 weeks in a fetus at 19 weeks of gestation.

3.1.2.2 Diagnostic tests of aneuploidy

Diagnostic testing allows patients to know with certainty whether the pregnancy is affected by a particular genetic condition. Abnormal screening tests in the first or second trimester must be followed up by diagnostic tests before any final decisions are made. Commonly performed diagnostic tests include chorionic villus sampling (Figure 10), and amniocentesis (Figure 11). Preimplantation genetic diagnosis is considered in known cases of familial syndromes or previously affected children with parents being carriers. Rapid aneuploidy testing using either quantitative fluorescent polymerase chain reaction (qfPCR) or fluorescent in-situ hybridization (FISH), will detect the major trisomies (13, 18, and 21) and Turners syndrome (45XO) and the results are issued in 1–3 working days. Full karyotyping is then performed following culturing of the cells. This takes 10–14 days and involves microscopic examination of cells and can detect other chromosomal rearrangements. However, as this approach will not detect very small submicroscopic changes, known as copy number variations (CNVs), chromosomal microarray (CMA) has replaced conventional karyotyping in identifying the CNVs.

Figure 10.

Transabdominal chorionic villus sampling.

Figure 11.

Amniocentesis.

The advantage of prenatal diagnosis is that when an anomaly or a genetic disease is diagnosed prenatally, it helps the obstetrician and neonatologist to counsel the family, discuss the available options, and to initiate a neonatal management plan even before delivery of the fetus. In certain cases, treatment may be instituted in utero. Although diagnostic testing is recommended to be available to all women, regardless of maternal age, patients should be counseled regarding types of invasive procedures, including the expected benefits, risks, and technical aspects of the test.

The indications of diagnostic testing are as follows:

1. Positive screening test for common trisomies.

2. Previous pregnancy complicated by fetal trisomy.

3. At least one major or two minor fetal structural anomalies in the current pregnancy abnormalities.

4. A desire to have the most reliable information about the fetal karyotype.

5. A desire to have a comprehensive genetic analysis that will detect both autosomal and sex chromosome aneuploidy and pathological copy number variants.

3.1.2.3 Chorionic villus sampling (CVS)

CVS is the only diagnostic test available in the first trimester and allows for diagnostic analyses, including quantitative Fluorescent Polymerase Chain Reaction (qFPCR), karyotype, microarray, molecular testing, and gene sequencing. CVS is performed between 10 and 14 weeks of gestation. Early CVS which was performed before 9 weeks in the past is no longer recommended as it is shown to increase the risk of limb deformities and oromandibular malformations.

Under ultrasonographic guidance, a sample of placental tissue is collected for genetic evaluation through a catheter placed through either the transcervical or transabdominal route without entering the sac (Figure 10). CVS allows for earlier prenatal diagnosis and earlier pregnancy termination if desired. A disadvantage of CVS is confined placental mosaicism (CPM) which is noted in 1–2% of CVS results. Pregnancy loss attributed to CVS is approximately 1 in 450 according to recent data [49, 50, 51].

3.1.2.4 Amniocentesis

Amniocentesis is a technique by which amniotic fluid is withdrawn from the amniotic sac using a needle under continuous ultrasound guidance via a transabdominal approach to obtain a sample of fetal exfoliated cells, transudates, urine, or secretions. It can be performed from 16 weeks of pregnancy onwards (Figure 11). The various tests which can be performed in the amniotic sample are chromosomal, biochemical, molecular, and microbial studies, the most common being prenatal diagnosis of chromosomal abnormalities, single-gene disorders, and fetal infection The procedure has a risk of fetal loss of approximately 0.5% (range, 0.06–1%) [50, 51].

3.1.2.5 Preimplantation genetic diagnosis

Preimplantation genetic diagnosis (PGD) is a test to detect the abnormality before embryo transfer so that only unaffected embryos are transferred to the patient. This helps in the earlier detection of chromosomal and genetic abnormalities. After in vitro fertilization (IVF) a polar body or a single cell from the blastocyst is removed and examined for aneuploidies/genetic disorders (Figure 12). However, it is recommended that all pregnancies conceived with IVF/PGD should be offered confirmatory testing with CVS or amniocentesis as false negative reports are possible [52, 53].

Figure 12.

Preimplantation genetic diagnosis.

