Clinical experience of the expanded carrier screening for recessive genetic diseases in a large cohort study in Southern central China

Abstract

Recessive monogenic disorders represent a significant cause of congenital malformations and disabilities in pediatric populations. The present study aims to provide the first comprehensive assessment of clinical experience with expanded carrier screening (ECS) panels in a large cohort from the Jiangxi province of Southern Central China. An ECS panel encompassing 147 genes associated with 155 genetic disorders was initially performed on 5,104 pre-gestational/prenatal females using next-generation sequencing. Following the identification of autosomal recessive conditions in female partners, sequential genetic testing was offered to 1,351 male partners, which included either the same ECS panel or other appropriate genetic tests. Comprehensive reproductive counseling was provided to all the identified at-risk couples (ARCs). Overall, 6,308 participants accepted ECS for 155 conditions (Female: 5,104, Male: 1,204) and approximately 38.43% (2,424/6,308) of them were detected as carriers for at least one of the 155 genetic conditions. The top four prevalent conditions identified in Jiangxi Province were α-thalassemia, GJB2-associated hearing loss, Krabbe disease and Wilson’s disease. Among the participated cohort, 1,960 females were identified with AR variants and 1,351 male partners received sequential testing, at a recall rate of 68.93%. Among the tested couples, a total of 36 ARCs (36/1,357, 2.65%) were identified with the same AR (n = 27) or X-linked conditions (n = 9). Our study represents the first large-scale demonstration of the substantial feasibility of ECS in the Jiangxi population of Southern Central China. Based on our findings, we propose that incorporating genes with a carrier frequency threshold of 1/2,000 in the screening panel could serve as an optimal criterion. Our findings may contribute significantly to facilitating future clinical implementations of ECS.

Supplementary Information

The online version contains supplementary material available at 10.1038/s41598-025-99253-9.

Introduction

It has been reported that there are more than 1,300 recessive inherited diseases worldwide, with those severe ones usually leading to intrauterine or neonatal mortality as well as chronic disability, which accounts for approximately 20% of infant mortality and 10% of pediatric hospitalizations in developed countries^1,2. Cumulatively, autosomal recessive (AR) and X-linked disorders are estimated to impact approximately 1 in 300 pregnancies, indicating that nearly 1–2 in 100 couples are at risk of conceiving a child affected by a recessive condition³. Since the majority of children affected by recessive conditions are born to asymptomatic carrier parents who lack overt phenotypic manifestations, preconception and prenatal carrier screening focused on specific high-prevalence conditions has emerged as an effective strategy to significantly reduce the incidence of disability and disease^4,5.

Carrier screening was initially adopted in the 1970s as a preventive genetic testing approach aiming at identifying at-risk reproductive couples who had an increased likelihood of bearing offspring with prevalent and clinically significant genetic disease within specific ethnic groups⁶. For instance, carrier screening programs for cystic fibrosis have been widely implemented in numerous Western countries, while comprehensive β-thalassemia screening initiatives have been established in several Middle East countries. Additionally, targeted Tay-Sachs disease screening has been systematically carried out among the Ashkenazi Jewish population^7,8. However, conventional carrier screening approaches are primarily confined to the evaluation of a restricted set of highly prevalent genetic variants in ethnically-targeted gene panels. In recent years, the advent of next-generation sequencing technologies coupled with their progressively decreasing costs has facilitated the development of comprehensive expanded carrier screening (ECS) panels, enabling simultaneous genotyping of multiple Mendelian recessive genes^9,10. ECS has incorporated a comprehensive panel of recessive disease-causing genes, significantly improving carrier detection rates and representing a clinically valuable alternative to conventional carrier screening approaches. The American College of Obstetricians and Gynecologists (ACOG) has recently updated its guidelines, endorsing ECS as a universally recommended approach for preconception and prenatal genetic screening across all ethnic populations¹¹. The American College of Medical Genetics (ACMG) has established that carrier screening plays a crucial role in reproductive decision-making processes, encompassing in vitro fertilization coupled with pre-implantation genetic diagnosis, the use of donor gametes, adoption considerations, and early prenatal diagnostic testing¹². The ECS screening demonstrate considerable heterogeneity in panel dimensions and detection methodologies, leading to marked variability in clinical detection rates influenced by the spectrum of genetic conditions screened, population-specific carrier frequencies, and technical performance characteristics of the screening assays.

