# Congenital abnormalities associated with microtia: A 10-YEARS retrospective study

Antoine Paul, Sophie Achard, François Simon, Nicolas Garcelon, Erea Noel

Garabedian, Vincent Couloigner, Charlotte Celerier, Françoise Denoyelle

► To cite this version:

Antoine Paul, Sophie Achard, François Simon, Nicolas Garcelon, Erea Noel Garabedian, et al. Con-genital abnormalities associated with microtia: A 10-YEARS retrospective study. International Journal of Pediatric Otorhinolaryngology, 2021, 146, pp.110764. 10.1016/j.ijporl.2021.110764. hal-04360132

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# CONGENITAL ABNORMALITIES ASSOCIATED WITH MICROTIA:
## A 10-YEARS RETROSPECTIVE STUDY

**Authors** : Antoine Paul MD¹, Sophie Achard MD¹, François Simon MD¹, Nicolas Garcelon MD², Erea Noel Garabedian PhD¹, Vincent Couloigner PhD¹, Charlotte Celerier MD¹, Françoise Denoyelle PhD¹.

### Affiliations

1. Pediatric otolaryngology and head and neck surgery department, Necker Enfants-Malades Assistance Publique - Hôpitaux de Paris, Université de Paris, Paris, France

2. Plateforme data science - institut des maladies génétiques Imagine, Inserm, centre de recherche des Cordeliers, UMR 1138 équipe 22, institut Imagine, université Sorbonne-Paris Cité, Paris, France.

Corresponding author: Antoine Paul, pediatric otolaryngology and head and neck surgery department, Necker Enfants-Malades Assistance Publique - Hôpitaux de Paris, Paris, France.

Phone number 0033(0)171396784; e-mail: antoine.paul@wuzcap.fr

All authors disclosed any actual or potential conflicts of interest, including any financial, personal or other relationships with other people or organizations within that could have inappropriately influenced this work.
1 Abstract

2 **Objective:** Microtia is a congenital auricular malformation, often part of a syndromic form
3 (35% to 55% of cases). The accurate prevalence of associated malformations remains to be
4 determined with regard to the heterogeneous results of the previous studies. This study aims
5 to describe in a large population cohort the abnormalities associated with microtia and to
6 determine the most suitable assessment for these children.

7 **Methods:** This is a retrospective and observational cohort study collecting data from the
8 medical records of children affected by microtia, diagnosed or followed-up between 2007 and
9 2017. Data were collected via a computer database. Clinical data, as well as imaging or
10 genetic results, were noted.

11 **Results:** Six hundred ninety four children were included, 587 (84.6%) with unilateral and 107 (15.4%) with bilateral microtia. Inner ear malformations were observed in 14.1% of the ears.
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13 The main associated anomalies were hemifacial microsomia (29%), velopharyngeal insufficiency (9%), ophthalmologic (6.2%), vertebral (5.9%), cardiac (5.5%) and kidney (3%) abnormalities. Main identified entities were Goldenhar, Treacher-Collins and Guion-Almeida syndromes.
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17 **Conclusion:** A comprehensive clinical assessment must be completed when microtia is
18 diagnosed. Besides screening well-known oculo-auriculo-vertebral spectrum malformations,
19 velopharyngeal insufficiency should be systematically sought. Specialized care must be
20 provided to the very frequently associated hemifacial macrosomia. Mild forms of this last
21 malformation may correspond to Guion-Almeida syndrome, especially in cases of learning
22 disability.

24 **Key-words:** Microtia, malformations, genetic, children, syndrome.
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# 1. Introduction

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Microtia is defined as a congenital auricular malformation, ranging from a small-sized pinna to a complete absence of auricular structure called anotia[1]. This abnormality is usually associated with external ear canal and middle ear malformations. Microtia is a rare malformation with an incidence rate between one and three out of 10000 births[2], which may vary according to the ethnicity of the studied population as Hispanic and Asian populations are more frequently affected.

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A significant part of cases are syndromic forms, for which prevalence rates range from 35% to 55%[3]–[5]. The most often reported abnormalities are orofacial, vertebral, ophthalmologic, or kidney malformations, and some specific syndromes are often reported such as Goldenhar, Treacher-Collins, Branchio-oto-renal or Nager syndromes. The results of previous studies regarding the frequency of the malformations associated with microtia are heterogeneous which may be explained by various differences such as population size, demographic characteristics, or study methodology. Thus, it remains difficult to estimate the accurate prevalence of the various associated anomalies. In the past few decades, a greater amount of work has been done in the genetic etiology of microtia, which will probably lead to the classification of various forms of this disorder.

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The aim of this work is to describe, in a large cohort on a ten years period, the rate of the abnormalities associated with microtia in order to help physicians in the assessment of this congenital malformation.

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# 2. Materials and methods

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This is a retrospective and observational cohort study. Data were collected from the medical records of affected children managed in the pediatric ENT department of Necker
Enfants-Malades children hospital in Paris, France. We included all children under 18 years of age with microtia-anotia diagnosed or followed-up between January 2007 and December 2017.

