Carrier Screening and Prenatal Diagnosis for Spinal Muscular Atrophy in Ningde City, Fujian Province

Abstract

ABSTRACT

Objective

This study aimed to screen for SMN1 (OMIM#600354) deletion carriers in 6035 pregnant women in the eastern part of Fujian Province, to provide a theoretical basis for genetic counseling and further prenatal diagnosis of the disease, and to explore the carrier frequency and clinical significance of spinal muscular atrophy (SMA) in the eastern part of Fujian Province.

Methods

Pregnant women treated at the prenatal diagnosis institution of Ningde Municipal Hospital of Ningde Normal University from February 2022 to October 2023 were selected as research subjects. The exons 7 and 8 (E7 and E8) of the survival of motor neuron 1 gene (SMN1) from 6035 pregnant women were detected using real‐time fluorescence quantitative PCR (qPCR). Spouses of pregnant women with positive results were recalled for gene detection, and prenatal diagnosis was performed for both partners as carriers.

Results

A total of 100 SMA carriers were detected in 6035 pregnant women, including 98 with heterozygous deletions of E7 and E8 of the SMN1 gene and two with heterozygous deletions of E7 only. The carrier frequency was 1.66%. One couple was both identified as SMA carriers, and their fetus with a homozygous deletion of E7 and E8 of the SMN1 gene was finally detected by prenatal diagnosis and gene analysis.

Conclusion

The frequency of SMA mutation in the Ningde area of Fujian province has been identified, which can provide the basis for genetic counseling and prenatal diagnosis. Interventional prenatal genetic diagnosis for high‐risk fetuses can effectively prevent the birth of children with SMA and is crucial for preventing and controlling birth defects.

Introduction

Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disease caused by the degeneration and loss of alpha motor neurons in the anterior horn of the spinal cord, resulting in progressive proximal muscle atrophy, paralysis, respiratory failure caused by repeated respiratory infections, and infant death, which is the second most deadly autosomal genetic disease in children (Yeo and Darras 2020). The incidence of SMA is approximately 1 in 6000 to 1 in 10,000 live births, with an overall carrier frequency of 1/40–1/60 in the general population (D'Amico, Mercuri, and Tiziano 2011). According to the onset time and clinical indications of the disease can be divided into Types I–IV. Type I SMA (Werdnig–Hoffman disease, OMIM#253300) is the most serious; the disease occurs within 6 months after birth, and most patients die of respiratory failure within 2 years of age. Type II SMA (intermediate, OMIM#253550) is an intermediate type, and the onset of type II occurs 6–18 months after birth; most patients can only live to adulthood. Type III SMA (Kugelberg–Welander disease, OMIM#253400) is a mild form with onset in late childhood and survival into adulthood; however, the patient may gradually lose the ability to walk with age. Type IV SMA (OMIM#271150) has a mild course of onset at 35 years of age or older and is usually associated with a normal life expectancy (Russman 2007; Rudnik-Schoneborn, Forkert, and Hahnen 1996).

In 1990, Brzzustowicz et al. used polymorphism loci linkage analysis to confirm that all clinical forms of SMA map to chromosome 5q11.2 to q13.3 (Sun, Yang, and Lu 2019). In 1995, Lefebvre et al. (1995) identified the survival of motor neuron 1 (SMN1) as the disease-causing gene of SMA. The SMN1 gene encodes a functional full-length SMN protein required for motor neuron survival in the anterior horn of the spinal cord. There are two highly homologous SMN gene (Version Number: NC 000005.10) copies in the human genome: SMN1 (NC_000005.10: 70924941.0.70966375) on the telomeric copy and SMN2 (NC 000005.10: 70049523.0.70090528) on the centromeric copy (Burglen et al. 1996). Only five bases differ between the two genes at their 3' end, two of which are located in exons 7 and 8, and the other three bases are in introns 6 and 7 (Butchbach 2021). A critical single nucleotide mutation in exon 7, a C to T substitution, creates an exonic splicing silencer or disrupts an exonic splicing enhancer, leading to an SMN2 gene product with a truncated SMN protein lacking exon 7 (Kashima and Manley 2003; Cartegni and Krainer 2002). Approximately 90% of the truncated protein produced by SMN2 is unstable and rapidly degraded, which leads to various severities of clinical SMA. The more copies of SMN2, the less severe the symptoms (Mailman et al. 2002; Prior et al. 2004).