Although an increasing number of studies have explored some aspects of ECS, ranging from panel assessment to clinical utility, the existing evidence remains largely confined to Western populations. Despite the growing recognition of expanded carrier screening (ECS) as a valuable strategy for identifying at-risk reproductive couples, its implementation in large-scale Chinese cohorts still remains limited, with few studies systematically evaluating its clinical utility. The lack of comprehensive phenotypic data and limited public awareness regarding the importance of carrier screening have hindered its widespread adoption. Furthermore, the relatively higher cost of ECS compared to traditional ethnicity-based screening methods presents a significant barrier to its clinical application in China. To address these challenges, further large-scale ECS studies are urgently needed to improve the clinical understanding of recessive genetic disorders. Such researches will not only enhance the management of high-risk reproductive couples but also facilitate informed genetic counseling regarding reproductive options. These efforts will provide critical evidence to guide clinical practice and policy-making in China, ultimately improving reproductive outcomes and reducing the burden of genetic diseases. In this large-scale study, we implemented the first expanded carrier screening (ECS) program in Jiangxi Province, Southern Central China, targeting 147 genes associated with 155 recessive genetic disorders. Our findings not only reveal the epidemiological characteristics and prevalence patterns of these genetic conditions in the study population, but also provide evidence-based management strategies to facilitate informed reproductive decision-making. This study underscores the clinical significance and practical utility of population-based ECS programs in genetic disease prevention.

Materials and methods

Study cohort

This prospective cohort study was conducted at Jiangxi Provincial Maternal and Child Health Hospital from October 2021 to March 2024. Participants were recruited through multidisciplinary referrals, including obstetricians, perinatologists, reproductive endocrinologists, and clinical geneticists. The study cohort comprised individuals who were either planning pregnancy or in early gestation (≤ 16 weeks) seeking genetic risk assessment for their offspring as part of reproductive planning. All participants were recruited from Jiangxi Province, with comprehensive demographic and clinical data systematically collected. Exclusion criteria included: (1) consanguineous relationships between couples, (2) any documented family history of inheritable genetic disorders, and (3) blood relationships among recruited subjects. These stringent criteria were implemented to minimize potential confounding genetic factors in the study population. A total of 6,308 individuals (5,104 females and 1,204 male reproductive partners) were enrolled in ECS for 155 genetic conditions. Additionally, 147 male partners of female carriers identified through ECS for 155 genetic conditions underwent sequential targeted genetic testing in a cost-efficient approach to assess their corresponding carrier status. The mean age of female participants was 25 ± 2 years, while male participants had a mean age of 35 ± 1 years. All the enrolled subjects voluntarily completed the recommended genetic testing and provided written informed consent prior to participation. This study was approved by the Ethics Committee of Jiangxi Provincial Maternal and Child Health Hospital.

Selection of conditions included in the ECS panel

In this study, ECS was performed for 147 genes associated with 155 clinically significant disorders. The selected conditions were prioritized based on stringent criteria including high population prevalence, early disease onset, clearly defined phenotypic manifestations and substantial detrimental effects on quality of life. The ECS panel comprised a total of 147 genes associated with diverse disease systems. The predominant proportion of these genes were associated with inherited metabolic disorders (IMD, n = 76, 49.03%), followed by neurological and musculoskeletal diseases (NMD, n = 23, 14.84%). The remaining genes were classified into distinct disease categories: skin conditions (SD, n = 19, 12.26%), blood disorders (BD, n = 9, 5.81%), immune and endocrine system disorders (IED, n = 8, 5.16%), digestive and urinary system diseases (DUD, n = 8, 5.16%), respiratory and hearing system diseases (RHD, n = 7, 4.76%) and other multisystem disorders classifications (MSD, n = 12, 7.74%) (Table S1). In the ECS panel dataset, the mode of inheritance for the identified genes was predominantly autosomal recessive (AR, n = 137; 93.20%), with a smaller proportion attributed to X-linked conditions (XL, n = 10; 6.80%).

Variants reported in the ECS panel

The ECS panel was designed to cover both exon regions and flanking intronic sequences spanning 30 base pairs upstream and downstream of the coding regions across 147 target genes. Overall, 10,449 pathogenic or likely pathogenic variants of 147 genes were reported as primary findings. These variants encompassed small nucleotide variants (SNVs), small inserts or deletions (Indels) within targeted regions and specific copy number variations (CNVs) including exon deletions/duplications in the DMD gene, exon 7 deletion in the SMN1 gene, as well as large deletions --SEA, -α3.7, -α4.2, Chinese, SEA-HPFH, FIL, THAI and Taiwanese in the HBA/HBB genes. Notably, pathogenic or likely pathogenic virulent variants, including nonsense variants, frameshift variants and canonical splicing variants (± 2), were also reported as secondary findings based on the 2015 ACMG/AMP classification guidelines¹³.

Screening pattern

In this study, a sequential approach was adopted for the ECS program based on the consideration of partner testing compliance and cost-effectiveness. Specifically, male partners were recommended for further testing only if their female reproductive partners were tested positive for AR conditions. The subsequent testing of male partners involved either the standard ECS panel for 155 conditions or other targeted detection methods at lower detection cost. The clinical workflow of this study was illustrated in the Figure S1.