A standard malformative assessment was performed during this period including cardiac and renal ultrasonography, vertebral and temporal bone imaging, and ophthalmologic examination.

Data collection was performed via a computer database Dr Warehouse[6] gathering the pediatric ENT department documents as well as the results or records from the radiology, neonatology, or other specialized departments involved in the care of the affected children.

The sample selection was done in Dr Warehouse by a keyword search: "microtia," "anotia," and "pinna malformation" were used. With this method, we were able to retrieve all patients with records mentioning these keywords. Then the medical records of each patient were studied to confirm their inclusion, and from the included records, all clinical data and results of complementary exams were noted. Two pediatric otolaryngologists were responsible for the cohort selection. After they agreed about which records to include, one of them collected the data, after which another ENT physician rechecked their accuracy, trying to improve their comprehensiveness and quality.

Clinical data included sex, affected side, family history, associated facial palsy, preauricular pit or tag, and other associated congenital abnormalities. Homolateral pits or tags were not noted because they are considered as part of the pinna malformation.

Results of additional exams were also registered: temporal bone imaging results (computed tomography and/or magnetic resonance imaging) searching for associated middle or inner ear malformations and genetic results. We registered the orofacial, ophthalmologic, cardiac, vertebral, renal, neurological anomalies as well as those of the digestive tract or limbs.

Any non-isolated microtia was considered a syndromic microtia. Specific identified syndromes are noted when they were named in the medical records (ex: Goldenhar syndrome).
The relationship between qualitative variables were analyzed through Chi-square and Fisher's exact tests depending on the population size studied (more or less than 5 cases respectively). *p* < 0.05 was considered statistically significant.

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All children and parents were given oral and written comprehensive information about the study modalities, explaining that it is a retrospective study without any influence on their care, and that all data would be anonymously published. They were aware of the possibility that inclusion of their data could be canceled according to their will. This study was approved by the ethics committee CENEM (Comité d'Ethique de Necker-Enfants Malades, referring code: 2019-06-FD).

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## 3. Results

Six hundred ninety four children were included. Sex-ratio was 1.4 (284 girls, 410 boys). Cases of unilateral and bilateral microtia were registered in 587 (84.6%) and 107 (15.4%) children, respectively. Ears were separately studied when analyzing the rates of middle and inner ear malformations associated with microtia on temporal bone imaging. Thus, data of 801 ears were analyzed. In 31/587 cases of unilateral microtia (5.3%), contralateral preauricular pits or tags were noted, (27 preauricular tag, 2 preauricular pit and 2 preauricular and cheek tag). Facial palsy was diagnosed in 44 children (6.3%): 38 unilateral microtia with homolateral Facial palsy(FP), 6 cases of bilateral microtia with unilateral or bilateral FP in 3 cases for each. In this retrospective study, it was difficult to determine whether FP was congenital or acquired as it was not systematically specified in the medical records.

Temporal imaging results were available for 311 affected ears (261 children with 211 unilateral and 50 bilateral microtia). Ossicular malformations were observed in all ears: in
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281/311 ears (90.4%), ossicular dysplasia was diagnosed with all 3 ossicles identifiable,

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whereas ossicular agenesia was noted in 29/311 ears (9.3%) (table 1). Window dysplasia was

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visualized in 2.6%. Inner ear malformations were observed in 44/311 ears (14.1%), mainly

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isolated vestibular malformations (table 2a).

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Inner ear malformations were statistically more frequently observed in children having

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associated anomalies (p < 10⁻⁶). No difference was observed between unilateral and bilateral

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cases (table 2b).

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281 children were registered as having microtia associated with other abnormalities,

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the prevalence of which was as follows: hemifacial microsomia (HM) in 202/694 (29%,

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IC95%[25.8-32.6]), velopharyngeal insufficiency with and without cleft lip and/or palate in

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63/694 (9%, IC95%[7-11.5]), ophthalmologic in 43/694 (6.2%, IC95%[4.5-8.3]), vertebral in

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41/694 (5.9%, IC95%[4.3-7.9]), cardiac in 38/694 (5.5%, IC95%[3.9-7.4]), kidney in 21/694

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(3%, IC95%[1.9-4.6]), limbs in 19/694 (2.7%, IC95%[1.7-4.2]), digestive tract and neurologic

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abnormalities in 14/694 (2%, IC95%[1.1-3.4]) for both. Malformations are detailed in table 3.

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Among those 281 syndromic cases of microtia, the main associations registered were

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isolated HM in 44.8% (n=126) or HM associated with other abnormalities in 27.8% (n=78).

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The main anomalies related to HM were velopharyngeal, palate or lip (VPL) defects in 6.5%

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(n=18), vertebral malformations in 4.3% (n=12) or ophthalmologic affection in 3.6% (n=10).