3.2 Preeclampsia/SGA screening

Prediction of PE and SGA can be done in the first trimester by a combination of maternal demographic characteristics, uterine artery pulsatility index (Ut art PI), mean arterial pressure (MAP), and maternal serum biochemical markers serum PAPP-A and PlGF [54].

SGA is defined as birth weight below the 10th centile for the gestational age though there are cutoffs varying between the 3rd and 10th centile. The prevalence of SGA is estimated to be 8–11%. The SGA babies are prone to develop complications like prematurity, neonatal asphyxia, hypothermia, hypoglycemia, hyperbilirubinemia, hypocalcemia, polycythemia, sepsis, and death [54]; and long-term morbidities like learning difficulties, cognitive, and behavioral defects.

Preeclampsia (PE) is a multisystem disorder of pregnancy [55, 56] and develops in 2–5% of pregnant women and is one of the leading causes of maternal and perinatal morbidity and mortality [57, 58]. The International Society for the Study of Hypertension in Pregnancy (ISSHP) definition is the accepted one by international bodies, [59] which defines gestational hypertension as systolic blood pressure (SBP) at ≥140 mm Hg and/or diastolic blood pressure (DBP) at ≥90 mm Hg on at least two occasions measured 4 h apart developing after 20 weeks of gestation in previously normotensive women. PE is defined as gestational hypertension accompanied by ≥1 of the following conditions at or after 20 weeks of gestation: (a) Proteinuria (≥30 mg/mol protein: creatinine ratio; ≥300 mg/24 h; or ≥ 2 + dipstick) (b) Maternal organ dysfunction, including acute kidney injury (creatinine ≥90 μmol/L; 1 mg/dL) liver involvement (elevated transaminases, e.g. alanine aminotransferase or aspartate aminotransferase >40 IU/L) with or without right upper quadrant or epigastric abdominal pain, neurological complications (e.g. eclampsia, altered mental status, blindness, stroke, clonus, severe headaches, and persistent visual scotomata) or hematological complications (thrombocytopenia—platelet count <150,000/μL) or c) uteroplacental dysfunction (fetal growth restriction, abnormal umbilical artery Doppler waveform analysis, or stillbirth).

PE can be further subclassified into [59]:

1. Early-onset PE (with delivery <34 weeks GA)

2. Preterm PE (with delivery <37 weeks GA)

3. Late-onset PE (with delivery ≥34 weeks GA)

4. Term PE (with delivery ≥37 weeks GA)

Most common maternal complications include placental abruption, HELLP syndrome, acute pulmonary edema, respiratory distress syndrome, acute renal failure, intracranial hemorrhage, and death [60, 61]. The early perinatal complications are fetal growth restriction, nonreassuring FHR during labor, oligohydramnios, intrauterine fetal death (IUFD) preterm birth, low Apgar scores, need for NICU admission, and long-term complications are cerebral palsy, hearing loss, visual impairment, insulin resistance, diabetes mellitus, coronary artery disease, and hypertension.

Thus, the occurrence of PE and SGA contributes significantly to adverse pregnancy outcomes. Hence screening at 11–14 weeks GA, is of paramount importance as one can identify the patients prone to develop these disorders, prevent them to a considerable extent by starting on prophylactic Aspirin and be prepared to tackle the maternal and perinatal morbidity associated with it.

Maternal risk factors for PE and SGA prediction are nulliparity, age ≥ 40 years, BMI ≥35 kg/ m², family history of PE, interpregnancy interval > 10 years, hypertensive disease in a previous pregnancy, chronic hypertension, chronic renal disease, diabetes mellitus, or autoimmune disease [62]. Based on the history and presence of risk factors, the detection rate is only 39% for preterm PE and 34% for term PE at a 10.3% false positive rate. Thus, though history-based screening is useful in identifying at-risk women in clinical practice, it is not a sufficient tool for the effective prediction of PE.

Combined risk assessment for both early PE and preterm SGA is based on maternal characteristics, assessment of biophysical markers like MAP, uterine artery pulsatility index (UTPI), and biochemical markers, namely placental growth factor (PLGF) and PAPPA.