Pretest and post-test counselling

Prior to the testing, all individuals received professional pretest genetic counseling from professional geneticists regarding the scope, significance, limitations and residual risks associated with the genetic testing. Relevant medical personal, family and pregnancy data were meticulously documented. Variation in regions highly homologous, with pseudogenes, genomic structural variation such as translocation/inversion, chimeric mutation in germ cells, and dynamic mutation were not detected in the ECS panel. During post-test counseling sessions, participants were informed of the variants detected and their implications, cascade screening of family members and reproductive choices.

DNA extraction

Genomic DNA was extracted from peripheral blood samples collected into the EDTA anticoagulant tube using DNeasy Blood & Tissue Kits (QIAGEN). The quantity and quality of DNA in each sample were determined by a Nanodrop 1000 spectrophotometer following the manufacturer’s instructions.

Targeted regions of the ECS panel and variants validation

Sequence-enrichment DNA probes were used to capture coding exons, flanking intronic sequences spanning 30 base pairs upstream and downstream of the coding regions, part of the deep intronic regions and the promoter region of the targeted 147 genes. Based on extensive previous data from the detection system, we have established robust validation-free criteria as follows: (1) SNVs with a sequencing depth of ≥ 40× and an allelic ratio of ≥ 40%; (2) InDels with a sequencing depth of ≥ 60× and an allelic ratio of ≥ 45%. Variants meeting these predefined criteria are considered validated without further testing whereas those failing to meet the thresholds were conducted with Sanger sequencing validation. Furthermore, CNVs in DMD and SMN1 identified by NGS were validated using quantitative PCR (qPCR) or multiplex ligation-dependent probe amplification (MLPA). Additionally, eight large deletions associated with thalassemia (--SEA, -α3.7, -α4.2, Chinese, Taiwanese, SEA-HPFH, FIL, and THAI) detected by NGS were further confirmed using Gap-PCR. Massively parallel sequencing was performed on the MGISEQ-2000 platform (BGI, Shenzhen, China).

Sequential testing of the male partners

The subsequent testing of male partners involved either the standard ECS panel for 155 conditions or other targeted detection methods at lower detection cost. - If the female partner carries one or more variants in any of the 10 genes (ATP7B, GAA, GJB2, MMACHC, MMUT, OCA2, PAH, SLC26A4, SMN1, TYR), the ECS for 155 conditions or a smaller screening panel for these 10 conditions by NGS could be selected.- If the female partner carries a variant specifically in any one of the SMN1, GJB2, or SLC26A4 gene, the ECS for 155 conditions or targeted genetic testing for the specific gene can be chosen.- If the female partner is identified as a carrier of thalassemia, her spouse may opt for the ECS for 155 conditions or thalassemia gene testing.

Variant interpretation and annotation

The classification and interpretation of variants were performed in accordance with the guidelines issued by the American College of Medical Genetics (ACMG). The variants were categorized as ‘Pathogenic (P)’, ‘Likely Pathogenic (LP)’, ‘Variants of Uncertain Significance (VUS)’, ‘Likely Benign (LB)’ or ‘Benign (B). However, only the P or LP variants were reported in our ECS panel. Likely benign variants, benign variants and variants of unknown significance (VUS) were excluded from our ECS panel to mitigate potential subject anxiety. The ANNOVAR software (http://www.openbioinformatics.org/annovar/), the population database gnomAD v2.1.1 (http://gnomad.broadinstitute.org), disease-related database HGMD (https://my.qiagendigitalinsights.com/bbp/view/hgmd/pro/start.php), and variant database Clinvar (https://www.ncbi.nlm.nih.gov/clinvar) were employed for variant annotation and interpretation in this study. Furthermore, online software Polyphen-2, SIFT, PROVEN, Mutation Taster, and GeneSplicer were used for in silico analysis on pathogenicity. The sequence variants were described following the HGVS nomenclature.

Statistical analysis

The average age was expressed as mean ± standard deviation. Proportions were compared using Chi-squared test or Fisher’s exact test. Statistical analysis of the collected data from 6,308 participants who underwent ECS was primarily conducted using SPSS 23.0 software. Comprehensive descriptive statistical analysis was conducted on 6,308 participants who underwent ECS, including the total carrier rate of the targeted diseases and prevalence of pathogenic or likely pathogenic variants.

Results

Overall carrier rate in the ECS panel

Among the 6,308 participants included in the study, the cohort comprised 5,104 (80.9%, 5,104/6,308) females and 1,204 (19.1%, 1,204/6,308) males. Genetic analysis demonstrated that 2,424 individuals (38.43%, 2,424/6,308) carried at least one pathogenic or likely pathogenic variant. Among them, 1,895 individuals (1,895/6,308, 30.04%) carried only one variant while 529 participants were genotyped as carriers for more than one variant (Table 1). Genetic screening revealed 1,089 pathogenic/likely pathogenic variants across 136 genes, demonstrating diverse systemic involvement as depicted in Fig. 1. Additionally, 2,414 individuals were identified with AR conditions, 7 with X-linked conditions and 3 participants with both autosomal recessive and X-linked recessive conditions. Particularly, the detection of three females on their X-linked variants was impeded due to the suggested presence of X chromosome number anomaly by NGS, which was subsequently confirmed as 47, XXX by karyotype analysis.

