# Prenatal Assessment of Fetal Vertebrae and Ribs by Three-Dimensional Ultrasound and the Association with Fetal and Neonatal Outcome

Pauline Schut, Titia Cohen-Overbeek, Tom JM Van Dooren, Annelieke Hijkoop, Marjolein H.G. Dremmen, Erwin Brosens, Frietson Galis, Alex Eggink

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Pauline Schut, Titia Cohen-Overbeek, Tom JM Van Dooren, Annelieke Hijkoop, Marjolein H.G. Dremmen, et al.. Prenatal Assessment of Fetal Vertebrae and Ribs by Three-Dimensional Ultrasound and the Association with Fetal and Neonatal Outcome. Fetal Diagnosis and Therapy, 2025, pp.1-12. 10.1159/000548341. hal-05351446

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# Prenatal Assessment of Fetal Vertebrae and Ribs by Three-Dimensional Ultrasound and the Association with Fetal and Neonatal Outcome

Pauline C. Schutª Titia E. Cohen-Overbeekª Tom J.M. van Doorenᵇ,ᶜ
Annelieke Hijkoopª Marjolein H.G. Dremmenᵈ Erwin Brosenseᵉ
Frietson Galisᶜ Alex J. Egginkª

ªDivision of Obstetrics and Fetal Medicine, Department of Obstetrics and Gynecology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands; ᵇCNRS, Institute of Ecology and Environmental Sciences iEES Paris, Sorbonne University Paris, Paris, France; ᶜNaturalis Biodiversity Center, Leiden, The Netherlands; ᵈDivision of Pediatric Radiology, Department of Radiology and Nuclear Medicine, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands; ᵉDepartment of Clinical Genetics, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands

**Keywords**

Supernumerary rib · Cervical rib · Lumbar rib · Rudimentary rib · Three-dimensional ultrasound

**Abstract**

**Introduction:** The presence of an abnormal vertebral pattern and (rudimentary) cervical ribs in particular has been associated with adverse fetal and neonatal outcomes, such as intrauterine fetal death and structural or chromosomal abnormalities. The feasibility and potential added value of prenatal assessment of the vertebral pattern and the presence of cervical ribs are currently unclear. Aims of this study were to evaluate the feasibility of prenatal assessment of the fetal vertebral pattern and cervical ribs using three-dimensional ultrasound and determine whether an abnormal vertebral pattern is associated with adverse fetal and neonatal outcome. **Methods:** A total of 1,138 women referred for an advanced ultrasound examination were included, and volume data sets of the spine were acquired. The vertebral pattern was reassessed on postnatal radiographic examinations, when available. Associations between adverse outcomes and abnormalities of the vertebral pattern that had at least a good agreement between prenatal and postnatal as-

sessments were studied. **Results:** Agreement between prenatal and postnatal assessment of the presence of cervical ribs was poor, which also applied to assessment of the complete vertebral pattern. Moderate to fair agreement existed between prenatal and postnatal assessments of thoracic rib number. Prenatal and postnatal assessments of lumbar ribs had a very good agreement. Lumbar ribs were rare (10/768, 1.3%) and associated with female gender, but not with any other variable. **Conclusion:** Lumbar ribs could be assessed very well on prenatal three-dimensional ultrasound and were not associated with adverse outcomes. Prenatal and postnatal agreement between the presence of cervical ribs, number of thoracic ribs, and the vertebral pattern was insufficient.

© 2025 The Author(s).
Published by S. Karger AG, Basel

**Introduction**

The presence of an abnormal vertebral pattern has been associated with adverse fetal and neonatal outcomes, such as intrauterine fetal death and structural or chromosomal abnormalities [1-5]. Variations of the

<footnote>karger@karger.com
www.karger.com/fdt</footnote>

<footnote>© 2025 The Author(s).
Published by S. Karger AG, Basel</footnote>

<footnote>This article is licensed under the Creative Commons Attribution 4.0 International License (CC BY) (https://karger.com/Services/OpenAccessLicense). Usage, derivative works and distribution are permitted provided that proper credit is given to the author and the original publisher.</footnote>