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When no hemifacial microsomia was noted, the most frequently abnormalities associated with

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microtia were VPL defects in 7.5% (n=21), ophthalmological anomalies in 3.6% (n=10),

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cardiac affection in 3.2% (n=9) or vertebral malformation in 1.8% (n=5). The main

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associations are summarized in the figure 1.

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VPL, ophthalmologic, or vertebral anomalies were more frequently observed in

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children with hemifacial macrosomia (table 4a). Ophthalmological, vertebral and kidney
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anomalies were significantly associated with unilateral microtia, as compared to bilateral cases (table 4b).

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The syndromic forms most frequently identified and reported in the medical records were Goldenhar, Treacher-Collins and Guion-Almeida syndromes. Goldenhar syndrome was noted in 45% of syndromic cases. The other entities are detailed in the table 5. Overall, genetic testing was performed in 147 children (21.2%). In 12 cases, a genetic abnormality was revealed: mutation in the *EFTUD2* gene (MIM: 610536) in four cases, terminal deletion on chromosome 18 in two children, 22q11 microdeletion (MIM:192430) in two cases, EDNRA gene (MIM: 616367) deletion in one case, and three cases of trisomy (one case for each trisomy 21 (MIM: 190685) and 22, and one unspecified trisomy reported in the medical records). Genetic findings are detailed in the appendices.

## 4. Discussion

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This study examines a remarkable population of patients who have been diagnosed with microtia over a period of 10 years. Collecting these data provides a clearer picture of the various forms of microtia that have been clinically assessed with current knowledge. Moreover, all the main malformations usually noted in this context were comprehensively analyzed.

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Middle ear malformations are very frequent in microtia cases, and temporal bone imaging was systemically performed in children with hearing loss, which revealed ossicular abnormalities. Inner ear malformations were much less frequently noted than middle ear
anomalies, in accordance with the different embryological origin between both structures
(branchial arches versus the otic capsule)[7][8]. The frequency of inner ear malformations
noted in this study is similar to that of Mayer *et al* (13% versus 10.9% in our study)[9].

Diagnosing inner ear malformations should lead to pneumococcal conjugate vaccination for
the prevention of meningitis and to a vestibular assessment.

Thus, temporal bone imaging is mandatory in the assessment of microtia with hearing
loss, keeping in mind that this type of imaging is most useful when the mastoid aeration is
developed enough (from a mean age of 5 years-old, with a completely pneumatized mastoid at
about 11 years-old). However, it remains useful at an earlier age in some cases: sensorineural
hearing loss, otorrhea from a narrow canal, or fistulae located in the atretic ear, to search for
an external ear cholesteatoma.

Hemifacial microsomia was the most frequently observed malformation in this study,
29%, similar to previous work[5][10][11]. It represented in 45% of all syndromic cases. This
malformation affects the mandibular bone in most cases, but the upper and midface must be
assessed as well; Kheogh *et al*. reassessed 100 cases of microtia initially considered as
isolated: 40% of them were associated with HM such as mandibular, orbital or malar
hypoplasia[12]. We show here that most of multiple malformations cases presented with the
oculo-auriculo-vertebral (OAV) spectrum anomalies which correspond to a large and
heterogeneous range of manifestations mainly affecting aural, oral, and mandibular (or
maxillary) developments. It includes the Goldenhar syndrome defined as the association of
HM, ophthalmologic affection (mainly epibulbar cyst), and vertebral anomalies. This clinical
spectrum is related to impaired development in the first and second branchial arches involving
the neural crest cells migration[13].

Among craniofacial anomalies, we reported many cases of velopharyngeal
insufficiency, mostly without obvious morphological anomaly. Indeed, they correspond to
muscular hypoplasia or hemiparesia of the soft palate or submucosal cleft leading to velar insufficiency. Westerflier *et al*, reported 96% of soft palate dysfunction, and velopharyngeal insufficiency was observed in 60% and 85% of unilateral and bilateral microtia, respectively[14]. Similarly, Kolodzynski *et al*. recently found velar palsy in 51/83 cases of microtia[15]. Thus, a systematic analysis of velar and palate contractility or morphology should be completed in all children with microtia.

Two other malformations were highly associated with microtia, being part of the so-called OAV spectrum: ophtalmologic abnormalities (mainly coloboma 26%, and epibulbar dermoid cyst 9%). Strabism prevalence was not different to the general population: 1.5-4% according to Schuster *et al*, and Torp-Pedersen *et al*[16][17], and vertebral malformations (prevalence similar to those of Luquetti *et al* or Stoll *et al*[18][19]). Among patients with vertebral anomalies, specific syndromic forms were identified in 15/42: 10 Goldenhar, 2 Guion-Almeida, one CHARGE, one trisomy, and one Klippel-Feil syndrome.

Other malformations could be observed, but they didn't seem to be specifically linked to a well delineated syndrome. The most prevalent were as following:
- cardiopathy such as interventricular or interauricular defects, similar to the incidence in the general population.
- kidney disorders such as agenesia/hypoplasia, with one case of Goldenhar syndrome and one Fraser syndrome.