Table 1.

The overall positive rates of the 155 recessive diseases in the ECS panel.

Allele counts of Conditions	Number (n)	Percentage (%)
0	3,884	61.57
1	1,895	30.04
2	449	7.12
3	71	1.13
4	8	0.13
6	1	0.01
Total	6,308	100.00

Fig. 1.

Information on the different disease systems involved in the ECS panel. IED: immune and endocrine disorders; DUD: digestive and urinary disorder; MSD: multi-system disorder; NMD: neurological and musculoskeletal disorder; SD: skin disorder; RHD: Respiratory and hearing disorder; BD: blood disorder; IMD: inherited metabolic disorder.

The most prevalent AR genes and conditions in our cohort

The most prevalent disorder identified was alpha-thalassemia, with a carrier frequency of 4.11% (259/6,308). DFNA1 ranked as the second most common disease, with a carrier frequency of 2.85% (180/6,308). Among the cohort, the ten genes most frequently detected were HBA1/HBA2, GJB2, GALC, ATP7B, SLC26A4, SLC22A5, PAH, SMN1, HBB and ABCG5. These top 25 genes accounted for approximately 66.49% of all variants observed. Furthermore, twenty-three out of the total 147 identified genes exhibited a carrier frequency exceeding 1/200 in our studied population (Table 2). Notably, one male participant was detected with EX14_30 DEL in DMD.

Table 2.

The recessive genes with a carrier rate over than 1/200 in our ECS panel.

No.	Disease	Gene	Number (n)	Carrier Frequency (%)
1	Alpha-thalassemia	HBA1/HBA2	259	4.11
2	Deafness, autosomal recessive 1 A	GJB2	180	2.85
3	Krabbe disease	GALC	165	2.60
4	Wilson’s disease	ATP7B	153	2.43
5	Deafness, autosomal recessive 4, with enlarged vestibular aqueduct	SLC26A4	136	2.24
6	Carnitine deficiency, systemic primary	SLC22A5	120	1.90
7	Phenylketonuria	PAH	118	1.87
8	Spinal muscular atrophy	SMN1	108	1.71
9	Beta-thalassemia	HBB	94	1.49
10	Albinism, oculocutaneous, type IA	TYR	77	1.22
11	Sitosterolemia 2	ABCG5	60	0.95
12	Albinism, oculocutaneous, type II	OCA2	54	0.86
13	Glutaric acidemia IIC	ETFDH	46	0.73
14	Methylmalonic aciduria and homocystinuria, cblC type	MMACHC	45	0.71
15	Hypophosphatasia, childhood	ALPL	44	0.70
16	Methylmalonic aciduria, mut (0) type	MMUT	43	0.68
17	Muscular dystrophy, limb-girdle, autosomal recessive 1	CAPN3	40	0.63
18	Hyperphenylalaninemia, BH4-deficient, A	PTS	40	0.63
19	Joubert syndrome 5	CEP290	39	0.62
20	Glycogen storage disease II	GAA	38	0.60
21	Hemophagocytic lymphohistiocytosis, familial, 2	PRF1	37	0.59
22	Cystic fibrosis	CFTR	32	0.51
23	Epidermolysis bullosa dystrophica inversa	COL7A1	32	0.51

The most prevalent variants identified in our study

A total of 1,089 variants associated with 136 targeted genes were detected, and the top ten variants were HBA1/HBA2: -α3.7, GALC: c.1901T > C (p.Leu634Ser) GJB2: c.235delC (p.Leu79Cysfs*3), HBA1/HBA2: --SEA, SMN1: EX7DEL, SLC26A4: c.919–2 A > G, SLC22A5: c.1400 C > G (p.Ser467Cys), HBB: c.316–197 C > T, ABCG5: c.1166G > A(p.Arg389His) and OCA2: c.1363 A > G (p.Arg455Gly). The detailed information involved in different systems was as listed in Fig. 1. Notably, one male individual was detected with a variant DMD EX14_30 DEL. The remaining prevalent variants were shown in Table 3. Among all these variants, the inherited metabolic disorder (IMD) accounts for approximately 45.37%, whereas the immune and endocrine disorders (IED) represent the smallest proportion of 2.20%.

Table 3.

The top twenty variants detected in the ECS panel in our study.