<footnote>Correspondence to:
Pauline C. Schut, p.schut@erasmusmc.nl</footnote>
cervical vertebral pattern in particular seem to be as-
sociated with disturbed fetal development, and most
individuals with cervical ribs are not expected to reach
reproductive age due to indirect selection against as-
sociated pleiotropic effects [6]. This hypothesized indi-
rect selection against variations occurring in the cervical
vertebral region can explain the striking stability of the
presence of seven cervical vertebrae in nearly all
mammals, regardless of the length of their neck [1, 7-9].
This selection probably occurs because disturbances of
the establishment of the vertebral pattern arise at a
developmental stage where they affect other developing
organ systems easily [6]. The remarkably high rates of
abnormalities of the vertebral pattern, including cervical
ribs, in deceased fetuses and neonates with multiple
structural abnormalities and in children with an ano-
rectal malformation and/or esophageal atresia and as-
sociated abnormalities support this hypothesis [1, 10].
Prenatal assessment of the vertebral pattern using three-
dimensional ultrasound seems possible [11-14], but
assessment of the number of vertebrae and ribs during
fetal ultrasound examinations is currently not routinely
performed. Studies assessing the complete vertebral
pattern, including the presence of cervical ribs, by ul-
trasound in a large cohort of fetuses both with and
without different structural, chromosomal, or genetic
anomalies, are lacking. The feasibility and potential
added value of prenatal assessment of the vertebral
pattern and of the presence of cervical ribs therefore
remain unclear. Aims of the study were to determine
whether the number of vertebrae and ribs, including the
presence of cervical ribs, can be reliably assessed in the
prenatal period using three-dimensional ultrasound, by
comparing this prenatal assessment with postnatal ra-
diographs, magnetic resonance imaging (MRI), or
computed tomography (CT) scans, when available and
to assess whether abnormalities of the vertebral pattern
which have been reliably assessed prenatally are asso-
ciated with an adverse fetal or neonatal outcome, such as
structural, chromosomal, or genetic abnormalities or
fetal or neonatal death.

Methods

Study Participants and Design

The study was a prospective single-center study,
conducted at the Department of Obstetrics and Gyne-
cology at the Erasmus University Medical Center, Rot-
terdam, a tertiary referral center in the Netherlands.
Pregnant women who were at least 18 years old, referred

for an advanced ultrasound examination in the period
between May 2014 and February 2018, were included
after written informed consent. Indications for referral
were a suspected fetal structural abnormality or in-
creased risk of fetal structural abnormalities due to
factors such as family history, maternal diabetes, or the
use of teratogenic medication. Fetuses with and without
structural, chromosomal, or genetic abnormalities were
included.

The ultrasound examinations were performed using
Voluson E8 or E10 systems (GE Healthcare, Waukesha,
WI, USA), equipped with transabdominal 1-7 MHz
transducers. During the advanced ultrasound exami-
nation, volume data sets of the spine were acquired
according to the study protocol (online suppl. file; for all
online suppl. material, see https://doi.org/10.1159/
000548341). All sonographers received instructions
and were trained in obtaining the correct volume data
sets of the spine. The volumes were saved, and the
number of vertebrae and ribs was independently assessed
by two reviewers using 4DView (version 14 Ext. 3, GE
Healthcare Austria GmbH & Co OG), without knowl-
edge of the indication of the ultrasound examination, the
ultrasound findings, and fetal outcome.

The vertebral patterns were classified according to
Ten Broek et al. [1] (Fig. 1). This classification system
categorizes abnormalities based on the vertebral region
involved and includes changes at the cervicothoracic
boundary, the thoracolumbar boundary, the lumbosacral
boundary, or across multiple boundaries. The seventh
cervical vertebra was identified by counting downward
from the first cervical vertebra in a sagittal plane. In
4DView, the sagittal, transverse, and coronal views were
visualized simultaneously. When a bone-white structure
was visualized separate from the ossification centers of
the center or vertebral arches, it was classified as a
cervical rib (Fig. 2). If the structure was less than half of
the length of the first thoracic rib, it was considered a
rudimentary cervical rib. Ribs on the first or last thoracic
vertebra were rudimentary if their length was less than
half of the adjacent thoracic rib. In case of discrepancies
between the assessment of the number of vertebrae and/
or ribs by both reviewers, the volumes were reassessed
together and if agreement was not reached the number of
vertebrae or ribs was classified as “unknown.” To assess
the reliability of prenatal ultrasound, the number of ribs
and the vertebral pattern were studied by a single ex-
perienced radiologist (MD) on postnatal radiographs,
MRI, or CT scans, when available. The radiologist was
blinded to the outcome of the prenatal assessment of the
vertebral pattern. Agreement between prenatal and
<figure><img src="image_2.png" /><figcaption>**Fig. 1.** Overview of different vertebral patterns. From left to right: R, regular pattern; CT, shift at the cervicothoracic boundary; CT_TL, shift at the cervicothoracic and thoracolumbar boundary; CT_TL LS, shift at cervicothoracic, thoracolumbar, and lumbosacral boundary.</figcaption></figure>

