Clinical utility of next-generation sequencing (NGS) technology in paediatric diseases: a Romanian experience
Belengeanu Valerica, MD1, 3, Diana Marian, MD2 and Cristina Popescu, PhD1, 3*
- Vasile Goldis Western University of Arad, Faculty of Medicine, Department of Genetics, Liviu Rebreanu Street 86, Arad, Romania
- Vasile Goldis Western University of Arad, Faculty of Medicine, Department of Dentistry, Liviu Rebreanu Street 86, Arad, Romania
- AMS Laborator Genetic Company, Division of Clinical Genetics, Turturelelor Street 62, Bucharest, Romania.
Abstract: Next-generation sequencing techniques have had a major contribution over the last decade through identification of disease-causing mutations in paediatric diseases. Rare diseases are typically severe, with genetic origin and in the vast majority of cases children are affected. Techniques such WES or WGS, have changed the approach toward diagnostic of rare disease in children. We evaluated 6 children with rare diseases using a high-density array, exon-sequencing and whole genome sequencing. We found that 80% of our patients harboured genetic variants classified as pathogenic or likely pathogenic, that contribute to their phenotype.
Keywords: NGS; WES; WGS; paediatric rare disease
It is estimated that 200 million people worldwide have unresolved rare genetic disease [1,2]. Rare diseases affect 6-8% of the EU population. Approximately 80% of rare diseases have a genetic origin, and new genomic technologies have important contributions in establishing the diagnosis . Approximate ~4% of births worldwide every year will be affected by a genetic disorder . Most of rare genetic diseases do not have a particular or specific clinical feature in the child. In clinical practice, genetic testing is approached step by step in order to achieve molecular diagnosis in patients with suspected genetic diseases. However, such a testing strategy is often time consuming, costly, and stressful for the families, particularly in clinically difficult cases. In some situations, after years of testing, patients and their families still have no answer.
Recent advances in modern genomic diagnostic technologies demonstrates the molecular implications in paediatrics pathology. The rapid evolution and widespread use of gene sequencing in clinical laboratories has allowed incredible progress in the genetic diagnosis of several paediatric disorders [5,6]. These new sequencing technologies, with a broad medical application, such as preimplantation, diagnostics, oncology, infectious diseases, have focused on massive parallelization, improved automation, speed and very importantly, price reduction. Next Generation Sequencing (NGS) services for DNA and RNA sample analysis is a powerful technique due to the ability to sequence samples with a much higher coverage and detect novel or rare variants. NGS offers several sequencing approaches, such as whole genome sequencing (WGS), whole exome sequencing (WES), transcriptome sequencing, methylome analysis, and more. Whole-exome sequencing in clinical practice comprises just over 1% of the genome providing sequence information for protein-coding regions [7-9]. The advantages of WGS testing can detect a wider range of genetic variations are the ability to discover new genes, highlighting new genetic variants including, structural variant such as numbered variants (CNV) and translocations, single nucleotide variants (SNV). Another potential advantage is to highlight variants present in the mitochondrial genome and new discovery of genes and molecular mechanisms associated with Mendelian disease. [10-13]. Recently, the largest cohort study performed up to date, recommends that WGS should be considered as standard second line or even first-line stand-alone test . In this article we present the genetic architecture of complex disorders in a group of children, based on NGS technologies, emphasizing part of the Romanian experience.
Materials and methods
Children with rare diseases, for whom no cause was identified, were investigated by WES or WGS due to referral from the Romanian specialist between 2019-2021. Children with various pathologies reported here, have been identified with genomic abnormalities in nuclear DNA or mitochondrial DNA. Consent for genetic testing and genetic diagnostic was given by patients, parents, or referring physicians. Analysis was performed in specialized lab according to stringent quality criteria and validation processes for variants detected by NGS.