3.2.1 Measurement of mean arterial pressure (MAP)

MAP should be measured by validated automated and semiautomated devices. Women should be seated, with their arms well supported at the level of their heart and an appropriate-sized cuff should be used according to the mid-arm circumference (small, medium, or large). After resting for 5 min, blood pressure is recorded in both arms simultaneously and two sets of similar recordings are made at 1-minute intervals (Figure 13). The four sets of SBP and DBP measurements are included in the risk calculator and the final average MAP measurement is used for the calculation of patient-specific risk. The formula for the calculation of MAP is DBP + (SBP − DBP)/3 [63].

Figure 13.

Measurement of mean arterial pressure. Courtesy: Perkin Elmer life and analytical sciences (Wileyonline library.com).

3.2.2 Measurement of Uterine artery pulsatility Index (UTPI)

UTPI is measured along with an NT scan when the fetal CRL is between 45 and 84 mm and the GA between 11 and 13 + 6 weeks according to the criteria put forward by Fetal Medicine Foundation. For this measurement, a sagittal section of the uterus is obtained identifying the cervical canal and internal cervical os by transabdominal USG. Keeping the transducer in the midline and gentle tilting to both sides will identify each uterine artery in color flow mapping alongside the cervix at the level of the internal os. Pulsed-wave Doppler is then applied with the sampling gate at 2 mm to cover the whole vessel and the angle of insonation should be less than 30° (Figure 14). When three to five consecutive waveforms are obtained, the UTPI is measured and the mean UTPI of the left and right arteries is calculated [64, 65]. The first trimester abnormal UTPI is defined as greater than the 90th percentile, achieving a detection rate of 48%, at an 8% false positive rate, for the identification of early-onset PE. However, the detection rate for predicting late-onset PE reduces to 26% at a 7% false positive rate [64].

Figure 14.

Identification of the uterine artery at the level of the internal os and the demonstration of typical waveforms of the uterine artery Doppler in the first trimester of pregnancy.

3.2.3 PLGF and PAPPA

PLGF and PAPPA are glycoproteins secreted by trophoblastic cells and changes in their levels have been implicated in the development of PE [66, 67]. Women who are prone to develop PE have significantly lower maternal PLGF and PAPPA concentrations in the first trimester than those with normal pregnancies. These biomarkers alone have a detection rate of 55% and 33%, respectively, at a 10% false positive rate, for the identification of both early and late-onset PE [68, 69, 70, 71]. However, the detection rate with the combined approach is 90% for early PE, 75% for preterm PE, and 45% for term PE with a false positive rate of 10%. The detection rate of preterm SGA is 55% and term SGA is 44% with a false positive rate of 10%.

The ASPRE trial concluded that administration of low-dose aspirin, resulted in a 62% reduction in the incidence of preterm PE, when compared to placebo but did not have a significant reduction in the incidence of term PE [72]. However, it is pertinent to note that this combined first trimester screening of PE is less effective at predicting and preventing preeclampsia developing >37 weeks of gestation and hence the need for second trimester screening methods for PE.

3.2.4 Second trimester PE prediction

This is based on the concept that uteroplacental dysfunction occurs due to an imbalance in angiogenic and antiangiogenic factors. Circulating levels of the antiangiogenic protein, soluble fms-like tyrosine kinase-1 (sFlt-1) is increased, proangiogenic factor, PlGF is decreased and the sFlt-1/PlGF ratio is elevated before the onset of PE. Therefore, measurement of angiogenic markers, either alone or combined as part of the sFlt-1/PlGF ratio, has a significant value in preeclampsia prediction [73, 74, 75].

The prospective PROGNOSIS study [76], aimed to investigate the value of using the sFlt-1/PlGF ratio to predict the absence of PE within 1 week and to predict the presence of PE within 4 weeks in women with clinical suspicion of PE. sFlt-1/PlGF ratio cutoff of ≤38 was shown to have an NPV of 99.3% for ruling out development of PE within 1 week and a ratio > 38 demonstrated a PPV of 36.7% for ruling in preeclampsia within 4 weeks in a cohort of 700 women. The PPV for the occurrence of a combined endpoint of preeclampsia/eclampsia/HELLP syndrome, maternal and/or fetal adverse outcomes within 4 weeks was 65.5% [76]. Similar results were obtained in a separate study involving Asian women [77]. Thus, sFlt-1 and PlGF can be valuable biomarkers for the short-term prediction and detection of evolving preeclampsia in women with clinical signs and symptoms of the disorder, demonstrating a high NPV for ruling out preeclampsia, although the PPV remains relatively low. However, more research is needed to elucidate the benefits of the second trimester PE screening considering perinatal and maternal risk reduction and resource optimization.