- limb malformations mainly corresponding to hypoplasia of the thumb or varus equinus of the foot: these affections were never the only malformation associated with a microtia case in our study

- digestive tract anomalies, less frequent, corresponding to esophageal atresia in a few cases.
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Among neurologic anomalies, microcephaly was the most frequent. Three out of six cases of microcephaly corresponded to Guion-Almeida syndrome. Screening for mutations of the *EFTUD2* gene[20] related to Guion-Almeida syndrome, seems interesting in cases of microtia associated with microcephaly, especially when hemifacial microsomia is associated.

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The high prevalence of microcephaly in the Hungarian study of Paput *et al* may question whether some cases of Guion-Almeida syndrome are part of them[21]. Microcephaly is one of the main differences between Guion Almeida and Treacher Collins syndromes (TC), both having many similarities such as malar hypoplasia and microtia[22]. The facial characteristics are nevertheless different on several points: in Guion Almeida syndrome, auricular malformations correspond mostly to large squared pinna, while TC cases present with auricular atresia or anotia. Downward-slanting palpebral fissures are specific to TC syndrome while Guion Almeida cases present with upward-slanting palpebral fissures.
Developmental delay is very often reported in Guion-Almeida syndrome and it was noted in all of our cases. The *EFTUD2* gene may be implicated in children without microcephaly: physicians and geneticists must be aware of this when children present with a severe delay in speech, discordant with their hearing level. Guion Almeida syndrome is probably frequently underdiagnosed.

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As a retrospective study, many limitations were expected such as incomplete data. Indeed, several imaging reports were unavailable, and some malformations could have been missed, either because of an incomplete assessment at the time of diagnosis, or because the information had not been noted in computerized medical records.

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On the other hand, the study is difficult to compare to those of previous authors. Indeed, there were many differences between the studies' design:

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- the population size, varying from 120 (Forrestier *et al*, 2005) to more than 818 cases

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(Luquetti *et al*, 2013)[18][23].

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- the range of years considered by the study, varying from 2 to 42 years (Wu *et al* 2010,

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Luquetti *et al* 2013)[5][18].

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- the inclusion criteria, leading to different population characteristics. For example, Luquetti

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*et al*. included stillborn infants which led to higher prevalence of severe malformations[18]. In

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Suutarla and Paput's studies, only syndromic forms without identified diagnoses were

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presented[10][21]. Our study was done in a French referring pediatric department and

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included all microtia forms.

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- the comprehensiveness of the results: it was surprising to observe that no hemifacial

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microsomia was noted in Forrester and Stoll's studies[19][23]. Velopharyngeal insufficiency

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is rarely reported, this disorder is probably underestimated as its evaluation is difficult in

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young children. Moreover, details are lacking in many studies: cardiac or vertebral

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malformations are not always specified in previous work.

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- the data analysis period. It is indeed difficult to compare results of the last few years with the

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earlier works as genetic knowledge has significantly improved as well as the corresponding

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phenotype description, mainly due to the recent development of "next generation sequencing."

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Guion Almeida syndrome and the *EFTUD2* gene have been well delineated since Line's and

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Gordon's works[20][24]. This syndrome has been probably underdiagnosed so far.

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## 5. Conclusion

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In conclusion, microtia is a rare malformation, the diagnosis of which must lead to a

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comprehensive clinical assessment. Beyond the well-known OAV spectrum malformations,

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some specific anomalies must be systematically considered due to their prevalence in this
242 population, such as hemifacial microsomia, velopharyngeal insufficiency, or milestone
243 development. The associated malformations may not be obvious. Thus, besides the
244 audiological and aesthetic considerations, it is essential for the associated anomalies to be
245 screened by pediatricians and otolaryngologists who are at the forefront of these children's
246 assessment.

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249 This research did not receive any specific grant from funding agencies in the public,
250 commercial, or not-for-profit sectors.

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[1] Y. Meurman, "Congenital microtia and meatal atresia: observations and aspects of treatment," *AMA Arch. Otolaryngol.*, vol. 66, no. 4, pp. 443-463, Oct. 1957.

[2] A. T. Hoyt *et al.*, "Sociodemographic and hispanic acculturation factors and isolated anotia/microtia," *Birt. Defects Res. A. Clin. Mol. Teratol.*, vol. 100, no. 11, pp. 852-862, Nov. 2014, doi: 10.1002/bdra.23282.

[3] M. A. Canfield, P. H. Langlois, L. M. Nguyen, and A. E. Scheuerle, "Epidemiologic features and clinical subgroups of anotia/microtia in Texas," *Birt. Defects Res. A. Clin. Mol. Teratol.*, vol. 85, no. 11, pp. 905-913, Nov. 2009, doi: 10.1002/bdra.20626.

[4] P. Mastroiacovo, C. Corchia, L. D. Botto, R. Lanni, G. Zampino, and D. Fusco, "Epidemiology and genetics of microtia-anotia: a registry based study on over one million births," *J. Med. Genet.*, vol. 32, no. 6, pp. 453-457, Jun. 1995.