No.	Variant	Gene	Disease	Number (n)	Ratio (%)
1	-α3.7	HBA1/HBA2	Alpha-thalassemia	135	2.14
2	c.1901T > C (p.Leu634Ser)	GALC	Krabbe disease	132	2.09
3	c.235delC (p.Leu79Cysfs*3)	GJB2	Deafness, autosomal recessive 1 A	122	1.93
4	EX7 DEL	SMN1	Spinal muscular atrophy	103	1.71
5	--SEA	HBA1/HBA2	Alpha-thalassemia	92	1.63
6	c.919–2 A > G	SLC26A4	Deafness, autosomal recessive 4	56	0.89
7	c.1400 C > G(p.Ser467Cys)	SLC22A5	Carnitine deficiency, systemic primary	49	0.78
8	c.316–197 C > T	HBB	Beita thalassemia	33	0.62
9	c.1166G > A(p.Arg389His)	ABCG5	Sitosterolemia 2	30	0.52
10	c.1363 A > G(p.Arg455Gly)	OCA2	Albinism, oculocutaneous, type II	30	0.48
11	c.299_300delAT(p.His100Argfs*14)	GJB2	Deafness, autosomal recessive 1 A	27	0.48
12	c.2120 A > G(p.Asp707Gly)	CAPN3	Muscular dystrophy, limb-girdle, autosomal recessive 1	27	0.43
13	c.760 C > T(p.Arg254*)	SLC22A5	Carnitine deficiency, systemic primary	25	0.43
14	c.2333G > T(p.Arg778Leu)	ATP7B	Wilson’s disease	23	0.40
15	c.95 C > T(p.Ala32Val)	MLC1	Muscular dystrophy, limb-girdle, autosomal recessive 2	23	0.36
16	c.259 C > T(p.Pro87Ser)	PTS	Hyperphenylalaninemia, BH4-deficient, A	20	0.32
17	-α4.2	HBA1/HBA2	Alpha-thalassemia	20	0.32
18	c.2975 C > T(p.Pro992Leu)	ATP7B	Wilson’s disease	18	0.29
19	c.1838 C > G(p.Ser613*)	HPS3	Hermansky-Pudlak syndrome 3	17	0.27
20	c.2605G > A(p.Gly869Arg)	ATP7B	Wilson’s disease	16	0.25

At-Risk couples and pregnancy outcomes

In total, 1,966 female individuals were identified with genetic variants, including 1,957 females with autosomal recessive (AR) conditions, 6 with X-linked conditions, and 3 presenting both AR and X-linked conditions. Within our cohort of women positive for AR conditions (n = 1,960), a subset of 1,351 male partners (1,351/1,960, 68.93%) underwent sequential testing. Among these couples, a total of 1,204 accepted the ECS panel consisting of 155 condition tests while 147 male partners opted for alternative genetic testing options at lower costs. In the latter group, 45 individuals opted for thalassemia genetic testing, while 32 individuals chose GJB2 direct sequencing and 17 individuals selected SLC26A4 sequencing. Additionally, 45 individuals preferred a smaller ECS panel encompassing 10 conditions and 10 participants chose SMN1 detection. A total of 27 couples at risk for autosomal recessive (AR) disorders and 9 couples at risk for X-linked (XL) disorders were identified, demonstrating an ARC rate of 2.65%. The overall screening results in the cohort were shown in Fig. 2. These ARCs received preconception (n = 19) and early pregnancy genetic counseling (n = 17), respectively. Following comprehensive genetic counseling, 16 of the 17 pregnant couples (94.12%) proceeded with prenatal diagnosis (PND) at appropriate gestational weeks. However, one couple with HBB: c.316–197 C > T and ααα anti3.7 declined further PND, separately. Among the sixteen fetuses analyzed, four were identified as high-risk cases for distinct genetic disorders: Familial Hemophagocytic Lymphohistiocytosis Type 2, GJB2-associated hearing loss, X-linked severe combined immunodeficiency and DMD respectively. In all the four cases, the families opted for pregnancy termination following genetic counseling. Moreover, among the 19 pre-conception couples, 10 (52.6%) opted for PGT-M (Table S2).

Fig. 2.

The overall results of ECS and the ARCs. The number of subjects was indicated in brackets. AR: autosomal recessive; XL: X-linked.