Thus, the guideline to prevent PE is following the first trimester screening and assessment for preterm PE, women identified at high risk should receive aspirin prophylaxis commencing at 11–16 weeks of gestation at a dose of 150 mg to be taken every night until either 36 weeks of gestation, when delivery occurs, or when PE is diagnosed [78].

3.3 Screening for preterm labor

Approximately 11% of infants worldwide are born preterm, and the majority of cases occur in low-income countries [79]. Preterm birth (PTB) continues to be one of the leading causes of perinatal morbidity and mortality worldwide [80, 81]. Two-thirds of PTB cases are attributed to spontaneous PTB (SPTB) and the remaining one-third are medically indicated, due to maternal or fetal complications [82]. SPTB is defined as birth between 20 and 37 weeks of gestation following the spontaneous onset of labor, preterm prelabor rupture of membranes, or premature dilation of the cervix [83].

Preterm babies require prolonged hospitalization and are at high risk of adverse outcomes, including respiratory difficulty, necrotizing enterocolitis, feeding difficulties, blindness, deafness, intraventricular hemorrhage, higher risk of death at the age of 5 years, and neurodevelopmental sequelae when compared to their term counterparts [80, 84]. Thus, they need immense and prolonged health care, and hence for both the family and society PTB constitutes a major public health problem. Considering these issues, screening and early detection of pregnancies at the highest risk for SPTB will guide us in the implementation of management options and secondary prevention of morbidities associated with SPTB.

3.3.1 Identification of maternal risk factors

Demographic risk factors like African race, low socioeconomic status, and maternal characteristics like low BMI have been identified as poor risk factors with a relative risk (RR)—<2 in identifying women who are destined to develop SPTB. Other maternal risk factors are further subclassified into prior risk factors and pregnancy-specific risk factors. The prior risk factors are previous h/o preterm birth, a short interpregnancy interval of <6 months, family h/o preterm labor, congenital uterine malformations, infections of urinary and genital tracts, maternal smoking, and drug abuse. Pregnancy-specific risk factors are mid trimester short cervix <2.5 cm and bleeding per vaginum in the first or second trimester [82, 85]. Though the greatest risk factor for SPTB is a history of the previous SPTB, prediction of SPTB beyond that is very challenging considering the heterogeneous nature of risk factors and etiology.

3.3.2 USG screening

Universal cervical length screening is controversial due to its concern about cost-effectiveness and the possibility of unnecessary interventions. The most important risk factor for SPTB is a combination of short cervical length in a woman with previous h/o SPTB, which contributes to a relative risk of 3.3 [86, 87]. Cervical assessment is done by transvaginal ultrasound measurement of cervical length which is a safe, reliable, and highly reproducible tool when performed by trained providers [88]. In a mid trimester (16–24 weeks of gestation) scan, a cervical length of 2.5 cm corresponds to the 10th centile for the period of gestation, and hence if the transvaginal cervical length is <2.5 cm, it is considered to be short [89] (Figure 15). According to the guidelines put forth by the Society for Maternal-Fetal Medicine and the American College of Obstetricians and Gynecologists, serial cervical length surveillance is indicated for pregnant women with prior h/o SPTB from 16 to 24 weeks gestation though studies have shown that 82% of women who developed SPTB did not have a short cervical length during screening by transvaginal ultrasound [90].

Figure 15.

Transvaginal cervical length measurement showing (a) normal cervix (3.8 cm) and (b) short cervix (1.6 cm).

3.3.3 Fetal fibronectin measurement

Fetal fibronectin (fFN) is an extracellular matrix glycoprotein that is present at the maternal-fetal interface of the amniotic membrane and is found in minimal quantity (<50 ng/ml) in the cervicovaginal secretions between 22 and 35 weeks of GA [91] and hence levels >50 ng/mL at >22 weeks gestation is associated with an increased risk of SPTB [92]. However, one should keep in mind that false positive test results can be noted in sexual intercourse, vaginal bleeding, and vaginal lubrication or douching [93].