Due to the global occurrence of SMA with no significant population and regional differences, the severe clinical symptoms, the high carrier rate, and a clear genetic cause, in 2008, the American College of Medical Genetics (ACMG) suggested that regardless of ethnicity, SMN1 gene carrier screening should be carried out in all gestational age groups, and prenatal diagnosis or preimplantation diagnosis should be carried out for high-risk fetuses, to reduce the birth of SMA children (Prior TW, Professional Practice and Guidelines Committee 2008). The American College of Obstetricians and Gynecologists (ACOG) put forward advocacy in 2009, suggesting that countries and regions around the world as far as possible to control SMA epidemiological data, and aim to establish an accurate and inexpensive test for SMA and incorporate it into the pre-pregnancy screening (ACOG Committee on Genetics 2009). In 2017, ACOG recommended that all women who are considering pregnancy or are pregnant should be screened for SMA carriers (ACOG Committee on Genetics 2017).

Several strategies are used to estimate SMN1 allele frequency, and the availability and convenience of real-time quantitative polymerase chain reaction (qPCR) make it possible to screen SMA carriers as a routine strategy. In this study, qPCR was used to screen the SMN1 mutation frequency in pregnant women in eastern Fujian Province for the first time and to provide a reference for subsequent genetic counseling and prenatal diagnosis. In addition, prenatal genetic diagnosis should be carried out for high-risk fetuses to prevent the birth of children with SMA and to promote public health in eastern Fujian.

Materials and Methods

Ethical Compliance

Studies involving human participants were reviewed and approved by the Medical Ethics Committee of the Ningde Municipal Hospital of Ningde Normal University (NSYKYLL-2023-028), and all pregnant women who underwent carrier screening signed a written informed consent form.

Sample Collection

A total of 7050 pregnant women with a normal phenotype who visited the Prenatal Diagnosis Center of Ningde Municipal Hospital between January 2022 and October 2023 were selected as study participants. The pregnant women had no SMA phenotype, reproductive history, or family history. All pregnant women received SMA carrier screening education, and 6035 of them voluntarily accepted SMA carrier screening, with an acceptance rate of approximately 85.60%. The average age of pregnant women was (29.38 ± 5.08) years (range from 15 to 50 years).

Genomic DNA Extraction

We collected 2 mL peripheral blood samples from pregnant women using ethylenediamine tetraacetic acid (EDTA) anticoagulant tubes, and DNA was extracted using a DNA extraction kit (Beijing Tiangen Biochemical Technology Co. Ltd.). DNA concentration was measured using a MIULAB Ultramicroamount UV–visible spectrophotometer, the extracted DNA was assessed for purity (absorbance ratio of 260/280 nm between 1.8 and 2.0), and the DNA concentration was adjusted by dilution with ultrapure water to 10–20 ng/L. We collected 10 mL of amniotic fluid from pregnant women who underwent prenatal diagnosis, and genomic DNA was extracted from the amniotic fluid using a microDNA extraction kit (QIAGEN, Germany).