<figure><img src="image_3.png" /><figcaption>**Fig. 2.** Sagittal and transverse ultrasound view of the vertebral column. In 4DView, the transverse view at the level of the seventh cervical vertebral column can be visualized simultaneously, corresponding with the yellow dot in the sagittal view. The arrows point to the bone-white structures that were classified as bilateral rudimentary cervical ribs. OC, occiput; C1, first cervical vertebra; C2, second cervical vertebra.</figcaption></figure>

postnatal assessments was calculated as percentage agreement, using Gwet's AC1 agreement coefficient [15]. This statistic is a chance-corrected agreement coefficient

which aims to circumvent undesired behavior of Cohen's kappa. The benchmarking method used takes the estimation error of the statistic into account to determine
whether agreement should be categorized as poor (AC1 <0.2), fair (0.4> AC1 ≥0.2), moderate (0.6> AC1 ≥0.4), good (0.8> AC1 ≥0.6), or very good (AC1 ≥0.8) [16]. A single (imprecise) estimate does not necessarily predict agreement well across repeated trials. Therefore, using the estimated value and the standard deviation of each AC1 statistic, we determined for each of these ordinal categories what the probability was that an estimate would fall within it. It was then rejected that agreement would be lower than a given category if the summed probabilities of all categories below it would be smaller than 0.05. Equivalently, one can then conclude that agreement was at least as good as according to that category. Thus, the category is reported for which the cumulative probability of obtaining an AC1 estimate at the categories' lowest boundary or larger is at least 0.95 and the probability of observing a lower agreement category is at most 5% [15]. We interpreted these agreements as degrees of accuracy to which vertebral traits can be determined using three-dimensional ultrasound.

Data regarding pregnancy, delivery, fetal and neonatal outcome were collected from electronic patient files or requested from other hospitals and/or midwifes. The presence of structural abnormalities was based on the following (in hierarchically descending order): autopsy reports, postnatal imaging reports, postnatal surgery, postnatal external inspection. If postnatal examinations were not performed, it was based on the findings of advanced ultrasound examinations. Structural abnormalities were categorized using the Eurocat classification [17].

Genomic single-nucleotide polymorphism (SNP) array (Illumina Infinium BeadChip 0.15 Mb) and/or karyotyping were performed on material collected in patients opting for invasive prenatal or postnatal diagnostic testing. This included amniocentesis, chorion villus biopsy, and skin biopsy. Additional DNA testing or whole exome sequencing was performed when requested by a clinical geneticist or for specific indications (e.g., cystic fibrosis in case of echogenic bowel on advanced ultrasound scan). DNA copy number alterations were determined using genotyping arrays (Illumina, San Diego, CA, USA) as described previously [18] and classified according to diagnostic guidelines released by the American College of Medical Genetics and Genomics [19].

Statistical analysis was performed using SPSS Statistics (IBM Corp., Released 2016, IBM SPSS Statistics for Windows, Version 24.0., Armonk, NY: IBM Corp.) and R [20]. Descriptive analyses were used to describe and compare the incidence of maternal, fetal, and neonatal characteristics. Chi-square or Fisher's exact tests were used to determine

incidence differences between groups. Throughout, *p* values <0.05 were considered statistically significant. One-way ANOVA *F* tests were used to compare continuous response variables between two or more groups when residuals were normally distributed. We studied associations between the occurrence of abnormalities of the vertebral pattern and other variables by means of log-linear modeling [21], using Poisson generalized linear models for counts in R. Per vertebral trait, a maximal model was fitted with all three-way associations (interactions) of the presence of that trait and two categorical explanatory variables. This model was then simplified by means of model comparisons using likelihood ratio tests, until a minimum adequate model remained which only contained significant effects. This last model per trait included at least all main effects of the trait and the explanatory variables. During model simplification, nonsignificant three-way interactions were removed first; two-way interactions were removed subsequently. We removed nonsignificant interactions involving the vertebral trait last within each group of interactions (hence those only containing explanatory variables first). We also removed interactions when the estimated effects did not allow us to interpret them, i.e., with very wide errors of the estimates, due to zero occurrences in small samples with specific combinations of variables. Explanatory variables for which there were many missing observations were not included in the maximal model, but tested with respect to the minimum adequate model retained. We tested for associations with the number of organ systems affected by structural abnormalities using logistic regression models containing that number and the variables above as explanatory variables, and including two-way interactions between the number of organ systems affected and the other explanatory variables. Model selection was again carried out as above.

We only report the results of model selection procedures on vertebral traits for which the agreement between prenatal and postnatal assessments was at least good. When agreement was lower than that, we assumed that the prenatal vertebral trait had not been determined reliably and we would therefore be modeling assessment errors confounded with the trait. Anticipating on some of the results on agreement below, we performed model selection procedures to determine associations of explanatory variables with the presence of a lumbar rib. To reduce the number of parameters to be estimated, explanatory variables were fitted with a low number of categories each. We fitted variables for maternal smoking (at intake), use of folic acid (pre- or postconceptional), assisted conception (ovulation induction, insemination, IVF, and IVF/ICSI), the occurrence of previous miscarriages, fetal gender, the presence of structural
abnormalities, birth outcome (stillbirth, termination of pregnancy, immature delivery, or live birth), and the occurrence of neonatal death. The possible association with an abnormal array or with a different karyotype (both many missing values) was studied by adding these variables to the selected models. When they had a significant effect in such a model, their association was also tested with a marginal Fisher's exact test.