Estimated yield of ECS panel

Our ECS Panel encompasses 155 severe recessive disorders associated with 147 genes, including 137 AR genes and 10 X-linked genes. The yield of screening 137 autosomal recessive (AR) genes in our ECS panel of 6,308 individuals without family history was estimated based on a study by Guo and Gregg’s study in 2019¹⁴ since the 10 X-linked recessive genes were not accessible due to inherent limitations in Guo’s model. The American College of Obstetricians and Gynecologists (ACOG) recommends carrier screening for genetic disorders with a prevalence rate of ≥ 1/100, whereas the consensus in China suggested screening on monogenic disorders with a carrier rate exceeding 1/200. The variability in range sizes of included conditions leads to differences in cumulative carrier rates (CCR) and at-risk couple rates (ACR). In our work, when applying screening criteria limited to autosomal recessive (AR) disorders with a carrier frequency ≥ 1/100, the CCR and ACR of AR genes was 22.42% and 1.33%, respectively. Notably, expansion of the screening panel to include 23 AR genes with carrier frequencies exceeding 1/200 resulted in a substantial increase in both the CCR (31.14%) and ACR (1.85%). When expanding the screening panel to include 113 autosomal recessive (AR) genes with a cumulative carrier rate (CCR) ≥ 1/2,000, only marginal increases in both the overall CCR (41.61%) and ACR (2.00%) were observed, alongside the identification of all ARCs. Nevertheless, extending the screening panel to incorporate genes with carrier frequencie lower than 1/2,000 yields no significant enhancement in either the CCR or ACR. The CCR and ACR for different ranges of AR conditions were calculated and summarized in Table 4, with corresponding graphical representations provided in Fig. 3 (3a and 3b). A comprehensive systematic review of existing studies on the ECS in the Chinese population was conducted, with the key findings from prior Chinese studies summarized in Table 5.

Table 4.

Yield of ECS panel performance for different gene set sizes.

Gene sets (GCR rate)	Gene Numbers (n)	Number of ARCs (n)	CCR (%)	ACR (%)
≥ 1/100	10	18	22.42	1.33
≥ 1/200	23	25	31.14	1.85
≥ 1/500	55	26	41.61	1.92
≥ 1/1000	87	26	45.91	1.92
≥ 1/2000	113	27	47.61	2.00
All the 137 AR genes	137	27	48.10	2.00

Fig. 3.

The overall clinical performance of CCR and ACR of the ECS panel in our study. Cumulative carrier rates (CCR) and at-risk couple rate (ACR) across varying numbers of screened genes. (a) The CCR and number of genes in different GCR cutoffs. (b) ACR for the varying number of selected AR genes. Genes are ranked in descending order based on gene carrier rate (GCR) among the 6,308 individuals in our cohort.

Table 5.

Previous studies on ECS and the carrier frequency in China.

PMID	Area	Genes (n)	Subjects (n)	Method	Positive rate (%)	ARCs Rate (%)
35079019	Jiangsu	10	3,830	CE	18.5%	1.57%
37031186	Anhui	15	327	CE	20.31	1.19%
33805278	Hong Kong	11	1, 321	NGS + Other	19.23%	0.83%
30275481	Multi- Center	11	20,952	NGS	27.49%	2.43%
33602879	Hong Kong	104	143	NGS	58.70%	7.31%
32573981	Shanghai	201	2,923	NGS	46.73%	2.26%
37450097	Shanghai	330	600	NGS	64.83%	3.00%
39030774	Sichuan	334	2,168	NGS	65.87%	11.76%
This study	Jiangxi	147	6,108	NGS	38.43%	2.65%

Abbreviations NGS: next generation sequencing; CE: capillary electrophoresis.

Discussion

Since its introduction in 2009, ECS has gained widespread acceptance as a robust approach for identifying reproductive risks associated with dozens to hundreds of diseases owing to its versatile range of panel sizes and assay technologies^14,15. The number and the types of genetic conditions included in ECS panel has been a subject of extensive discussion. Numerous professional societies have issued recommendations and suggestions on careful vetting of the clinical and population characteristics of each disease rather than indiscriminately encompassing a wide range of conditions. The selection of genes included in the ECS panel may exhibit population-specific variations, potentially leading to differences in both detection rates and ARC rate across diverse populations. To date, the majority of expert opinions on ECS have been based on research data from European countries. Current epidemiological data indicate that the prevalence of birth defects in China reaches approximately 5.6%, with monogenic disorders contributing to nearly 30% of these cases^16,17. So far, the majority of researches in population carrier screening have been focused on newborns^18,19. Therefore, the implementation of ECS in China remains inconsistent across regions, and there is a paucity of robust epidemiological data regarding the prevalence of genetic disorders in the population. In addition, the consensus on whether ECS should be universally or selectively applied in China still remains elusive to a certain extent. In order to optimize the clinical benefits of ECS, it is imperative to conduct further large-scale investigations into carrier frequencies for recessive genetic disorders in China. Although several previous studies on ECS have demonstrated a high carrier rate of monogenic disease in certain areas of China, these investigations remain limited by either the scope of screened conditions or their sample sizes^20–23.