Real-Time Fluorescence Quantitative PCR

In this study, a survival of motor neuron gene 1 (SMN1) exon deletion detection kit of (Shanghai Medicore Technology Co. Ltd.) was used. The human RPP40 gene was used as an internal standard, and exons 7 and 8 of the SMN1 gene (target gene) were amplified using MGB probe multiplex real-time fluorescence quantitative PCR, and the copy number was quantified. The specific operation was performed as per the standard operating procedure; three gradients (1:2:4) normal control, heterozygous control, blank control, and 4.0 μL of specimen DNA were added to the E7 and E8 reaction systems, and the target gene (FAM channel signal) and the internal standard gene (VIC channel signal) were amplified using an ABI StepOne Plus real-time PCR instrument. The ΔCt values between target genes and internal standard genes in the E7 and E8 reactions of each gradient control were calculated, ΔCt = Ct_FAM-Ct_VIC. The average ΔCt values of the three gradients of the E7 and E8 reaction normal controls were calculated and denoted as ΔCt_a. The ΔCt values between the target gene and internal standard gene of test specimens in the E7 and E8 reactions were calculated, ΔCt = Ct_FAM-Ct_VIC, which was denoted as ΔCt_s. ∆∆Ct = ∆Ct_s-∆Ct_a. In the E7 and E8 reactions, ∆∆Ct ≤ −0.55 indicated normal, and −0.45 < ∆∆Ct ≤ 0.45 indicated a heterozygous deletion. In the E7 reaction, ∆∆Ct > 0.8 indicated a homozygous deletion of E7, and in the E8 reaction, ∆∆Ct > 1.5 indicated a homozygous deletion of E8.

Quantitative Fluorescence PCR and MLPA for Prenatal Diagnosis in High-Risk Fetuses

Prenatal diagnostic amniotic fluid and maternal peripheral blood samples were compared using quantitative fluorescence polymerase chain reaction (QF-PCR) analysis of short tandem repeat (STR) sites. After excluding maternal cell contamination, SMN1 was detected using qPCR. PCR amplification was performed per the manufacturer's instructions (Shanghai Medicore Technology Co. Ltd.) protocol. Multiplex ligation-dependent probe amplification (MLPA) was used to verify the presence of E7 and E8 copy number variations in SMN1.

Results

Results of Screen

In this study, 7050 pregnant women received SMA carrier screening education from January 2022 to October 2023, and 6035 of them voluntarily accepted SMA carrier screening, with an acceptance rate of approximately 85.60%. The ages of these women ranged from 15 to 50 years, with an average age of 29.38 ± 5.08 years (Table 1). Of the 6035 pregnant women tested (Table 2), 100 asymptomatic SMA carriers were identified using qPCR, with a carrier rate of 1.66%.

TABLE 1 Information on the 6035 pregnant women participating in the screening program.

	2022.01–2022.12	2023.01–2023.10	Total
(n = 3551)	(n = 2484)	(n = 6035)
n	%	95% CI	n	%	95% CI	n	%	95% CI
Age, years
≤ 25	417	11.74	10.72–12.84	299	12.04	10.82–13.38	716	11.86	11.07–12.70
26–34	2545	71.67	70.17–73.13	1807	72.75	70.96–74.46	4352	72.11	70.96–73.23
≥ 35	589	16.59	15.40–17.85	378	15.21	13.86–16.69	967	16.03	15.12–16.97

TABLE 2 Outcome of population-based SMA carrier screening.

Variable	2022.02–2022.12	2023.01–2023.10	Total
Received education women (n)	3600	3450	7050
Screened women (n)	3551	2484	6035
Acceptance rate, %	98.64	72	85.65
95% CI, %	98.21–98.97	70.48–73.47	84.76–86.40
Number of carriers (n)	50	50	100
Carrier rate, %	1.41	2.01	1.66
95% CI, %	1.07–1.85	1.53–2.64	1.37–2.01
Partner tested (n)	45	50	95
Recall rate, %	90	100	95
95% CI, %	78.64–95.65	92.87–100	88.82–97.85
Carrier couples (n)	0	1	1
Carrier rate of partner, %	0	2.00	1.00
Prenatal diagnoses (n)	0	1	1
Affected cases (n)	0	1	1
Pregnancies terminated (n)	0	1	1
Termination rate, %	0	100	100

Among the 100 identified SMA carriers with a heterozygous deletion of E7 in SMN1, 98 pregnant women had both E7 and E8 heterozygous deletions, and two had an E7 heterozygous deletion with a normal E8. Of the remaining 5935 women, 36 had normal E7 levels with a heterozygous deletion of E8, and the other 5899 pregnant women were normal within the range tested (Table 3). There were 95 spouses of these pregnant women recalled for genetic testing of SMN1. The overall recall rate was 95%. The recall rates are 90% in 2022 and 100% in 2023 (Table 2). Among the tested spouses, the deletion states of E7 and E8 of the SMN1 gene were detected. Finally, one couple was identified as SMN1 deletion carriers, and their child was at a high risk of developing SMA after birth. The couple already had a 5-year-old daughter who, for safety reasons, was also screened for SMA and turned out to be a carrier (Table 4), which has no impact on her life, but it is recommended that her spouse be tested for the deletion of the SMN1 gene when she marries or gives birth.