A qualitative assay involves doing a swab test that detects whether fetal fibronectin is present in the cervicovaginal secretion. A positive fFN test (≥50 ng/mL) has low sensitivity and a positive predictive value [94]. However, a negative fFN test has a high negative predictive value up to 35 weeks gestation and strongly suggests that SPTB will not occur within the following 2 weeks [95]. Due to its limited predictive ability, the American College of Obstetricians and Gynecologists (ACOG) discourages the use of this test as a screening strategy in asymptomatic women, as there is a lack of evidence for better perinatal outcomes.

Quantitative assays are tests that will measure the amount of fetal fibronectin in the cervicovaginal secretions and studies have demonstrated that increasing concentration of qfFN is directly proportional to the rate of SPTB. A threshold of 10 ng/ml has high sensitivity (96%) and negative predictive value (98%) to detect those women unlikely to deliver preterm. The higher the qfFN concentration, the greater the need for surveillance and intervention. It is predicted that quantitative fetal fibronectin measurements enhance the accuracy in the identification of women at risk of preterm delivery [87]. However, studies have shown that combined fFN assay and cervical length screening had low sensitivity to predict SPTB before 35 weeks gestation which was ratified in a systematic review by Berghella et al. [91].

3.3.4 Role of Insulin-like growth factor binding protein (IGFBP-1)

Insulin-like growth factor binding protein-1 (IGFBP-1) is one of the major secretory proteins of the decidualized endometrium and is present in large amounts in the amniotic fluid. Decidua contains more phosphorylated IGFBP-1 (phIGFBP-1) and amniotic fluid contains more nonphosphorylated IGFBP-1. So, when there is a detachment of the fetal membrane, phIGFBP-1 may leak into cervical secretions and trigger the cascade of SPTB. A strong phIGFBP-1-positive result which is an immunochromatography-based dipstick test predicted delivery before 35 completed weeks with a sensitivity of 72.7%, a specificity of 83%, a PPV of 47%, and a negative predictive value of 93.6% [96]. The advantage of this test over fFN is that IGFBP-1 is less prone to influence by sexual intercourse [97].

3.3.5 Role of placental alpha microglobulin 1 (PAMG-1)

PAMG-1 is another glycoprotein synthesized by the decidua and is present in the amniotic fluid in high concentrations. There is a transudation of PAMG-1 through chorioamniotic pores in fetal membranes during uterine contractions due to the inflammatory process of labor or infection. An immunoassay bedside ‘dipstick test’ is done by a vaginal swab between 20 and 37 weeks to obtain the result within 5 min. This test has a high specificity of 97.5% and NPV of 97.5% and the advantage is that the test results will not be affected by vaginal examination, and thus can be used shortly after the vaginal examination [98].

3.3.6 Role of biomarkers

Certain pro-inflammatory cytokines, such as interleukins, tumor necrosis factor-alpha (TNF-α), C-reactive protein (CRP), granulocyte colony-stimulating factor (G-CSF), soluble intercellular adhesion molecule-1 (sICAM-1), alkaline phosphatase, stromal cell-derived factor-1a (SDF-1a), interferon-c, and matrix metalloproteinase-8 (MMP-8) are hypothesized to respond to infection at the maternal-fetal interface and stimulate the release of prostaglandins thereby causing uterine contractility and subsequent cervical change triggering SPTB. Based on this concept, an assay of these biomarkers should predict spontaneous preterm birth in women with singleton pregnancies with no symptoms of preterm labor [99]. However, multiple studies and a subsequent meta-analysis by Agudelo et al. have proven that none of the novel biomarkers are clinically useful for predicting SPTB [95, 98] and more research is needed to clarify their efficacy as predictors.

3.3.7 Preventive strategies for SPTB

1. Though OPPTIMUM trial which was designed to determine the role of progesterone in preventing SPTB concluded that progesterone supplementation did not reduce the incidence of preterm birth [100], a subsequent systematic review of randomized controlled trials has proven that vaginal progesterone supplementation starting in the mid trimester to 37 weeks gestation remains the best-known strategy to prevent SPTB in women with a history of prior PTB [101, 102, 103, 104, 105, 106, 107, 108, 109]. In addition, vaginal progesterone administration was associated with a reduction in the risk of admission to the neonatal intensive care unit (NICU), respiratory distress syndrome (RDS), composite neonatal morbidity and mortality, and birthweight <1500 g. Vaginal progesterone has been recommended for patients with a singleton gestation and a short cervix by the Society for Maternal-Fetal Medicine (SMFM), the American College of Obstetricians and Gynecologists (ACOG), the International Federation of Gynecology and Obstetrics (FIGO), and the National Institute for Health and Care Excellence (NICE) [110, 111, 112, 113].