Exome and genome sequencing (WES/WGS)
For WES analysis double stranded DNA capture baits against approximately 36.5 Mb of the human coding exome (targeting >98% of the coding RefSeq from the human genome build GRCh37/hg19) are used to enrich target regions from fragmented genomic DNA with the Twist Human Core Exome Plus kit. The generated library was sequenced on an Illumina platform to obtain at least 20x coverage depth for >98% of the targeted bases. An in- house bioinformatics pipeline, including read alignment to GRCh37/hg19 genome assembly, variant calling, annotation and comprehensive variant filtering were applied. All variants with minor allele frequency (MAF) of less than 1% in gnomAD database, and disease-causing variants reported in HGMD®, in ClinVar or in CentoMD® were considered. The investigation for relevant variants is focused on coding exons and flanking +/-20 intronic nucleotides of genes with a clear gene-phenotype evidence (based on OMIM® information). All potential modes of inheritance patterns are considered. In addition, provided family history and clinical information are used to evaluate identified variants with respect to their pathogenicity and causality. Variants are categorized into five classes (pathogenic; likely pathogenic; VUS; likely benign; benign). All variants related to the phenotype of the patient are reported. Variants with low quality and/or unclear zygosity are confirmed by orthogonal methods. Consequently, a specificity of >99.9% for all reported variants is warranted.
For WGS analysis genomic DNA was enzymatically fragmented, and libraries are generated by PCR-mediated addition of Illumina compatible adapters. The libraries are paired-end sequenced on an Illumina platform to yield an average coverage depth of ~30x. An in-house bioinformatics pipeline including read alignment to GRCh37/hg19 genome assembly, variant calling and annotation is used. Structural variant (SV) calling is based on the DRAGEN pipeline from Illumina. All variants with minor allele frequency (MAF) of less than 1% in gnomAD database, and disease-causing variants reported in HGMD®, in ClinVar or in CentoMD® are considered. While the evaluation is focused on coding exons and flanking +/-20 intronic bases, the complete gene region is interrogated for candidate variants with plausible association to the phenotype.
For the mitochondrial genome, sequence reads are aligned to the Revised Cambridge Reference Sequence (rCRS) of the Human Mitochondrial DNA (NC_012920) and variant calling is performed using validated in-house software. The pipeline confidently detects heteroplasmy levels down to 15%. Structural variant (SV) calling is based on the DRAGEN pipeline from Illumina. All identified variants are evaluated with respect to their pathogenicity and causality. Variants are categorized into five classes (pathogenic; likely pathogenic; VUS; likely benign; benign). All variants related to the phenotype of the patient are reported. Consequently, a specificity of >99.9% for all reported variants is warranted.
Case #1 Gardner Syndrome
A 12-year-old girl was sent for genetic evaluation from a dental clinic because the panoramic radiograph showed multiple anomalies Panoramic X ray reveals dental inclusions of 2.3, 2.5, 3.5 and 4.5, persistence of the primary teeth beyond the age by which they normally are shed and replaced by the permanent teeth (6.3, 6.5, 7.5 – with a complicated caries lesion, 8.5.). Close to the included teeth 2.3 and 2.5 which have a deformed root, there is a supernumerary tooth with an aspect of complex odontoma. At the level of both mandibular angles there are sclerotic lesions with exofitic extensions, heterogeneous structure and lobular contour – osteoma- the right one being more developed and more radio-opaque. Also, we observed diffuse and radio- opaque masses between the lower border of the mandible and the alveolar ridge, especially in the area between 3.6 and 4.6 and on the maxilla in the medial and inferior part of the right zygomatic posterior arch and in the left anterior part of the nasal bone. The maxillary sinuses have a radio-opaque structure (Figure 1). After dental investigation, a clinical suspicion of Gardner’s syndrome was raised, and further complex craniofacial CT investigations and scintigraphy were performed.
Figure 1. A panoramic image shows well-defined multiples teeth anomalies in suspected Gardner syndrome.
Case #2 Wiskot-Aldrich
The baby was born by cesarean section at 38 weeks, with an Apgar score of 8 and a weight of 2240 grams. (Rh incompatibility, rh positive baby). The affected skin peels, but on the 2nd, 3rd day his skin was almost reepithelialized. He presented 85,000 low platelets and increased CRP. The first dermatological diagnosis was ichthyosis. Latter then based on skin biopsy the diagnostic of bullous epidermolosis was concluded (Figure 2). Interestingly, skin manifestations were cyclical since they disappeared with cortisone treatment. At the age of 4 months, a blood count and low anemia were found and the infant had no symptoms of petechiae. The suspicion of WISCOTT-ALDRICH syndrome and neonatal herpes was raised, the mother having a positive IgM for herpes in pregnancy. The child did 7 days treatment with antibiotics but in parallel with cortisone and erythrocyte mass (since maternity).
Figure 2. The ulceration on lower abdomen (a), multiple vesiculobullous lesions (b), baby normal appearance at the age of 14 months (c) in Wiskot-Aldrich syndrome.
