Haemophilia management programme: Transformation during COVID-19

Background & objectives: Haemophilia is a debilitating bleeding disorder with significant comorbidities affecting the quality of life. In India, the management of these individuals is still limited to on-demand institutional treatment with coagulant factors. In this study, we highlighted the problems faced by these patients in the COVID-19 period due to nationwide lockdown. Methods: A retrospective study was done to ascertain the trend in the number of patients with haemophilia A and B visiting the hospital, those succumbing to haemophilic complications and indications for factor requirement in the pre-COVID (October 2019-March 2020) and during the COVID-19 period (April-September 2020). Representative cases with unusual complications were described along with significant challenges faced in providing standard care of treatment to these individuals due to the COVID-19 pandemic. Results: A total of 818 and 162 individuals with haemophilia A and B, respectively, were registered with the department. The overall number of patient visits to the hospital significantly reduced from an average of 6.9 outpatient department (OPD) visits per patient in the pre-COVID-19 period to an average of 3.9 OPD visits per patient and admissions reduced to 50 per cent during the COVID-19 period. This led to a reduction in utilization of factors VIII and IX except VIIa for haemophilia with inhibitors. There was no factor utilization for elective surgeries during the COVID-19 period. A total of eight patients succumbed to haemophilia-related complications during the COVID-19 period due to delay in reaching the hospital. The challenges faced in the management of three cases with musculoskeletal bleeds, one case with scrotal haematoma and one with haemothorax during the COVID-19 period were also highlighted. Interpretation & conclusions: COVID-19 pandemic has unveiled the need for on-demand home treatment with coagulant factors and has also brought to light the existing need for primary prophylaxis, especially for younger individuals with haemophilia.

Congenital Hyperinsulinemia of Infancy: Role of Molecular Testing in Management and Genetic Counseling

Congenital hyperinsulinemia (CHI) is a genetically and clinically heterogenous disorder. In addition to the standard care of management of the proband, genetic counseling regarding the risk of recurrence in the future siblings is an important part in the management of the disorder. The counseling needs identifcation of accurate etiology and is challenging due to the complexity of the molecular mechanisms of CHI. This case highlights the importance of molecular testing which not only helped in planning the management of the proband with CHI but also helped in providing genetic counseling for which the family had consulted the medical genetics department.

Prenatal diagnosis of congenital high airway obstruction syndrome: our experience from a tertiary care center

Congenital high airway obstruction syndrome (CHAOS) is an extremely rare and life-threatening condition. It occurs due to obstruction in fetal respiratory tract and is characterized by typical ultrasonographic findings. Risk of recurrence is low, so antenatal diagnosis can help in counselling regarding risk of recurrence. A retrospective record review of all cases referred to our institution for antenatal ultrasound over a period of 5 years from January, 2014 to December, 2018 was done. Cases diagnosed as CHAOS were reviewed in detail regarding the radiological findings, information regarding delivery, fetal karyotype and postnatal/ fetal examination. Between the period of 2014 to 2018 three fetuses with CHAOS were identified. All of them had characteristic radiological features. Two of them were associated with hydrops and one fetus had oligohydramnios. All the pregnancies were terminated after antenatal diagnosis. Amniocentesis was done in 2 out of 3 cases and fetal karyotype was found to be normal. Fetal autopsy was done in one case and site of upper airway obstruction was identified. Confirmation of diagnosis by antenatal ultrasound and if possible, by post-mortem examination is essential for providing estimation of risk of recurrence to the family and genetic counselling.