2. Cervical cerclage, a stitch inserted into the cervix, introduced way back in 1902, is still considered one of the standard options for prophylactic intervention to prevent preterm birth and second trimester fetal loss. Revised nomenclature has been proposed by the NICE in their recent guideline based on the indication of cervical cerclage [114, 115].

1. History indicated—a cervical suture which is performed as a prophylactic measure in asymptomatic women but with a history of three or more preterm births or mid trimester losses, is usually inserted as a planned procedure at 11–14 weeks of gestation.

2. USG indicated—women with a previous history of one or more spontaneous preterm births or mid trimester losses who are undergoing ultrasound surveillance of cervical length should be offered cerclage if the cervical length is <25 mm < 24 weeks.

3. Emergency cerclage (rescue cerclage)—insertion of cerclage as a salvage measure in the case of premature cervical dilatation with exposed fetal membranes in the vagina identified by a speculum or USG can be performed up to 28 weeks.

The different types of cervical stitch are McDonald cerclage which involves placing a transvaginal purse-string suture at the cervical isthmus junction, without bladder mobilization [116]. High transvaginal or Shirodkar cerclage involves placing a transvaginal purse-string suture above the level of the cardinal ligaments following bladder mobilization, [117] and transabdominal cerclage which involves placing the suture at the cervicoisthmic junction by laparotomy or laparoscopy [118]. Transabdominal cerclage can be performed in women with previous unsuccessful transvaginal cerclage and is done in the preconception period or early pregnancy.

3.4 Screening for diabetes

Screening and prediction of diabetes in pregnancy are advisable as it causes increased morbidity, namely fetal macrosomia, trauma during birth, induction of labor, increased chance of cesarean section, shoulder dystocia, neonatal hypoglycemia, and perinatal death. Early diagnosis will ensure the patient follows medical nutritional therapy along with exercise and if glycemic control is not achieved, early recourse to oral hypoglycemic agents or Insulin can be undertaken thereby preventing the abovementioned morbidity.

The risk factors for the development of diabetes in pregnancy are:

• BMI above 30 kg/m²

• previous macrosomic baby weighing 4.5 kg or above

• previous h/o gestational diabetes

• family history of diabetes

• ethnicity with a high prevalence of diabetes

Screening is done by the 75-g 2-h oral glucose tolerance test (OGTT) to test for gestational diabetes in women with risk factors. Women who had gestational diabetes in a previous pregnancy, can either be offered early self-monitoring of blood glucose or a 75-g 2-h OGTT as soon as possible after booking (whether in the first or second trimester) and a further 75-g 2-h OGTT at 24–28 weeks if the results of the first OGTT are normal. In women with no risk factors, OGTT is offered at 24–28 weeks. Gestational diabetes mellitus (GDM) is diagnosed if the fasting plasma glucose level is 5.6 mmol/L or above or a 2-h plasma glucose level is 7.8 mmol/L or above and the woman is advised a dietician consultation and medical nutrition therapy (MNT-Diet/exercise) is initiated. If her glycemic control is inadequate within 2 weeks, she should be referred to a diabetologist for the start of oral hypoglycemic agents (OHA)/Insulin. In a woman with preexisting DM, multidisciplinary team care should be offered for optimal glycemic control and adequate end-organ assessment from preconception to delivery [119].

3.5 USG to screen for anomalies

USG has been established as an essential modality in the prenatal assessment of the fetus and thus obtain an optimal outcome for the mother and fetus. As a majority of fetal abnormalities occur in the low-risk group, targeted imaging of fetal anomalies is offered to all pregnant women. The mid trimester USG is done between 18 and 24 weeks of gestation according to the local protocol regarding the legal limit of termination of pregnancy. The sensitivity in detecting anomalies improves when done in close to 24 weeks. The request for the scan should originate from the primary obstetrician and the pregnant woman should be counseled regarding the potential benefits and limitations of a second trimester fetal ultrasound scan and a consent form should be signed before the evaluation. Mid trimester USG should be performed by trained professionals, and it includes a detailed and systematic evaluation of the external and internal anatomy of the fetus. The established accuracy in diagnosing fetal anomalies according to the EUROFETUS study is 55–60% [120].