The baby eruptions after the interruption of cortisone were more and more severe (herpes-specific vesicular eruptions), now located only at the level of the scalp (Figure 3). Because he did not have the opportunity to feed, he was initially fed through the tube and then through the gastrostomy. It should be remembered that the baby never bled, had no otitis, urinary or respiratory tract infections, and no patches.
During his evolution, the infant was fed through the probe until the age of 13 months, and from this age onwards, gastrostroma was performed. Baby was hospitalized 4 times many days until 12 months of age and managed with fresh whole blood transfusion, broad spectrum antimicrobials and prednisone. Due to the low number of platelets, WAS gene analysis was indicated. The first genetic analysis indicated a mutation in the WAS-Wiscott Aldrich gene (exon 10, c.1291_1292dupe (Ala433Glufs * 13)). This hemizogotic variation has not been described in clinical and population frequency databases, or in the literature. It is possible that the variant found did not resonate in the phenotype regarding hemorrhagic manifestations and eczema.
Latter, WES analysis was performed (including NGS-based CNV analysis) and the result highlighted A hemizygous likely pathogenic variant was identified in the WAS gene. This finding is consistent with the genetic diagnosis of X-linked Wiskott-Aldrich syndrome. The WAS variant c.1291_1292dup p.(Ala433Glufs*13) creates a shift in the reading frame starting at codon 433.
At 12 months of age the child was hospitalized in a specilized clinic Rome waiting for a bone marrow transplant. During hospitalization he was treated with immunosuppressant therapy prior bone marrow transplant. It is spectacular that at the skin level no lesions appeared (Figure 3, c) The baby has an appearance of health.
Figure 3. The dysmorphism including a coarse face thick eyebrows, broad nasal tip, wide mouth with tick lips, low-set ears (a, b) dental anomalies (a, c) in Coffin-Sirris case.
Case #3 Coffin- Siris syndrome
The female patient, 13 years 6 months of age, was referred to geneticist for assessment for short stature, cognitive and developmental delay, atypical autistic behaviour. Clinically she is diagnosed with Ehlers-Danlos syndrome and subclinical hypothyroidism. General examination of the girl showed their height (1.40 cm), weight (31 kg) and head circumference (52 cm) to be below the third percentile for age. Coarse facies, thick eyebrows, teeth abnormalities (microdontic) except the upper central incisor dysmorphic and low-set ears, broad nasal tip, wide mouth with tick lips, hypertrichosis, hypoplastic of all toenails with clinodactyly of 5th fingers and joint laxity, hypotonia, and scoliosis (Figure 4). Karyotype analysis showed an apparently balanced de novo reciprocal translocation involving chromosomes 7 and 22, 46, XY, t (7;22). The clinical diagnosis in our patients was set by WES analysis, which identified the presence of de novo mutations of ARID1B gene.
Figure 4. Hypertrichosis (a), hypoplastic of all toenails with clinodactyly (b) in Coffin-Sirris case.
Case #4 Autistic behaviour
3-year 11-month-old boy who was diagnosed with ASD and borderline intellectual development, he presented with delays in social communication skills (lack of conversational speech, poor eye contact), repetitive and stereotyped behaviour. WGS analysis revealed two genetic variants. The first gene affected is DLG4 variant c.934G>A p.(Asp312Asn) that causes an amino acid change from Asp to Asn at position 312. It is classified as variant of uncertain significance (class 3) according to the recommendations of ACMG and CENTOGENE. Second pathogenic variant was identified in the GAA gene: NM_000152.3: c.-32-13T>G (rs386834236 ) but it’s effect in currently unknown. Pathogenic variants in the DLG4 gene have been associated with the autosomal dominant intellectual developmental disorder type 62(MRD62 (OMIM®: 618793). Lelieveld et al. (2016) reported 3 unrelated patients with MRD62. Clinical details were limited, but all had mild to moderately impaired intellectual development and motor delay. Two patients each had language delay, nonspecific dysmorphic facial features, and visual impairment; 1 patient had behavioural abnormalities.