[5] J. Wu, R. Zhang, Q. Zhang, Z. Xu, W. Chen, and D. Li, "Epidemiological analysis of microtia: a retrospective study in 345 patients in China," *Int. J. Pediatr. Otorhinolaryngol.*, vol. 74, no. 3, pp. 275-278, Mar. 2010, doi: 10.1016/j.ijporl.2009.12.001.

[6] N. Garcelon *et al.*, "A clinician friendly data warehouse oriented toward narrative reports: Dr. Warehouse," *J. Biomed. Inform.*, vol. 80, pp. 52-63, 2018, doi: 10.1016/j.jbi.2018.02.019.

[7] M. Ozeki-Satoh, A. Ishikawa, S. Yamada, C. Uwabe, and T. Takakuwa, "Morphogenesis of the middle ear ossicles and spatial relationships with the external and inner ears during the embryonic period," *Anat. Rec. Hoboken NJ 2007*, vol. 299, no. 10, pp. 1325-1337, 2016, doi: 10.1002/ar.23457.

[8] H. Thompson, A. Ohazama, P. T. Sharpe, and A. S. Tucker, "The origin of the stapes and relationship to the otic capsule and oval window," *Dev. Dyn. Off. Publ. Am. Assoc. Anat.*, vol. 241, no. 9, pp. 1396-1404, Sep. 2012, doi: 10.1002/dvdy.23831.

[9] T. E. Mayer, H. Brueckmann, R. Siegert, A. Witt, and H. Weerda, "High-resolution CT of the temporal bone in dysplasia of the auricle and external auditory canal," *AJNR Am. J. Neuroradiol.*, vol. 18, no. 1, pp. 53-65, Jan. 1997.

[10] S. Suutarla, J. Rautio, A. Ritvanen, S. Ala-Mello, J. Jero, and T. Klockars, "Microtia in Finland: comparison of characteristics in different populations," *Int. J. Pediatr. Otorhinolaryngol.*, vol. 71, no. 8, pp. 1211-1217, Aug. 2007, doi: 10.1016/j.ijporl.2007.04.020.

[11] D. P. F. van Nunen, M. N. Kolodzynski, M.-J. H. van den Boogaard, M. Kon, and C. C. Breugem, "Microtia in the Netherlands: clinical characteristics and associated anomalies," *Int. J. Pediatr. Otorhinolaryngol.*, vol. 78, no. 6, pp. 954-959, Jun. 2014, doi: 10.1016/j.ijporl.2014.03.024.

[12] I. J. Keogh, M. J. Troulis, A. A. Monroy, R. D. Eavey, and L. B. Kaban, "Isolated microtia as a marker for unsuspected hemifacial microsomia," *Arch. Otolaryngol. Head Neck Surg.*, vol. 133, no. 10, pp. 997-1001, Oct. 2007, doi: 10.1001/archotol.133.10.997.

[13] C. Tasse *et al.*, "Oculo-auriculo-vertebral spectrum (OAVS): clinical evaluation and severity scoring of 53 patients and proposal for a new classification," *Eur. J. Med. Genet.*, vol. 48, no. 4, pp. 397-411, Dec. 2005, doi: 10.1016/j.ejmg.2005.04.015.

[14] C. van Hövell Tot Westerflier, I. C. Bracamontes, Y. Tahiri, C. Breugem, and J. Reinisch, "Soft Palate Dysfunction in Children With Microtia," *J. Craniofac. Surg.*, vol. 30, no. 1, pp. 188-192, Jan. 2019, doi: 10.1097/SCS.00000000000004907.
317

318

319

[15] M. N. Kolodzynski, B. T. van Hoorn, M. Kon, and C. C. Breugem, "Abnormal soft palate movements in patients with microtia," *J. Plast. Reconstr. Aesthetic Surg. JPRAS*, vol. 71, no. 10, pp. 1476-1480, 2018, doi: 10.1016/j.bjps.2018.06.004.

320

321

322

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[16] A. K. Schuster, H. M. Elflein, R. Pokora, and M. S. Urschitz, "[Childhood strabismus in Germany: Prevalence and risk groups : Results of the KiGGS survey]," *Bundesgesundheitsblatt Gesundheitsschutz*, vol. 60, no. 8, pp. 849-855, Aug. 2017, doi: 10.1007/s00103-017-2578-x.

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325

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[17] T. Torp-Pedersen et al., "Strabismus Incidence in a Danish Population-Based Cohort of Children," *JAMA Ophthalmol.*, vol. 135, no. 10, pp. 1047-1053, Oct. 2017, doi: 10.1001/jamaophthalmol.2017.3158.

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[18] D. V. Luquetti, T. C. Cox, J. Lopez-Camelo, M. da G. Dutra, M. L. Cunningham, and E. E. Castilla, "Preferential associated anomalies in 818 cases of microtia in South America," *Am. J. Med. Genet. A.*, vol. 161A, no. 5, pp. 1051-1057, May 2013, doi: 10.1002/ajmg.a.35888.