In our work, an extensive ECS Panel comprising 137 AR and 10 X-linked conditions was performed in a large cohort of 6,308 individuals, representing the largest sample size investigation on ECS in China. It was observed that 38.43% of individuals carried at least one of the 1,088 variants in 136 identified genes. The carrier frequency observed in our study was higher than that reported in a previous investigation (24% carrier rate for at least one variant)²³. The observed discrepancy in the prevalence estimation between our ECS findings and the study conducted by Zhao may be primarily attributable to our more comprehensive panel design, which encompasses a wider array of screened disorders. Our findings closely align with a previous retrospective study on the prevalence of 135 recessive diseases in the Chinese population (46.73% carried at least one variant)²². The spectrum of monogenic variants in Jiangxi area demonstrates notable similarities with those observed in other Chinese populations, including Shanghai, Anhui, and Southern China. For instance, DFNB1A (GJB2), Krabbe disease (GALC), EVA (SLC26A4) and Wilson disease (ATP7B) exhibited a high prevalence across multiple regions in China, suggesting their high prevalence throughout the country. Notably, consistent with previous studies, thalassemia demonstrates a high carrier rate of 5.60% in Jiangxi Province, ranking as the most prevalent monogenic disorder in this region. The prevalence of Carnitine deficiency, systemic primary (SLC22A5), is also frequent in the Jiangxi area, similar to its neighboring area- Hunan Province²⁴. This observation may be explained by the interregional population migration between these two regions. Given the extensive geographical expanse and ethnically diverse populations in China, the genetic architecture underlying recessive disorders demonstrates significant heterogeneity across this country. Therefore, the standardized disease screening panel utilized in our work might necessitate region-specific modifications to achieve optimal diagnostic efficacy across various Chinese populations.

The American College of Obstetricians and Gynecologists (ACOG) endorsed in March 2017 that the screened diseases in ECS panel should meet the carrier rate of 1/100 or greater^25,26. Furthermore, the American College of Medical Genetics and Genomics (ACMG) underscores the significance of prioritizing clinically relevant disorders over indiscriminate large-scale testing, and they recommend screening for genes with a carrier frequency exceeding 1/200 in any ethnic population as part of preconception or prenatal genetic testing protocols¹¹. Several previous ECS studies have demonstrated that employing a screening threshold for monogenic diseases with a prevalence > 1/100 may fail to detect 11–81% of the ARCs²⁷. In contrast, setting the frequency cutoff at > 1/200 could identify approximately 94% of ARCs or substantially increase the detection rate of these conditions²⁸. In order to enhance the cost-effectiveness and acceptance of expanded carrier screening (ECS) among populations, further optimization of the selected conditions included in the panel is necessary. A relatively high ARCs rate of 2.65% was identified in our work, which may be explained by the expanded gene scope in ECS. Based on our findings on carrier frequency data among reproductive-aged population in Jiangxi Province, it appears that the current 155-condition panel may not be optimally suited for this region. Our findings on the current ECS Panel revealed that only 10 genes exhibit a carrier frequency ≥ 1/100, 23 genes demonstrate a carrier frequency ≥ 1/200, 55 genes with a carrier rate ≥ 1/500, 87 genes show a carrier rate ≥ 1/1,000 while 113 genes with the carrier frequency ≥ 1/2000. Restricting the carrier screening panel to only the 10 conditions with carrier frequencies ≥ 1/100 would substantially reduce both the estimated detection rate (22.42% vs. 48.10%, a reduction of approximately 50%) and the ARCs detection rate (1.33% vs. 2.00%). In contrast, expanding the panel to include 113 genes with carrier frequencies ≥ 1/2,000 could maintain a comparable overall detection rate (47.61% vs. 48.10%), with all the ARCs being detected. Therefore, based on our findings, the screening strategy focusing on disorders with carrier frequencies ≥ 1/100 proves insufficient for detecting the majority of carriers and ARCs. We propose that incorporating genes with a carrier rate of 1/2,000 in the Panel would optimize the detection yield while maintaining a reasonable balance with potential psychosocial implications in Jiangxi Province. Through continuous and systematic investigations in the field of ECS, we aim to develop an optimized ECS platform designed to achieve enhanced detection sensitivity while maintaining cost-effectiveness for clinical applications. Among the seventeen gestational ARCs in our cohort, sixteen opted for prenatal diagnosis to prevent the birth of severely affected offspring and four couples subsequently chose to terminate their pregnancies following the prenatal diagnosis of affected fetuses. Notably, only 10 out of the total 19 ARCs pursuing preconception genetic counseling have currently opted for PGT-M to prevent affected pregnancies. The remaining 9 couples exhibited significant ambivalence toward either PGT-M or natural conception, a hesitation that may primarily stem from the considerable financial burden associated with PGT-M. Future reproductive decisions among these couples may be influenced by either increased awareness regarding selective termination of pregnancies affected by monogenic disorders, or reduced costs of in vitro fertilization (IVF).