TABLE 3 Results of preliminary screening using real-time fluorescence quantitative polymerase chain reaction.

SMN1 exon 7	SMN1 exon 8
Heterozygous deletion	Normal	Total	Rate, %	95% CI, %
Heterozygous deletion	98	2	100	1.66	1.37–2.01
Normal	36	5899	5935	98.34	97.99–98.63
Total	134	5901	6035	100

TABLE 4 Prenatal diagnosis of fetuses of spinal muscular atrophy carrier parents.

Family	SMN1 of mother	SMN1 of father	SMN1 of sister	SMN1 of fetus	Pregnancy outcomes
1	E7 and E8 heterozygous deletion	E7 and E8 heterozygous deletion	E7 and E8 heterozygous deletion	E7 and E8 homozygous deletion	Pregnancy terminated

Results of Prenatal Diagnosis

Based on informed selection and genetic counseling, information on the clinical characteristics, genetic patterns, reproductive risk, and treatment of SMA was obtained. The fetus at high risk for SMA underwent amniocentesis and prenatal genetic testing, and QF-PCR analysis of STR loci in maternal blood and amniotic fluid samples showed no maternal cell contamination (the allele of at least one STR locus in the maternal sample was not consistent with the corresponding locus in the fetus). The qPCR results revealed that the fetus had a homozygous deletion of E7 and E8 of SMN1 (Figure 1). An MLPA assay was used to verify the results of qPCR (Figure 2). In the MLPA reaction, each pair of probes hybridized with the target sequence of the sample to be tested after modification and separated multiple target sequences in a tube reaction through ligation, amplification with universal primers, and capillary electrophoresis, and then compared and analyzed the relative copy number. The gene copy number is a duplicate of the MLPA probe ratio. When both copies of the gene were present, the ratio of the MLPA probe was 1; when the sample gene is heterozygous and there is only 1 copy, the ratio of the MLPA probe is 0.5. In this test, the ratio of exon 7 and exon 8 probe is 0, and the copy number of exon 7 and exon 8 of the SMN1 gene was both 0 in the subjects; the probe ratio was 1, and the corresponding copy number of the SMN2 gene was 2 (Figure 2). SMN1 is the main pathogenic gene of SMA, and about 95%–98% of SMA patients have an SMN1 homozygous deletion of exon 7; the copy number of SMN2 is correlated with the severity and prognosis of SMA; the more copies of SMN2, the less severe the symptoms. This subject fits the genetic variation pattern of SMA caused by the homozygous deletion of exon 7 of the SMN1 gene. Therefore, the fetus was diagnosed with SMA. After adequate genetic counseling, the parents decided to terminate the pregnancy (Table 4).

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Discussion

SMA is an autosomal recessive genetic disease caused by SMN1 mutations. It is the most common fatal genetic disease in infancy, with a general carrier rate of 1/40–1/60. Given the high population carrier rate of SMA and the disease severity, screening during pregnancy is critical. Studies have shown that over 95% of SMA cases are caused by homozygous deletion of E7 or both homozygous deletions of E7 and E8 of the SMN1 gene (Qu, Bai, and Cao 2016; Wirth, Herz, and Wetter 1999). Therefore, copy number testing for E7 and E8 of the SMN1 gene has become a common method for screening SMA carriers.