331

332

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[19] C. Stoll, Y. Alembik, B. Dott, and M.-P. Roth, "Associated anomalies in cases with anotia and microtia," *Eur. J. Med. Genet.*, vol. 59, no. 12, pp. 607-614, Dec. 2016, doi: 10.1016/j.ejmg.2016.10.012.

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[20] M. A. Lines et al., "Haploinsufficiency of a spliceosomal GTPase encoded by EFTUD2 causes mandibulofacial dysostosis with microcephaly," *Am. J. Hum. Genet.*, vol. 90, no. 2, pp. 369-377, Feb. 2012, doi: 10.1016/j.ajhg.2011.12.023.

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[21] L. Paput, F. Bánhidy, and A. E. Czeizel, "Distribution of associated component abnormalities in cases with unclassified multiple ('syndromic') anota/microtia," *Int. J. Pediatr. Otorhinolaryngol.*, vol. 75, no. 5, pp. 639-647, May 2011, doi: 10.1016/j.ijporl.2011.01.041.

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[22] M. Vincent et al., "Treacher Collins syndrome: a clinical and molecular study based on a large series of patients," *Genet. Med. Off. J. Am. Coll. Med. Genet.*, vol. 18, no. 1, pp. 49-56, Jan. 2016, doi: 10.1038/gim.2015.29.

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[23] M. B. Forrester and R. D. Merz, "Descriptive epidemiology of anotia and microtia, Hawaii, 1986-2002," *Congenit. Anom.*, vol. 45, no. 4, pp. 119-124, Dec. 2005, doi: 10.1111/j.1741-4520.2005.00080.x.

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[24] C. T. Gordon et al., "EFTUD2 haploinsufficiency leads to syndromic oesophageal atresia," *J. Med. Genet.*, vol. 49, no. 12, pp. 737-746, Dec. 2012, doi: 10.1136/jmedgenet-2012-101173.

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359 **Appendices**

360 EFTUD2 mutations (NM_004247) : - c.1860 G>C, p.Lys629Asn

361 - c.1854del p.Thr619Profs*10

362 - c.1034del, p.Gly345ValfsX41

363 - c.2562-2_2562-1del

364 18q22 deletions : - 46,XY del 18(q22.3qter)

365 - 46,XX,del(18)(q22.2).ish del(18)(qter)

366 EDNRA mutation (NM_045958) : c.386A>T, p.Tyr129Phe

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369

370
<figure><img src="image_2.png" /><figcaption>**Figure 1: Main associations among syndromic cases of microtia**</figcaption></figure>

VLP: velopharyngeal/palate/lip abnormality

Miscellaneous: isolated cases of complex associations without hemifacial microsomia.
<table><caption>**Table 1: middle ear malformations associated with microtia.**</caption><thead><tr><th>Middle ear malformations</th><th>n (%)</th></tr></thead><tbody><tr><td><strong>Ossicular dysplasia - all ossicles identifiable</strong></td><td><strong>282 (90.7)</strong></td></tr><tr><td>including : OW agenesia</td><td>4</td></tr><tr><td>RW agenesia</td><td>1</td></tr><tr><td><strong>Ossicular agenesia</strong></td><td><strong>29 (9.3)</strong></td></tr><tr><td>Stapes / with OW dysplasia</td><td>10 / 2</td></tr><tr><td>Complete agenesia / with OW dysplasia</td><td>8 / 1</td></tr><tr><td>Malleus</td><td>4</td></tr><tr><td>Incus</td><td>3</td></tr><tr><td>Malleur and Incus</td><td>1</td></tr></tbody></table>

OW: oval window, RW: round window.
<table><caption>Table 2a: inner ear malformations associated with microtia.</caption><thead><tr><th>Inner ear malformations</th><th>n (%)</th></tr></thead><tbody><tr><td><strong>Vestibula</strong></td><td><strong>28 (9)</strong></td></tr><tr><td>Lateral SCC</td><td>15</td></tr><tr><td>Posterior SCC</td><td>5</td></tr><tr><td>Multiple SCC malformations</td><td>2</td></tr><tr><td>Vestibular dilatation</td><td>4</td></tr><tr><td>SCC + vestibule</td><td>2</td></tr><tr><td><strong>Cochlea</strong></td><td><strong>4 (1.3)</strong></td></tr><tr><td>Hypoplasia</td><td>1</td></tr><tr><td>Canal stenosis</td><td>1</td></tr><tr><td>Dysplasia</td><td>2</td></tr><tr><td><strong>CV malformations</strong></td><td><strong>6 (1.9)</strong></td></tr><tr><td>Cochlea + SCC dysplasia</td><td>2</td></tr><tr><td>Cochlea + SCC dysplasia + EVA</td><td>1</td></tr><tr><td>Unique vesicle</td><td>2</td></tr><tr><td>Cochlea canal stenosis + lateral SCC dysplasia</td><td>1</td></tr><tr><td><strong>Other</strong></td><td><strong>6 (1.9)</strong></td></tr><tr><td>SCC dysplasia + IAC dilatation</td><td>1</td></tr><tr><td>Cochlea and lateral SCC dysplasia + EVA</td><td>1</td></tr><tr><td>Cochlear nerve agenesia</td><td>1</td></tr><tr><td>CV malformation + CV nerve agenesia</td><td>1</td></tr><tr><td>Unique vesicle + cochlear nerve hypoplasia</td><td>1</td></tr><tr><td>Unknown</td><td>1</td></tr><tr><td><strong>No Inner ear malformations</strong></td><td><strong>267 (85.9)</strong></td></tr><tr><td><strong>TOTAL</strong></td><td><strong>311</strong></td></tr></tbody></table>