## Results

Informed consent was obtained in 1,138 pregnancies. After exclusion of pregnancies without volumes of sufficient quality or follow-up, 802 pregnancies were available for analysis. Women for whom it was technically not feasible to acquire the correct volumes and those with volumes of insufficient quality had a significantly higher BMI than those with available volumes of sufficient quality of at least one vertebral region (25.69 [s.e. 0.56] and 26.20 [s.e. 0.62] versus 23.71 [s.e. 0.14], overall test $F_{2, 960} = 17.34, p < 0.01$). No statistically significant differences were seen in the presence of structural abnormalities between pregnancies with and without volumes of sufficient quality (263/801, 32.8% vs. 100/322, 31.1%; $p = 0.57$, Fisher's exact), nor in gestational age at the time of volume acquisition (median 20.57 weeks [IQR 1.14] vs. 20.43 weeks [IQR 1.29], respectively). In one fetus with trisomy 21, intrauterine fetal death occurred at 19.4 weeks' gestation. Besides acquisition of data volume sets of the spine, the advanced ultrasound examination was not completed, nor was autopsy performed. Summary statistics on the included pregnancies are provided in Table 1.

Most pregnancies ($n = 730/802, 91.0%$) resulted in live births. Indications for termination of pregnancy before 24 weeks' gestation were structural and/or chromosomal abnormalities in most cases ($n = 58/60, 96.7%$). One pregnancy was terminated because of a severe fetal growth restriction and one because of premature rupture of membranes and oligohydramnios before 24 weeks' gestation. Intrauterine fetal death occurred in nine pregnancies (1.1%). In seven of these, chromosomal or structural abnormalities were present. One intrauterine fetal death was caused by placental abruption, and one fetal death at term occurred in a growth-restricted fetus with anhydramnios. Neonatal death occurred in 11 live births, including 10 patients with severe congenital abnormalities. The gestational age at birth, cause of death, and assessment of the vertebral pattern on prenatal ultrasound examination and postnatal radiographic

images (if available) in these deceased patients are shown in Table 2.

About one-third of the total number of included fetuses and neonates had a structural abnormality. The numbers of structural abnormalities according to affected organ system are shown in Figure 3. In more than a quarter of fetuses with a structural abnormality, more than one organ system was affected (77/264, 29.2%). Cardiovascular, craniofacial, and urogenital organ systems were most frequently affected.

A SNP array was available in 206/802 (25.7%) patients and a karyotype in two patients (0.2%). Aneuploidies were diagnosed in 19 fetuses, and other chromosomal abnormalities were identified in 31 fetuses. Additional DNA analysis was requested in 42 patients (5.2%) and showed an abnormality in 13 cases (31.0%, online suppl. Table S1).

An overview of the prenatal assessment of the number of vertebrae and ribs in the total population and in fetuses with and without structural abnormalities is provided in online supplementary Table S2. Postnatal radiographs, MRI, or CT scans were available in 132 fetuses and neonates.

### Cervical Ribs

In the majority of fetuses (399/611, 65.3%), echogenic structures at the level of the 7th cervical vertebrae, which were interpreted as (rudimentary) cervical ribs, were found. An example is shown in Figure 2. Most of these were rudimentary and bilateral (361, 90.5%), followed by rudimentary left (24, 6.0%), rudimentary right (13, 3.3%), and bilateral cervical ribs (1, 0.3%). The cervical region could be assessed both prenatally and postnatally in 85 patients. Prenatal and postnatal assessments were in fair or poor agreement (cervical rib categories estimate AC1 = 0.41 [s.e. 0.07], presence of cervical ribs estimate AC1 = 0.24 [s.e. 0.11]). Only in 35 of 60 patients (58.3%) in whom cervical ribs were identified on prenatal ultrasound, the presence of cervical ribs was confirmed on postnatal radiographic images. The absence of cervical ribs was confirmed in 16 of 25 patients (64.0%) in whom cervical ribs were prenatally not identified. An example of both absence and presence of cervical ribs on prenatal and postnatal images is shown in Figure 4.