For the mid trimester scans, a USG machine with the following capabilities should be used: real-time, grayscale transabdominal/transvaginal transducers, necessary software applications, color Doppler, power Doppler, adjustable acoustic power output controls with output display standards, freeze-frame capabilities, electronic calipers, and capacity to print/store images. High-end machines with elaborate software settings and the use of 3D/4D probes will hasten the diagnosis and reporting process in certain circumstances.

Though the safety of USG has been established in many studies [121, 122, 123], the evaluation time should be minimized, using the lowest possible power output needed to obtain diagnostic information, following the as low as reasonably achievable (ALARA) principle [124]. Apart from evaluation of cardiac activity, fetal number, fetal environment, placental appearance and location, evaluation of biometry to assess fetal growth is recommended in the mid trimester USG. Biometry includes biparietal diameter (BPD), head circumference (HC), abdominal circumference (AC), and femur length (FL). Details of evaluation and images of mandatory biometry are given in Table 8. The minimum evaluation and checklist which is recommended in the mid trimester USG are described in Table 9 [125].

Table 8.

Planes of evaluation and images of mandatory biometry in mid trimester scan.

Anatomic region	Structures assessed
Head	Intact skull, midline falx, cavum septi pellucidi, cerebral lateral ventricles, thalami, cerebellum, cisterna magna
Face	Both orbits, midsagittal facial profile, nose and mouth with an intact upper lip
Neck	Absence of cysts and masses
Spine	Sagittal, coronal, and axial views showing no open neural tube defects
Thorax	Normal shape/size of chest and lungs; Cardiac situs, Four-chamber view of heart, Aortic and pulmonary outflow tracts; No evidence of diaphragmatic hernia
Abdomen	Stomach in normal position, bowel not dilated, both kidneys present, Cord insertion site-normal
Extremities	Three segments of all four limbs with normal relationship, arms and feet present, normal muscle mass
Placenta	Position, no masses present, any accessory lobe
Umbilical cord	Three vessel cord

Table 9.

Minimum requirements recommended in 18–24 weeks USG (modified from ISUOG practice guideline for mid-trimester USG, 2011).

Evaluation of the cervix, uterine pathology like fibroids and adnexa also should be done to look for any pathology. A proper referral mechanism should be in place once a diagnosis of an anomaly is made and a detailed report including the name, date of USG, any relevant medical or obstetric conditions, the scan indication, the best estimate of gestational age, estimated delivery date, amniotic fluid assessment, BPD, HC, AC, and FL (in centiles), EFW in grams with centile graphs, Dopplers, diagnostic impression, and recommendations for follow up examination or management.

Thus, the inverted care pyramid model helps in the identification of low, intermediate, and high-risk antenatal mothers. The low-risk mothers continue their antenatal care in the general obstetrician’s clinic, the high-risk group is treated by a multidisciplinary team including perinatologists, genetic counselors, dieticians, endocrinologists, and USG experts. The intermediate group in their further visits is stratified as either high or low-risk groups and managed accordingly.

4. Postnatal period

Care during this period is based on the concept that the prevalence of various NCD, namely metabolic syndromes and premature cardiovascular diseases is increased in women with uteroplacental dysfunction. These women are referred to specialist care, namely diabetologists, nephrologists, endocrinologists, genetic counselors, cardiologists, and nutritionists thereby preventing future occurrence of NCDs. This can be achieved using lifestyle changes, exercise, and medication. Furthermore, the women who have preexisting medical morbidities like connective tissue disorders, renal disease, and neurological disease are referred to appropriate specialist physicians for a reevaluation of their medical condition and alteration of medication if indicated.

5. Conclusion

Incorporation of the above-mentioned protocols in the prenatal screening process helps in the standardization of antenatal care from the preconception period, into the pregnancy till delivery, and through the postnatal period. This structured care will help in the substantial reduction of adverse outcomes in pregnancy thus achieving an optimal perinatal outcome. Thus, prenatal screening helps us to predict the at-risk mother and fetus, and prevent the problem from occurring by means of prophylactic measures and timely interventions. Nevertheless, in unpreventable conditions, a multidisciplinary team-based approach is considered and relevant care is given to both mother and fetus to make the process of delivery and the postnatal period a less stressful and more pleasant one.

Acknowledgments

This chapter is dedicated to “Roshan Jethro Rollands, my son, my guardian angel.”