Caz #5 Autistic behaviour
2-year-old boy with delayed language development, attention deficit, unstable gait, axial hypotonia, joint laxity (Fig 5). Clinical phenotype without other informative elements. The mother claims that the child has bruises even without a cause (Figure 5). The analysis found the ITGA2 variant c.1975G>C p.(Ala659Pro) that causes an amino acid change from Ala to Pro at position 659. It is classified as variant of uncertain significance (class 3) according to the recommendations of ACMG and Centogene. Pathogenic variants in the ITGA2 gene are associated with autosomal dominant fetal and neonatal alloimmune thrombocytopenia. Foetal and neonatal alloimmune thrombocytopenia (NAIT) results from maternal alloimmunisation against foetal platelet antigens inherited from the father and different from those present in the mother, and usually presents as a severe isolated thrombocytopenia in otherwise healthy new-borns. NAIT has been considered to be the platelet counterpart of Rh Haemolytic Disease of the Newborn (RHD). Unlike RHD, NAIT can occur during a first pregnancy. The spectrum of the disease may range from sub-clinical moderate thrombocytopenia to life-threatening bleeding in the neonatal period. Mildly affected infants may be asymptomatic. In those with severe thrombocytopenia, the most common presentations are petechiae, purpura or cephalohematoma at birth, associated with a major risk of intracranial haemorrhage (up to 20% of reported cases), which leads to death or neurological sequelae (Orphanet – ORPHA:853). The result of the genetic analysis forced a blood count, which had platelets in normal numbers.
Figure 5. Clinical phenotype in a child with autistic behavior. (a) police laxity; (b) bruising.
Case #6 sever development delay
5-month-old female infant investigated for microcephaly, facial dysmorphia, generalized severe hypotonia, repeated seizures, failure to thrive, plus cerebellar vermis hypoplasia (Figure 6). No clinically relevant variants, including copy number variations, related to the described phenotype were identified by whole exome sequencing (WES). However, by analysing the mitochondrial genome dataset, a variant of uncertain significance was identified in the MT-CO3 gene. The MT-CO3 variant: m. 9480T>C: p.Phe92Leu has been identified in in a homoplasmic state (100%). This is classified as variant of uncertain significance (class 3) according to the recommendations of ACMG and Centogene. Pathogenic variants of the MT-CO3 gene are associated with Leigh syndrome. Routine hemogram and serum lactate were in normal limits. An important role in treatment is anticonvulsant drugs. Because the girl had multiple seizures, daily, a pharmacogenetic analysis test was performed to select the appropriate treatment. Current medication (according to the data completed in the application form): Cobazam, Depakene, Desitin, Lamictal, levetiracetam, melatonin, phenobarbital. Medication without effect: carbamazepine, risperidone, topiramate, valproic acid. The pharmacogenetic test indicated a cumulative drug-drug and drug-gene interaction. The impact was moderate regarding phenobarbital-CYP2C9 intermediate metabolizer-Depakene. Also, a moderate interaction was found between Lamictal- phenobarbital-Depakene. Recommendations for monitoring and dose adjustments were made. The girl’s evolution was and still is unfavourable and she is frequently hospitalized for seizures and failed to develop.
Figure 6. Facial dysmorphia in a baby girl suspected of Leigh’s syndrome: triangular face, hypertelorismm, thick lips, microretrognathia.
In clinical practice, the diagnosis is burdened both by phenotypic manifestations that are not always specific indicators of the disease but also by the genetic and phenotypic heterogeneity of the disease [15-18]. Therefore, the clinically oriented approach to the diagnosis of genetic diseases is often not successful, unless the phenotype is very specific to a genetic disease. In some cases, the results of the NGS analysis confirmed the suspected clinical diagnosis, but most of the time the genetic results led to a new diagnosis or to the enunciation of an unspecified clinical diagnosis. In some cases, DNA variants of undetermined significance have been identified but in other cases, we found clearly pathogenic variants such as, case #1, case #2, case #3, and they had a clear genotype-phenotype correlation, with albeit some atypical features in Wiskott-Aldrich syndrome.