In this study, a total of 7050 pregnant women received SMA screening education, among which 6035 pregnant women voluntarily accepted SMA screening, with an acceptance rate of 85.60%, which is much higher than the acceptance rate reported in Taiwan in 2011 (10.6%) (Su et al. 2011); Zhang et al. (2023) reported that 33,560 pregnant women were educated about SMA screening, and 4429 pregnant women voluntarily accepted SMA screening, with an acceptance rate of 13.2%. Zhang, Wang, Ma, et al. (2020) reported that 34.9% of 16,549 pregnant women in Nanjing underwent screening. SMA screening is increasingly performed in various regions of China; however, the acceptance rate is still much lower than that in other countries. Prior, Snyder, and Rink (2010) reported that 98.7% of the US population underwent SMA carrier screening. Ben-Shachar, Orr-Urtreger, and Bardugo (2011) reported that 93% of the Israeli population required screening for SMA carriers. The reasons for the low acceptance of SMA in other regions in China are mainly related to the lack of understanding of the disease; a few people believed that they had no family history and that the incidence rate of SMA was low. Another reason is related to the test cost, which has not been covered by insurance. Therefore, it is important to strengthen education on SMA screening and the implementation of related policies by government agencies to increase the acceptance of SMA screening.

In the primary screening stage, 100 SMA carriers were screened from 6035 pregnant women using qPCR. The overall carrier rate was 1.66%. These results are per published data from different regions of China. The population carrying rate of pregnant women in Ningde was closest to that in Yancheng (1.64%) (Zhang et al. 2023), which is higher than that in Liuzhou (1.23%) (Tan et al. 2018), Zhaoqing (1.44%) (Huang et al. 2023), and Hong Kong (1.58%) (Chan et al. 2004), and lower than that in Taiwan (2.10%) (Su et al. 2011), Shanghai (1.91%) (Gong et al. 2013), Sichuan (2.11%) (Zeng et al. 2014), Yunnan (2.03%) (Zhang, Wang, He, et al. 2020), Nanjing (1.77%) (Zhang, Wang, Ma, et al. 2020), Qingdao (1.71%) (Wang n.d.), Guiyang (2.29%) (Li, Luo, and Hu 2021), Taiyuan (2.19%) (Deng, Guo, and Zhang 2022), and other areas (Table 5). At the international level, the probability of SMA carriers in Europe ranges from 1.25% to 2.00% (Lyahyai, Sbiti, and Barkat 2012), and the carrier frequencies in Korea and Israel are approximately 2.13% and 1.67% (Joel, Itamar, and Ehud 2016; Tae-Mi, Sang-Wun, and Kwang-Soo 2004), respectively. In Australia, cystic fibrosis (CF), fragile X syndrome (FXS), and SMA genetic carrier screening were simultaneously performed on 12,000 subjects; 241 SMA carriers were found, with a probability of about 2.01% (Archibald, Melanie, and Trent 2018). In a pan-ethnic population, SMA carrier frequency research for six major ethnic groups in the United States, including African American, Hispanic, Ashkenazi Jewish, Asian, Asian Indian, and Caucasian, ranges from 0.98% to 2.02% (Sugarman, Nagan, and Zhu 2012). According to current reports, the frequency of SMA varied among different countries and regions, which ranges from 0.98% to 2.29%. SMA screening should be carried out according to the local situation.

TABLE 5 Spinal muscular atrophy carrier rates in the different regions of China.

Area	Survey population (n)	SMA carriers (n)	Carrier frequency of (%)	Literature reference
Taiwan	107,611	2262	1/48 (2.10)	Su et al. (2011)
Hong Kong	569	9	1/63 (1.58)	Chan et al. (2004)
Shanghai	4719	90	1/55 (1.91)	Gong et al. (2013)
Sichuan	427	9	1/47 (2.11)	Zeng et al. (2014)
Liuzhou, Guangxi	4931	61	1/80 (1.23)	Tan et al. (2018)
Yunnan	3049	62	1/49 (2.03)	Zhang, Wang, He, et al. (2020)
Zhaoqing, Guangdong	5200	75	1/69 (1.44)	Huang et al. (2023)
Nanjing	13,069	231	1/56 (1.77)	Zhang, Wang, Ma, et al. (2020)
Qingdao	1118	19	1/57 (1.71)	Wang (n.d.)
Guiyang	524	12	1/44 (2.29)	Li, Luo, and Hu (2021)
Taiyuan	910	20	1/46 (2.19)	Deng, Guo, and Zhang (2022)
Jiangmen, Guangdong	14,378	251	1/57 (1.75)	Li, Shi, and Sun (2018)
Lianyungang	1266	25	1/51 (1.97)	Yin, Zhang, and Zheng (2023)
Jiangsu	5776	100	1/58 (1.73)	Zhang, Wang, and Ma (2021)
Yanchen, Jiangsu	4429	73	1/61 (1.64)	Zhang et al. (2023)
Shijiazhuang, Hebei	4568	126	1/36 (2.76)	Meng, Sun, and Yu (2021)
This study	6032	99	1/60 (1.67)