### Thoracic Ribs

The number of thoracic vertebrae and ribs could be assessed in the majority of fetuses (93.3% and 92.9%, respectively). In 131/745 (17.6%) fetuses, fewer than 24 thoracic ribs were present. However, these were rarely
<table><caption>**Table 1.** The baseline characteristics of the included pregnancies</caption><thead><tr><th></th><th>N (%)</th></tr></thead><tbody><tr><td>Pregnancies</td><td>802</td></tr><tr><td>Singleton</td><td>792 (98.8)</td></tr><tr><td>Multiple</td><td>10 (1.2)</td></tr><tr><td>Maternal age, years</td><td>30 (18–46)</td></tr><tr><td>Preconception maternal BMI</td><td>23 (15.9–43.2)</td></tr><tr><td>Maternal smoking</td><td></td></tr><tr><td>Yes</td><td>64 (8.0)</td></tr><tr><td>Quit during pregnancy</td><td>64 (8.0)</td></tr><tr><td>Unknown</td><td>1 (0.1)</td></tr><tr><td>Nullipara</td><td>378 (47.1)</td></tr><tr><td>Method of conception</td><td></td></tr><tr><td>Spontaneous</td><td>646 (80.5)</td></tr><tr><td>ICSI</td><td>94 (11.7)</td></tr><tr><td>IVF</td><td>14 (1.7)</td></tr><tr><td>Other</td><td>34 (4.2)</td></tr><tr><td>Unknown</td><td>14 (1.7)</td></tr><tr><td>Indication for advanced ultrasound scan</td><td></td></tr><tr><td>Suspected structural abnormality</td><td>394 (49.1)</td></tr><tr><td>Family history</td><td>189 (23.6)</td></tr><tr><td>ICSI</td><td>85 (10.6)</td></tr><tr><td>Maternal diabetes mellitus</td><td>30 (3.7)</td></tr><tr><td>Fetal growth restriction</td><td>20 (2.5)</td></tr><tr><td>Enlarged nuchal translucency first trimester</td><td>18 (2.2)</td></tr><tr><td>Teratogenic medication</td><td>16 (2.0)</td></tr><tr><td>Other</td><td>50 (6.2)</td></tr><tr><td>GA at time of advanced ultrasound scan, week)</td><td>20.6 (15.7–29.3)</td></tr><tr><td>Fetal and neonatal outcome</td><td></td></tr><tr><td>Termination of pregnancy</td><td>60 (7.5)</td></tr><tr><td>Immature delivery</td><td>3 (0.4)</td></tr><tr><td>Intrauterine fetal death</td><td>9 (1.1)</td></tr><tr><td>Live birth</td><td>730 (91.0)</td></tr><tr><td>Neonatal death (&lt;28 days after birth)</td><td>11 (1.5)</td></tr><tr><td>GA at time of birth, weeks<sup>a</sup></td><td>39.3 (26.1–42.4)</td></tr><tr><td>Birth weight, g<sup>a</sup></td><td>3,310 (410–5,320)</td></tr><tr><td>Structural abnormality</td><td></td></tr><tr><td>Unknown<sup>b</sup></td><td>1</td></tr></tbody></table>

Data presented as median and range or as number and percentage. BMI, body mass index, ICSI, intracytoplasmic sperm injection; IVF, in vitro fertilization; GA, gestational age. <sup>a</sup>Stillbirths, terminations of pregnancy, and immature deliveries were excluded. <sup>b</sup>In one fetus with trisomy 21, intrauterine fetal death had occurred. Data volume sets of the spine were obtained, but the advanced ultrasound scan was not completed, nor was autopsy performed.

fewer than 22 (4/745, 0.5%). In only 4/745 fetuses (0.5%), more than 24 thoracic ribs were present.

The total number of thoracic ribs was determined both prenatally and postnatally in 110 patients. The assessments of an abnormal number of thoracic ribs had a percentage agreement of 70.9%, and the AC1 statistic indicated that