Case #1: Gardner’s syndrome is a clinical subgroup of Familial Adenomatosus Polyposis, is characterized by multiple intestinal polyps, osteomas, and epidermoid cysts. Supplementary, one can observe dental abnormalities, increased frequency of multiple odontomas, as well as supernumerary or unerupted teeth, impacted teeth and root anomalies. Skeletal abnormalities, the most common of which are osteomas, specific of Gardner syndrome they were also present in our case . Comparison of the identified mutation in our case, gene APC variant: c.4701_4707delinsGTTCT p. (Glu1568Phefs * 2) which creates a change in the reading frame starting from codon 1568. Comparing the result with data from the literature, we confirm a genotype-phenotype relationship. There was no evidence of familial inheritance. This case seems to represent sporadic mutation. The patient had the great chance for the disease to be suspected early and thus a personalized surveillance management was elaborated. Osteoma is a benign tumour of bone with slow but continuous growth and is the most common bone lesion seen in Gardner syndrome Osteomas appear in about half of Gardner’s syndrome patients presented with 3 or more osteomas in the mandible and maxilla as well as in other locations such as long bones and even phalanges may be affected. As it turned out in the investigations, the patient had multiple osteomas located at the craniofacial level and both joints of the ankle and at both joints of the hand. Osteomas often appear 10 years before the manifestations of polyposis and can be considered a clinical marker for early diagnosis of the disease [20, 21]. Similar to data literature we observed that vast majority of osteomas were located on the skull. Dentists can play an important role in diagnosing this syndrome. The presence of osteomas and unerupted teeth provides the dentist with a major pointer clinical suspicion four Gardner’s syndrome.
Cases #2: The Wiskott-Aldrich syndrome is a rare X-linked recessive disease, with immunodeficiency, micro-thrombocytopenia, eczema and increased susceptibility to pyogenic and opportunistic infections  and increased risk in developing autoimmunity and lymphomas . The typical skin lesions in this syndrome resemble acute or chronic eczema in appearance and distribution. But eczema develops in 80% of the patients and is heterogeneous in severity and persistence . Haemorrhages are frequent (80% incidence) in WAS and range from light manifestations (epistaxis, petechiae, purpura, oral bleeding) to severe manifestations, such as intestinal and intracranial bleeding . Our case did not have eczema and haemorrhagic manifestations. The correlation between phenotype and genotype in this syndrome is debatable, some studies support the correlation [25-27] other studies have not found this correlation . A Japanese study of 50 patients on the clinical features of WAS syndrome argued that the clinical context is dependent on the presence or absence of WAS protein because they found that the lack of WASP is directly or indirectly responsible for the eczema observed in WAS patients .
Case #3: Coffin-Siris Syndrome (CSS) is a rare, congenital malformation syndrome with a variable display of phenotype. The characteristic manifestations proposed the minimal criteria for diagnosis are mental retardation, coarse facial features, hypertrichosis and hypoplastic or absent fingernails or toenails . Our patient meets these criteria associated with other manifestations specific to the syndrome. Manifestations that hypothyroidism present in the case was documented in 19.0% of cases and hypotonia of the face occurred in 81.0% of patients . Mutations in the gene encoding AT-rich interactive domain-containing protein 1B (ARID1B) are associated with multiple syndromes with developmental delay and intellectual disability, in addition to non-syndromic intellectual disability. The major features associated with mutations the gene ARID1B are intellectual disability, speech delay, coarse facies, and hypertrichosis. Mutations in ARID1B and several other genes encoding components of the Brahma-associated factor (BAF, also referred to as switching defective and sucrose nonfermenting SWI/SNF-α) chromatin remodelling complex, were recently shown to cause Coffin-Siris syndrome (CSS) [32,33]. Data from the literature specify that ARID1B mutations are associated with a milder phenotype compared to the phenotype determined by mutations in the SMARCB1 or SMARCE1 genes that may be the cause of the syndrome .
Cases #4 and #5: genetic causes Autism Spectrum Disorder (ASD) it is only partially deciphered, due to a high degree of genetic and phenotypic heterogeneity. Although the implications of the genetic changes found in ASD vary from study to study, it is clear that they play an important role. Several studies have shown that a number of genes are associated with ASD susceptibility, but variants for a single gene have been estimated for approximately 400 genes or chromosomal regions involved in the predisposition to this pathology [35, 36]. According to other data bases these numbers are as follows: 913 genes catalogued and scored for their strength of association to ASD ; 1379 syndromic and non-syndromic autism-related genes; 5420 CNVs or structural variations (SVs), 11,669 single-nucleotide variations (SNVs)/insertions and deletions (InDels) and 172 linkage regions associated with ASD .
Data from the literature specify that the heterozygous mutations in the DLG4 gene that were identified in patients with MRD62 by Lelieveld et al. (2016) and Moutton et al. (2018), occurred de novo [39, 40]. Not so many clinical data are available, and they varied from case to case. In some patient language delay, nonspecific dysmorphic facial features, visual disturbance and behavioural dysfunctions was observed . Other patients had in addition to intellectual and speech impairment a marfanoid phenotype with highly arched palate, long face, long thin fingers, pectus or foot deformities, mild scoliosis, but and autism spectrum disorder and seizures . The variant identified in the DLG4 gene is not sufficiently known, being classified as the variant with uncertain significance. The complexities of penetrance and variable expressivity of this DLG4 gene in phenotype, might be partially explained by gene interacting with other molecules, and perhaps nongenetic factors that might impede their homeostasis. It is necessary and useful that after a period of reanalysing the data regarding the classification of variants in the DLG4 gene.