After the preliminary screening, the husband of the carrier pregnant woman was recalled for SMA testing. In this study, 95 spouses were recalled to undergo SMA screening. The recall rate is 90% in 2022 and is expected to increase to 100% by 2023. The recall rate was much higher than that reported by Zhaoqing (62.67%) and slightly higher than those reported by Nanjing (89.61%) and Taiwan (90.10%). It has been suggested that clinicians' education on SMA is well propagated, and SMA screening has been well carried out in this region. In the second stage of screening, one couple was demonstrated to be both carriers; therefore, their child was a high-risk fetus for SMA and required prenatal diagnosis to identify the state of the SMN1 gene. After detailed and adequate genetic counseling, amniocentesis was performed for genetic analysis of the fetus, which was confirmed to have a homozygous deletion of the SMN1 gene with two copies of the SMN2 gene, which may develop SMA type I after birth. After further genetic counseling and careful consideration, the couple chose to terminate the pregnancy. Our program prevented the births of children with SMA.

Accordingly, a standardized SMA screening process is critical for the clinical implementation of the program (Figure 3). A three-stage screening process can be conducted to prevent the birth of children with SMA. (1) Pregnant women should be educated so that they can fully understand the etiology, clinical manifestations, and severity of SMA and improve screening acceptance, and then the voluntarily pregnant women should sign an informed consent form to inform the purpose, method, and limitations of the test. (2) For SMA carriers with E7 or E7/E8 deletions of the SMN1 gene, their spouses should be advised to undergo SMA screening. (3) If both couples are SMA carriers, genetic counseling and prenatal diagnosis should be recommended, and the high-risk fetus should be tested for the SMN1 gene. Complete genetic counseling, comprehensive risk assessment, and adequate residual risk notification are particularly important during the screening process.

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SMN1 detection methods include real-time fluorescence qPCR, denaturing high-performance liquid chromatography (DHPLC), Multiplex ligation-dependent probe amplification (MLPA), and next-generation sequencing (NGS). MLPA has a high degree of precision and is considered the gold standard. However, both DHPLC and MLPA have the disadvantages of complex experimental operations, laboratory pollution, high requirements for experimental conditions, difficult interpretation of results, and relatively high prices, which make them difficult to carry out on a large scale. NGS counted the total number of SMN1 and SMN2 copies and then analyzed the ratio of SMN1 reads and SMN2 reads according to C.840 C>T and finally concluded that each person carried several copies of SMN1 and SMN2. However, due to the high homology of SMN1 and SMN2 and the length of NGS sequencing read, it is difficult for conventional comparison methods to distinguish SMN1 and SMN2. Even with bioinformatic analysis, other technical methods are still needed to verify the results. Therefore, at present, NGS has not become a routine detection method for SMA in China, and it is mainly used for the differential diagnosis of SMA. The qPCR technology used in this study has the advantages of lower cost, simplicity, and rapidity; the sample is not easily contaminated, and the results are accurate and intuitive. This method is suitable for large-scale population screening across various regions.