the agreement was fair (AC1 = 0.50, s.e. 0.09). For the number of thoracic ribs, the agreement was moderate (AC1 = 0.64 [s.e. 0.05]). In most patients (26/37, 70.3%) in whom prenatal and postnatal assessment differed, 24 thoracic ribs were identified on prenatal ultrasound volumes, while on postnatal radiographic images, the 12th
<table><caption>**Table 2.** Gestational age at birth, age at death, and cause of death of the included neonatal deaths</caption><thead><tr><th>GA at birth, weeks</th><th>Age at death, days</th><th>Cause of death</th><th>Cervical ribs (ultrasound)</th><th>Vertebral pattern (ultrasound)</th><th>Cervical ribs (radiograph)</th><th>Vertebral pattern (radiograph)</th></tr></thead><tbody><tr><td>26.1</td><td>2</td><td>Respiratory insufficiency due to pulmonary bleeding with thrombocytopenia<br>Unsuccessful resuscitation</td><td>Unknown</td><td>Unknown</td><td>No</td><td>R</td></tr><tr><td>27.7</td><td>5</td><td>Cardiorespiratory insufficiency, bowel perforation, dysmature</td><td>No</td><td>Unknown</td><td>No</td><td>R</td></tr><tr><td>29.6</td><td>21</td><td>Cardiorespiratory insufficiency. Hypotonia and intraventricular hemorrhage</td><td>No</td><td>TL</td><td>—</td><td>—</td></tr><tr><td>31.6</td><td>2</td><td>Noonan syndrome. Hydrops, hypertrophic cardiomyopathy, persisting pulmonary hypertension. Discontinuation of treatment</td><td>Unknown</td><td>Unknown</td><td>Yes, bilateral rudimentary</td><td>CT</td></tr><tr><td>32.9</td><td>2</td><td>Multiple abnormalities, including left diaphragmatic hernia and hypoplastic left heart syndrome. Palliative care</td><td>Yes, bilateral rudimentary</td><td>CT_LS</td><td>No</td><td>LS</td></tr><tr><td>33.6</td><td>0</td><td>Cardiorespiratory insufficiency, cord prolapse, asphyxia and severe pulmonary hypoplasia, left diaphragmatic hernia</td><td>Yes, bilateral rudimentary</td><td>Unknown</td><td>Yes, bilateral rudimentary</td><td>CT_TL</td></tr><tr><td>34.1</td><td>0</td><td>Multiple abnormalities, including right diaphragmatic hernia and complex cardiac abnormality. Palliative care</td><td>Yes, bilateral rudimentary</td><td>CT</td><td>—</td><td>—</td></tr><tr><td>34.9</td><td>0</td><td>Encephalocele</td><td>Yes, bilateral rudimentary</td><td>CT</td><td>—</td><td>—</td></tr><tr><td>35.1</td><td>0</td><td>Pulmonary hypoplasia, bilateral multicystic dysplastic kidney. Palliative care</td><td>Unknown</td><td>Unknown</td><td>—</td><td>—</td></tr><tr><td>35.4</td><td>1</td><td>Trisomy 13. Multiple abnormalities, including hypoplastic left heart syndrome. Palliative care</td><td>No</td><td>Unknown (but 11 thoracic rib pairs)</td><td>—</td><td>—</td></tr><tr><td>39.3</td><td>10</td><td>Multiple organ failure caused by circulatory shock due to minimal systemic flow with complex cardiac abnormality</td><td>Unknown</td><td>Unknown</td><td>Yes, bilateral rudimentary</td><td>CT</td></tr></tbody></table>

Assessment of the presence of cervical ribs and vertebral pattern on prenatal ultrasound and radiographs after birth (if available) is also provided. R, regular pattern; CT, shift at the cervicothoracic boundary; TL, shift at the thoracolumbar boundary; LS, shift at the lumbosacral boundary; CT_TL, shift at the cervicothoracic boundary and at the thoracolumbar boundary; CT_LS, shift at the cervicothoracic boundary and at the lumbosacral boundary.

thoracic rib was classified as unilateral or bilateral rudimentary, leading to a thoracic rib count of 23 or 22, respectively. When fewer than 24 thoracic ribs were identified prenatally, this was confirmed after birth in 15/18

patients (83.3%). Postnatal images were available in two of the four patients who were prenatally suspected to have 26 thoracic ribs, and this was confirmed in 1 patient; in the other, the 26th thoracic rib was considered rudimentary.
<figure><img src="image_4.png" /><figcaption>**Fig. 3.** Distribution of the different types of structural abnormalities (n = 264).</figcaption></figure>

## Lumbar Ribs

The presence or absence of lumbar ribs could be assessed in 95.8% of fetuses. Lumbar ribs (10/768, 1.3%) were infrequent, and all were rudimentary. The percentage agreement between prenatal and postnatal assessments of the presence of lumbar ribs was 98.8% (79/80), and the agreement was very good according to the AC1 statistic (0.99, s.e. 0.01). In one of the two patients with a lumbar rib that was seen on prenatal images of whom a postnatal radiograph was available, a discrepancy was noted, namely,

a unilateral lumbar rib that was only identified on prenatal images. Lumbar ribs were associated with female gender (9/397 = 2.3% versus 1/371 = 0.2%, $\chi^2(1) = 11.45$, *p* = 0.0007) and not with any other explanatory variables. Lumbar ribs were thus not significantly more frequent in fetuses with structural abnormalities (3/246, 1.2%) compared with those without structural abnormalities (7/526, 1.3%, Fisher's exact *p* = 1.0), nor in fetuses with an abnormal array (1/47, 2.1%) compared to those with a normal array (0/144, Fisher's exact *p* = 0.25).
<figure><img src="image_5.png" /><figcaption>**Fig. 4.** Example of sagittal and axial prenatal ultrasound images analyzed using 4DView and postnatal radiographic images of a fetus without (left) and with (right) bilateral rudimentary cervical ribs (orange arrows).</figcaption></figure>

### Presacral Number of Vertebrae

A normal number of 24 presacral vertebrae was counted in most fetuses (562/605, 92.9%). The presacral number of vertebrae could be assessed on postnatal images in 21 fetuses, and the percentage agreement was 76.2% (16/21), which was moderate, based on AC1 (AC1 = 0.73 [s.e. 0.12]).