Accurate and thorough genetic counseling plays an indispensable role in the clinical management of ECS. However, uncertainty regarding the correlation between phenotype and genotype remains a challenge in clinical practice. The observed phenomena of genotype-phenotype heterogeneity and incomplete penetrance underscore the crucial role of comprehensive genetic counseling in clinical practice. Variants in DMD are associated with two forms of muscular dystrophies, including Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD)²⁹. DMD is a severe X-linked recessive neuromuscular disorder characterized by progressive muscle weakness, which affects approximately 1 in every 5,000 live male births across diverse ethnic populations³⁰. Nevertheless, BMD are characterized by relatively milder clinical manifestations, occurring at an approximately one-third the incidence rate of DMD³¹. In our work, DMD variants exhibit a carrier frequency of 0.12% and constitute approximately 16.67% of the total ARCs in Jiangxi Province, which demonstrates a similar prevalence compared to a previous study by Zhao²³. This study represents the first comprehensive investigation of Duchenne muscular dystrophy (DMD) carrier rates among reproductive-age individuals in Jiangxi Province. Notably, one male participant was found to carry the EX14_30 DEL mutation in the DMD gene. Despite this genetic finding, comprehensive clinical evaluation at our institution revealed no significant phenotypic abnormalities. Genetic analysis of the proband identified an EX49_51 deletion in the DMD gene. Pedigree investigation revealed that this variant was paternally inherited. Notably, the 61-year-old father with the exon deletion exhibits no significant clinical abnormalities. Hence, we propose pedigree analysis on the relevant family members to ascertain the origin of the variant, thereby improving the accuracy of genetic counseling for this family. Additionally, the pathogenic variant GJB2 c.109G > A (p.Val37Ile) was excluded from our analysis due to its demonstrated low penetrance in hearing impairment³². The inclusion of the GJB2 c.109G > A would significantly improved both the positive detection rate and the ARCs rate in our work, consistent with the findings reported in previous studies by Chen^20,33.

According to the recommendations of ACOG, comprehensive genetic screening should be offered to all reproductive partners of female carriers with a specific genetic condition. This approach enables accurate genetic risk assessment and counseling for couples regarding the probability of having an affected offspring. In our cohort, approximately 68.93% (n = 1,351) of male partners underwent sequential testing, representing a significantly higher proportion compared to the approximately 50% rate previously reported by Pereira et al.³⁴ and a lower rate compared to the sequential testing rate (77%) observed in the previous study by Simone et al.³⁵. We observed an actual ARCs rate at 2.65% (36/1,357) among the tested couples. However, this finding may represent an underestimation of the true carrier rate due to the substantial proportion of cases (n = 605) where male partners declined sequential testing. The primary reasons for refusal were concerns about detection costs and limited awareness of genetic disorders. Cost reduction in testing procedures and enhancement of genetic education are imperative to facilitate the effective implementation of ECS. The barriers to implementing universal sequential testing of male partners in our study were multifactorial, including challenges in providing timely post-test genetic counseling for positive cases, the substantial financial burden associated with partner testing, residual risks inherent in current disease screening methodologies, psychological stress resulting from positive test outcomes, and logistical difficulties in coordinating testing timelines between positive individuals and their partners. To ensure the comprehensive implementation of ECS, it is essential to prioritize several critical measures, such as enhancing public and provider education on genetic disorders, reducing the cost of genetic testing, providing comprehensive pre- and post-test genetic counseling, optimizing clinical management for identified carriers and establishing systematic follow-up protocols.

Conclusion

Our study showed a notably high carrier rate of recessive disorders at 38.43% in the Jiangxi population, facilitating personalized reproductive options for these detected ARCs. To date, our study represents the first and largest clinical investigation to to assess the burden of carriers with recessive genetic disorders in the Jiangxi Province of Central Southern China, thereby offering valuable insights into potential implementation of ECS programs nationwide.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Author contributions

Lu pan and Haiyan Luo contributed to the study design and manuscript writing. Tingting Huang, Huizhen Yuan, Yongyi Zou, Qing Lu and Baitao Zeng performed the ECS panel testing and data analysis. Pengpeng Ma, Laipeng Luo, Yan Yang, Ting Huang, Danping Liu and Bicheng Yang assisted in clinical management and follow-ups of these individuals with positive status. Fen Fu, Jun Zou and Yanqiu Liu provided guidance for the paper writing. All the authors have reviewed and approved the final manuscript.

Funding

This study was supported by Jiangxi Province Key Research and Development Project (Grant No. 20232BBG70023 to YZ), Jiangxi Provincial Science and Technology Plan Projects of Traditional Chinese Medicine (Grant No. 20232B1258 to HL) Jiangxi Provincial Natural Science Foundation (Grant No. 20232BAB216025 to HL).

Data availability

The data sets generated and/or analysed during the current study are available in the Figshare repository[DOI 10.6084/m9.figshare.28122389, 10.6084/m9.figshare.28122392 and 10.6084/m9.figshare.28328357].

Declarations

Competing interests

The authors declare no competing interests.

Ethics approval and consent to participate

Ethics approval has been granted by the Ethics Committee of Jiangxi Provincial Maternal and Child Health Hospital in accordance with the Helsinki Declaration. All the patients have provided written informed consent.