However, the limitation of this technique is that there is a certain residual risk, and it can only detect the most common “1 + 0” carriers, and cannot identify the rare “2 + 0,”, “1 + 1^D,” and “2 + 1^D” carriers. “1 + 0” and “2 + 0” types refer to the presence of one or two genes on one chromosome and no genes on the other chromosome (Yuan, Xiong, and Chen 2010). “1 + 1^D” and “2 + 1^D” types were defined as one or two SMN1 genes on one chromosome and a point mutation or microdeletion of the SMN1 gene on the other chromosome. The result may be false-negative, resulting in a missed diagnosis. Therefore, the residual risk should be fully explained during screening, counseling, and prenatal diagnosis. Consequently, the actual carrier rate of SMA in this region may be slightly higher than the results of this study. Different techniques and methods have their own characteristics, and the combination of them can accurately detect SMN1 copy number variation and base mutation.

In recent years, research on the etiology and treatment of SMA has made rapid progress based on the genetics of the disease. Nusinersen, risdiplam, and zolgensma are the three disease-modifying treatments currently used in clinical practice. Nusinersen, the first FDA-approved SMA treatment, was officially approved by the US FDA in December 2016 (Clabom, Stevens, and Walker 2019). Nusinersen has the ability to modulate the splicing pattern of precursor mRNA derived from the SMN2 gene, thereby promoting an increased production of full-length SMN protein and augmenting the compensatory effect exerted by the SMN2 gene. Risdiplam is the first oral therapy approved for SMA and was approved in the United States in August 2020 (Dhillon 2020). As the first small molecule drug directly targeting RNA, its mechanism of action is to up-regulate the expression of SMN protein by targeting the splicing silencer N1 (ISSN1) sequence in the key splicing region of SMN2 intron 7. However, the two drugs are expensive and require lifelong administration; even if the drugs are included in medical insurance, numerous SMA families cannot afford the huge cost of treatment. In addition to possible drug therapy, gene therapy using viral vectors to replace SMN1 approaches has been evaluated for SMA. Currently, Zolgensma gene replacement therapy, which was approved for marketing in the United States in May 2019, is a therapy that introduces the cDNA of the SMN1 gene into the human body through adeno-associated viral serotype 9 (AAV9) as a carrier (Mendell, Al-Zaidy, and Rodino-Klapac 2021). This treatment works by introducing the normal SMN1 gene to produce the SMN protein that is lacking in SMA patients. Once treated, the body will produce SMN protein from the cDNA delivered by the AAV9, achieving a single dose, long-term effectiveness. However, one dose costs 2.125 million dollars. Both the drug therapy and gene therapy costs are expensive for treatment, making it unaffordable for most families. Therefore, carrier screening and prenatal diagnostics to avoid the birth of children are more economically significant than postnatal treatment.

Conclusion

In conclusion, this study aimed to provide a screening and diagnostic tool for SMA in pregnant women in the Fujian area, perform prenatal diagnosis for suspected SMA fetuses to determine whether it is SMA, and provide genetic guidance for the family according to the results to improve the eugenic rate of the family. Through an investigation of 6035 samples, this study preliminarily determined the carrier status of SMA mutations to be 1.66% in Ningde, Fujian Province, China. The prenatal genetic testing strategy used in this study avoided the birth of one SMA fetus and reduced the burden on society and families. In addition, the experiment established a simple, fast, and efficient SMA prenatal diagnosis process and further provided research ideas for the screening of SMA carriers in other regions to improve the rate of healthy populations.

Author Contributions

Lu JJ collected the data and wrote the initial draft; Zheng X carried out the experiment; Dong WX collected the samples and clinical information; Yang J analyzed the experimental data; Cao LY was in charge of article modification; and Fu XG was in charge of project guidance and experimental analysis; Zeng X, Wu Q, and Chen X provided genetic counseling.

Acknowledgments

The authors are very grateful to the pregnant women and their spouses who participated in this research and to the whole team in the laboratory of Prenatal Diagnosis. The authors thank the Editage editor for the valuable advice in editing the manuscript.

Disclosure

All the authors have read and approved the final version of the manuscript.

Consent

The authors have nothing to report.

Conflicts of Interest

The authors declare no conflicts of interest.

Data Availability Statement

The data that support the findings of this study are available from the corresponding author upon reasonable request.

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