SCC: semicircular canal, IAC: internal auditory canal, EVA: enlarged vestibular aqueduct.

<table><caption>Table 2b: comparison of inner ear malformation rates between unilateral/bilateral groups, and between children with isolated or syndromic microtia.</caption><thead><tr><th></th><th>Inner ear malformation (n)</th></tr></thead><tbody><tr><td>Unilateral microtia (n=587)</td><td>22</td></tr><tr><td>Bilateral microtia (n=107)</td><td>6</td></tr><tr><td>p-value</td><td>0.368</td></tr><tr><td>Syndromic microtia (n=281)</td><td>24</td></tr><tr><td>Isolated microtia (n=413)</td><td>4</td></tr><tr><td>p-value</td><td>1.07 x 10<sup>-7</sup></td></tr></tbody></table>

Chi2 and Exact Fisher's tests
<table><thead><tr><th></th><th>n (%)</th></tr></thead><tbody><tr><td><strong>Hemifacial microsomia</strong></td><td><strong>202 (29)</strong></td></tr><tr><td><strong>Velopharyngeal, palate/lip abnormalities</strong></td><td><strong>63 (9)</strong></td></tr><tr><td>Velopharyngeal insufficiency - no cleft</td><td>36</td></tr><tr><td>Cleft palate</td><td>17</td></tr><tr><td>Bifida uvula</td><td>7</td></tr><tr><td>Cleft lip and palate</td><td>3</td></tr><tr><td><strong>Ophthalmologic abnormalities</strong></td><td><strong>43 (6.2)</strong></td></tr><tr><td>Coloboma</td><td>11</td></tr><tr><td>Strabism</td><td>9</td></tr><tr><td>Oculomotor palsy</td><td>4</td></tr><tr><td>Dermoid cyst</td><td>4</td></tr><tr><td>Ptosis</td><td>3</td></tr><tr><td>Hypertelorism</td><td>2</td></tr><tr><td>Miscellaneous</td><td>10</td></tr><tr><td><strong>Vertebral abnormalities</strong></td><td><strong>42 (6)</strong></td></tr><tr><td>Hemivertebra</td><td>13</td></tr><tr><td>Vertebral fusion</td><td>11</td></tr><tr><td>Scoliosis</td><td>5</td></tr><tr><td>Torticollis + cervical malformation</td><td>2</td></tr><tr><td>Narrowed cervico-occipital hinge</td><td>2</td></tr><tr><td>Miscellaneous</td><td>5</td></tr><tr><td>Unspecified</td><td>4</td></tr><tr><td><strong>Cardiac abnormalities</strong></td><td><strong>38 (5.5)</strong></td></tr><tr><td>Interventricular defect (4 with valvular insufficiency, one with left ventricular hypoplasia)</td><td>19</td></tr><tr><td>Interauricular defect (one of which with valvular insufficiency)</td><td>5</td></tr><tr><td>Interventricular + interauricular defect</td><td>3</td></tr><tr><td>Fallot tetralogy</td><td>2</td></tr><tr><td>Patent foramen ovale</td><td>2</td></tr><tr><td>Miscellaneous</td><td>5</td></tr><tr><td>Unspecified</td><td>2</td></tr><tr><td><strong>Kidney abnormalities</strong></td><td><strong>21 (3)</strong></td></tr><tr><td>Kidney hypoplasia/agenesia</td><td>6</td></tr><tr><td>Kidney ectopia</td><td>4</td></tr><tr><td>Hydronephrosis</td><td>5</td></tr><tr><td>Horseshoe kidneys</td><td>3</td></tr><tr><td>Miscellaneous</td><td>3</td></tr><tr><td><strong>Limbs abnormalities</strong></td><td><strong>19 (2.7)</strong></td></tr><tr><td>Superior limbs (4 thumb malformations, 2 clinodactyly, 2 miscellaneous)</td><td>8</td></tr><tr><td>Inferior limbs (6 varus foot, 3 asymmetry, 4 miscellaneous)</td><td>11</td></tr><tr><td><strong>Neurologic abnormalities</strong></td><td><strong>13 (1.9)</strong></td></tr><tr><td>Microcephaly</td><td>6</td></tr><tr><td>Plagiocephaly</td><td>3</td></tr><tr><td>Miscellaneous</td><td>4</td></tr><tr><td><strong>Digestive tract abnormalities</strong></td><td><strong>13 (1.9)</strong></td></tr><tr><td>Oesophageal atresia</td><td>5</td></tr><tr><td>Anorectal malformation</td><td>4</td></tr></tbody></table>
<caption>**Table 3: Malformations associated with microtia.**</caption>