### Vertebral Pattern

The complete vertebral pattern could be assessed in 535 fetuses (66.7%). Inability to assess the vertebral pattern was caused by insufficient quality of the volumes, which could be due to fetal position, movement artifacts, high maternal BMI, or because the volumes did not include all regions of the vertebral column. Only 154 fetuses (28.8%) had a regular vertebral pattern. The most common deviation from a regular vertebral pattern was caused by shifts at the CT boundary (CT; 262, 49.0%), which was caused by the presence of echogenic foci that were interpreted as cervical ribs. An abnormal vertebral pattern including shifts at all three boundaries (CT-TL-LS) was seen in 12 (2.2%) of fetuses. The vertebral pattern was classified on both prenatal and postnatal images in 35 patients. The percentage agreement between these assessments was poor (estimate AC1 = 0.27 [s.e. 0.10]).

### Number of Affected Organ Systems and Vertebral Patterning Anomalies

We found that the presence of a lumbar rib depends in a gender-specific manner on the number of organ systems affected ($\chi^2(1) = 4.20, p = 0.040$). The parameter

estimates of the gender-specific effects were overall positive in males (95% confidence interval [-0.06, 2.8]) and not in females (c.i. [-2.2, 0.4]), entailing a correlation between the presence of a lumbar rib and the number of affected organ systems in males. This can be explained by the presence of only one male fetus with a lumbar rib that had abnormalities in three different organ systems.

## Discussion

### Main Findings

Prenatal assessment of the presence of lumbar ribs by three-dimensional ultrasound is feasible in most pregnancies. Determination of the presence of cervical ribs and determination of the complete vertebral pattern using three-dimensional ultrasound are more complex. This could not be assessed in approximately one-quarter and one-third of the included fetuses, respectively. In addition, compared to available postnatal images, prenatal assessment of the presence of cervical ribs, (rudimentary) thoracic ribs, presacral vertebrae, and the vertebral pattern could not be performed with sufficient accuracy, which makes this technique at this moment unsuitable for clinical use. The hypothesized indirect selection against variations occurring at the cervical vertebral region could therefore not be confirmed or rejected by the data from this study.

The high prevalence of echogenic structures that were interpreted as (rudimentary) cervical ribs in both fetuses
with and without structural abnormalities according to prenatal assessment on ultrasound volumes was unexpected and not in line with previous studies [1, 4, 10, 12]. An overestimated prevalence of cervical ribs could be due to the lower resolution of ultrasound, complicating accurate assessment of the cervical region, a region with a relatively high concentration of different anatomic structures. The discrepancy could also be explained by the possibility that these echogenic structures may not represent actual (rudimentary) cervical ribs. Most of these structures may have been too small to protrude beyond the transverse process of the first thoracic vertebra, a criterion that has been used to define cervical ribs in studies in which the presence of cervical ribs was assessed on radiographs [1, 10, 18]. Because the transition between vertebra and transverse process could not be precisely identified on prenatal ultrasound volumes, it was not possible to use this criterion. Instead of forming rudimentary cervical ribs, these structures may represent the pleurapophysis and fuse with the vertebral body at a later stage [22]. Alternatively, these echogenic foci could be artifacts or resolve later in fetal or neonatal development, which also applies to intra-abdominal microcalcifications or echogenic cardiac foci [23, 24]. In order to evaluate the evolution of these echogenic structures, a longitudinal follow-up study should be performed. A much better agreement of prenatal assessment lumbar ribs is probably caused by the larger size of these ribs, compared to rudimentary cervical ribs.

Overall, the incidence of fewer than 24 thoracic ribs was 11.4%, which is high in comparison with previous studies assessing fetal rib number on three-dimensional ultrasound [12, 25, 26]. This could be due to the fact that these studies only included absent ribs and did not describe the presence of rudimentary thoracic ribs. In the total study population, no association was found between an abnormal number of lumbar ribs and structural or chromosomal abnormalities, or adverse fetal or neonatal outcome. This corresponds with a previous ultrasound study [12] and also with the hypothesis that selection against abnormalities of the thoracic vertebral pattern is less strong, because this part of the vertebral column is formed later in fetal development compared to the cervical vertebral pattern [6].

In female fetuses, the presence of lumbar ribs occurred more frequently. Cervical ribs have been reported to occur more often in women [27, 28], but this has not been reported for lumbar ribs or an abnormal number of thoracic ribs [29, 30]. In many studies, the distribution of an abnormal rib number according to gender was not reported [31–33]. A potential explanation for the female

predisposition of the abnormalities in rib number in this study could be the hypothesized higher vulnerability of male embryos and fetuses to intrauterine fetal demise compared with female embryos and fetuses [34]. Abnormalities in embryonic development, such as abnormal development of the vertebral pattern, could make male embryos and fetuses more susceptible to intrauterine fetal death, compared with female embryos and fetuses, leading to a lower number of live males with abnormalities of the vertebral pattern. It remains questionable whether this so-called male disadvantage really exists, as this has been questioned by various studies [35, 36].