As for the pathogenic variant in the GAA gene that is in the heterozygous state, the status of carrier results in the child. The patient has no relevant symptoms at this time. It is perhaps especially interesting to note that the apparently unaffected father carried the same mutations as the ASD affected child.
Pathogenic variants in the ITGA2 gene found in our case, are associated with autosomal dominant fetal and neonatal alloimmune thrombocytopenia. Neonatal alloimmune thrombocytopenia (NAIT) occurs in one in 1,000–1,500 live birth, is characterized by maternal alloimmunization against fetal platelet antigens inherited from the father. Most cases of diseases are diagnosed incidentally, shortly after birth, the first-born infant is found to have thrombocytopenia with petechiae, ecchymosis, or bleeding. Depending on the severity of thrombocytopenia, NAIT varies the spectrum of disease and some neonates may be asymptomatic [41-43]. The child results from the mother’s first pregnancy. Based on the child’s personal history, correlated with the maternal history, we cannot claim that the genetic variant found in the gene is the cause of the manifestations in the child. It remains under investigation focused on NAIT signs, in order to overlap phenotype with the variant detected.
Case #6: Leigh’s syndrome, a mitochondrial disorder, is a rare inherited neurometabolic disorder affects the central nervous system, presenting in the infancy or childhood. It progresses rapidly and often causes death. The MT-CO3 gene encodes cytochrome c oxidase subunit III, which is 1 of 3 mitochondrial DNA (mtDNA) encoded subunits of respiratory Complex IV. Pathogenic variants in this gene are associated with Leber optic atrophy, mitochondrial complex IV deficiency, seizures and lactic acidosis among other phenotypes (OMIM: 516050). This variant found in this child was mentioned for the first time in a patient with sensorineural hearing impairment without any evidence . Various mutations in mtDNA-encoded tRNA genes have been described as causing Leigh syndrome . Several studies showed significant heterogeneity in the clinical findings of Leigh syndrome and it is known that certain genetic variations are also found in some nuclear genes, not just mitochondrial ones [46,47]. The clinical features of Leigh syndrome with the mtDNA mutations are not specific, showing the common features of developmental delay, muscle hypotonia, and seizures, manifestations that were present in our case as well. A number of factors influence the prognosis such as the age of onset of the disease, the mutation in the gene, the seizures, the cerebral abnormalities found, the failure to gain weight and the supervision by intensive care . We cannot say whether the variant present in the patient negatively influences the evolution of the disease, because the patient is very often hospitalized. Newly identified genetic causes are increasing, due to next-generation and whole-exome sequencing [49-51].
The complexities of penetrance and variable expressivity of this gene in ASD might partially explained by gene interaction with other molecules and perhaps nongenetic factors that might impede their homeostasis. Due to the widespread clinical and genetic heterogeneity and pleiotropism in ASD, genetic causes remain an area to be exploited. Thus, whole genome sequencing (WGS) seems to be the ideal method at present to identify undetected genes or new variations in known genes. Genetic variants in ASD reported in this study are extremely rare because they are not frequently present in public databases.
It is important in medical practice that these new technologies for affected families will bring data for better understanding the genetic basis of rare diseases, will contribute to a more accurate prognosis and management, genetic surveillance and counselling . NGS genetic testing has a wide application in many countries around the world. In Romania, it has been successfully implemented in the big cities where there are universities of medicine and geneticist. This situation highlights the unequal access of patients to genetic testing in Romania. The explanations are as follows: on the one hand, the lack of information of the population about the benefit of genetic testing, and the still prohibitive price for the population, especially the rural one, which exceeds the income of a family. NGS tests are still costly in Romania, because the health policy does not support at least partially the cost of the test.
Currently, the concern is to convert pathological genetic data into a diagnostic analysis that can adjust and finalize clinical decisions and patient management. We must also remember the decisive role in development of new and personalized therapies. Based on genetic data on the etiology of the disease, the patient can be properly informed of management such as hematopoietic stem cell transplantation, immunomodulation or gene therapy.
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