Unspecified: malformation not detailed in the reports. Miscellaneous: malformations observed in isolated cases.
<table><caption>**Table 4a: comparison of the malformations between children with and without hemifacial microsomia.**</caption><thead><tr><th></th><th>Hemifacial microsomia<br>(n=202)</th><th>No hemifacial microsomia<br>(n=492)</th><th>p-value</th></tr></thead><tbody><tr><td>VLP</td><td>36</td><td>26</td><td><strong>&lt; 10<sup>-6</sup></strong></td></tr><tr><td>Ophthalmologic</td><td>23</td><td>22</td><td><strong>0.001</strong></td></tr><tr><td>Vertebral</td><td>25</td><td>13</td><td><strong>&lt; 10<sup>-6</sup></strong></td></tr><tr><td>Cardiac</td><td>15</td><td>22</td><td>0.127</td></tr><tr><td>Kidney</td><td>7</td><td>14</td><td>0.69</td></tr><tr><td>Digestive tract</td><td>9</td><td>4</td><td><strong>0.004</strong></td></tr><tr><td>Neurological</td><td>9</td><td>7</td><td><strong>0.017</strong></td></tr></tbody></table>

Chi2 and Exact Fisher's tests

VLP : velopalatal and/or lip defect

<table><caption>**Table 4b: comparison of the malformations between children with unilateral and bilateral microtia.**</caption><thead><tr><th></th><th>Unilateral microtia<br>(n=587)</th><th>Bilateral microtia<br>(n=107)</th><th>p-value</th></tr></thead><tbody><tr><td>HM</td><td>170</td><td>35</td><td>0.43</td></tr><tr><td>VLP</td><td>43</td><td>19</td><td><strong>0.0005</strong></td></tr><tr><td>Ophthalmologic</td><td>23</td><td>22</td><td><strong>&lt; 10<sup>-9</sup></strong></td></tr><tr><td>Vertebral</td><td>30</td><td>8</td><td>0.32</td></tr><tr><td>Kidney</td><td>14</td><td>7</td><td><strong>0.02</strong></td></tr><tr><td>Limb</td><td>13</td><td>6</td><td><strong>0.04</strong></td></tr><tr><td>Cardiac</td><td>26</td><td>11</td><td><strong>0.013</strong></td></tr><tr><td>Digestive tract</td><td>9</td><td>4</td><td>0.137</td></tr><tr><td>Neurological</td><td>6</td><td>10</td><td><strong>&lt; 10<sup>-6</sup></strong></td></tr></tbody></table>

Chi2 and Exact Fisher's tests

HM: hemifacial macrosomia, VLP : velopalatal and/or lip defect
<table><caption>**Table 5: genetic findings in patients referred to genetic department.**</caption><thead><tr><th></th><th>n (%)</th></tr></thead><tbody><tr><td><strong>Syndromes reported in the medical records</strong></td><td><strong>86</strong></td></tr><tr><td>Goldenhar</td><td>40 (45)</td></tr><tr><td>Treacher Collins</td><td>26 (29)</td></tr><tr><td>Guion-Almeida</td><td>4 (5)</td></tr><tr><td>Pierre Robin sequence</td><td>3 (3)</td></tr><tr><td>Branchio-oto-renal</td><td>3 (3)</td></tr><tr><td>CHARGE</td><td>2 (2)</td></tr><tr><td>Najer</td><td>2 (2)</td></tr><tr><td>Wildervanck</td><td>1 (1)</td></tr><tr><td>Klippel Feil</td><td>1 (1)</td></tr><tr><td>Pfeiffer</td><td>1 (1)</td></tr><tr><td>VACTERL</td><td>1 (1)</td></tr><tr><td>Crouzon craniostenosis</td><td>1 (1)</td></tr><tr><td>Frazer</td><td>1 (1)</td></tr><tr><td><strong>Molecular diagnosis</strong></td><td></td></tr><tr><td>EFTUD2 mutations</td><td>4 (2.7)</td></tr><tr><td>18q22 deletions</td><td>2 (1.4)</td></tr><tr><td>22q11 microdeletions</td><td>2 (1.4)</td></tr><tr><td>EDNRA mutations</td><td>1 (0.7)</td></tr><tr><td>Trisomy 22</td><td>1 (0.7)</td></tr><tr><td>Trisomy 21</td><td>1 (0.7)</td></tr><tr><td>Unspecified trisomy</td><td>1 (0.7)</td></tr></tbody></table>

Mutations are detailed in the appendices.