Because the isolated presence of lumbar ribs does not seem to have clinical implications, routine prenatal assessment of the presence of lumbar ribs using three-dimensional ultrasound does not seem indicated. For better visualization of abnormalities of the vertebral column, such as hemivertebrae, three-dimensional ultrasound can be of added value [37]. When encountered with isolated lumbar ribs, based on the data of this study and on the current literature, reassurance seems appropriate. When isolated rudimentary or absent 12th thoracic rib is visualized on prenatal ultrasound, this finding should be interpreted with caution, due to the moderate agreement with postnatal imaging. Considering these results, patients should be referred to a prenatal diagnostic center if an abnormal number of fetal ribs is suspected, for appropriate counseling by a fetal medicine specialist.

Strengths of this study were the inclusion of a large study population with and without structural abnormalities. Only a small number of cases were lost to follow-up. All volumes were assessed by two observers, who were blinded to the fetal and neonatal outcome. In addition, postnatal images were available in a relatively large number of neonates, enabling determination of agreement between prenatal and postnatal assessments of the vertebral pattern.

A limitation of this study was the inability to acquire accurate volumes in part of the included pregnancies, in particular in women with a higher BMI, while abnormalities of the vertebral column may be more prevalent in this population. Additional limitations were the lower number of postnatal radiographs in healthy neonates to confirm the prenatal findings and the lack of longitudinal follow-up in the prenatal period. Due to the period of inclusion of patients, SNP array was most frequently used, while whole exome or genome sequencing is currently more often performed, which could lead to detection of previously missed genetic abnormalities.
Future research should include longitudinal assessment of the fetal vertebrae and ribs by 3D ultrasound, especially in fetuses with echogenic foci at the seventh cervical vertebrae, and determination of the feasibility of postnatal ultrasound to confirm presence or absence of cervical ribs, which would make it possible to assess the cervical region in healthy neonates without the use of radiographs. Prenatal MRI could also be of added value in the detection of fetal cervical ribs and an abnormal rib number, as the diagnostic accuracy of MRI has been shown to be significantly higher compared to ultrasound for the detection of fetal vertebral abnormalities, such as hemivertebrae. [38] By gaining more experience with prenatal evaluation of fetal rib number using 3-dimensional ultrasound and particularly by reviewing prenatal ultrasound images with the knowledge of the final definitive diagnosis, the diagnostic accuracy of prenatal ultrasound is expected to improve. Subsequently, if ultrasound studies confirm the association between vertebral pattern abnormalities and specific chromosomal and structural abnormalities, as identified in studies involving postnatal radiographic assessments, this technique could be incorporated into the array of noninvasive prenatal diagnostic options.

### Clinical Implications

If lumbar ribs are detected on three-dimensional ultrasound as an isolated finding in the prenatal period, reassurance seems appropriate. In this study, adequate volumes of the fetal spine could not be obtained in the entire population, which hinders the use in clinical settings. As detection of lumbar ribs improves with growing expertise and advancements in ultrasound technology, this could become possible in the future.

### Conclusions

The presence of lumbar ribs could be reliably assessed on prenatal three-dimensional ultrasound volumes. Lumbar ribs are not associated with structural or

chromosomal abnormalities, nor with fetal or neonatal death. The reliability of assessment of the vertebral pattern, the presence of (rudimentary) cervical, and an abnormal number of thoracic ribs was insufficient. The prevalence of prenatal cervical ribs seemed overestimated, and correct identification on prenatal ultrasound images did not seem feasible.

### Statement of Ethics

This study protocol was reviewed and approved by the Ethics Committee of the Erasmus University Medical Center, Approval No. MEC-2014-098. Written informed consent was obtained from participants to participate in the study.

### Conflict of Interest Statement

The authors have no conflicts of interest to declare.

### Funding Sources

This study was not supported by any sponsor or funder.

### Author Contributions

P.C.S.: conceptualization, methodology, formal analysis, investigation, writing – original draft, and visualization. T.E.C.-O. and A.J. E.: conceptualization, methodology, writing – review and editing, and supervision. T.J.M.D.: formal analysis, visualization, and writing – review and editing. A.H., M.H.G.D., and E.B.: investigation and writing – review and editing. F.G.: conceptualization, methodology, and writing – review and editing.

### Data Availability Statement

All data generated or analyzed during this study are included in this article and its supplementary material files. Further inquiries can be directed to the corresponding author.

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