Recurrent miscarriage in North Indian population: a study of association of polymorphisms in genes coding for the natural killer: cell receptor natural killer group 2, member D and its ligand MHC class I chain-related protein A

Background: The objective of this present study was to investigate the possible association of natural killer group (NKG) receptors gene polymorphisms and MHC class I chain-related protein A (MICA) gene polymorphism with recurrent spontaneous abortion (RSA).Methods: Three single-nucleotide polymorphism (SNPs) in NKG2D gene (rs2255336, rs2617160 and rs2617170) and one SNP in MICA gene (MICA129) rs1051792 were assessed in 100 controls and 100 patients employing polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and agarose gel electrophoresis.Results: NKG2D (rs2617160) and MICA 129 (rs1051792) variants are associated with RSA risk in North Indian women.Conclusions: The NKG2D and MICA129 gene polymorphisms may influence the success of pregnancy in North Indian women population.

Hypotonic infant with Pallister–Killian syndrome diagnosed by cytogenetic microarray, without pigmentary skin changes and malformations

Pallister–Killian syndrome (PKS) is a rare genetic developmental disorder characterized, by intellectual disability, seizures, streaks of hypo- or hyperpigmentation and characteristic dysmorphic features. PKS is characterized by the presence of cytogenetic abnormality in form of a supernumerary isochromosome 12p, in a tissue limited mosaicism. The isochromosome 12p is usually not detected in karyotype done from peripheral blood. Presence of patchy pigmentary skin lesions suggest the possibility of mosaicism and karyotype from skin is done which clinches the diagnosis. We describe an infant with severe hypotonia in whom trisomy 12p was detected by chromosomal microarray performed on peripheral blood. The karyotype from blood was normal and combining this information with three copies of 12p in microarray suggests the possibility of tetrasomy12p in mosaic form. The infant did not have any skin patchy pigmentary changes and malformations and hence, the diagnosis of PKS was not clinically suspected. Cytogenetic microarray is the first test for evaluation of cases with developmental delay and intellectual disability, PKS diagnosis may come as a surprise in unsuspected cases without characteristic skin pigmentary abnormality and malformations.

Computer-aided Facial Analysis in Diagnosing Dysmorphic Syndromes in Indian Children

Objective: To assess the utility of computer-aided facial analysis in identifying dysmorphicsyndromes in Indian children. Methods: Fifty-one patients with a definite molecular orcytogenetic diagnosis and recognizable facial dysmorphism were enrolled in the study andtheir facial photographs were uploaded in the Face2Gene software. The results provided bythe software were compared with the molecular diagnosis. Results: Of the 51 patients, thesoftware predicted the correct diagnosis in 37 patients (72.5%); predicted as the first in thetop ten suggestions in 26 (70.2%). In 14 patients, the software did not suggest a correctdiagnosis. Conclusions: Computer-aided facial analysis is a method that can aid indiagnosis of genetic syndromes in Indian children. As more clinicians start to use thissoftware, its accuracy is expected to improve.

Vici Syndrome with a Novel Mutation in EPG5

Background: Vici syndrome is a neurodevelopmental disorder of the autophagy pathway.Almost all cases reported have the cardinal features of agenesis of corpus callosum,cataract, cardiomyopathy, immunodeficiency and hypopigmentation. Casecharacteristics: 8-month-old boy with developmental delay, myoclonic jerks, repeatedrespiratory infections, coarse facial features, cataract and hypopigmented hair.Echocardiography revealed dilated cardiomyopathy and magnetic resonance imaging ofbrain suggested agenesis of corpus callosum. Exome sequencing detected a novelhomozygous nonsense mutation in the EPG5 gene. Outcome: Establishing a definitediagnosis helped in proper prognostication, providing genetic counseling and prenataldiagnosis to the family. Message: Though uncommon, presence of the characteristicfeatures makes Vici syndrome a clinically recognizable cause of developmental delay.

Cytogenetic microarray in structurally normal and abnormal foetuses: a five year experience elucidating in creasing acceptance and clinical utility

The aim of the present study was to evaluate the diagnostic yield of prenatal cytogeneticmicroarray (CMA) in structurally normal and abnormal foetuses and record the acceptance rate of CMA for prenatal diagnosis over a course of five year. In 128 structurally normal and abnormal foetuses, CMA was performed along with foetal karyotype, after exclusion of aneuploidy by quantitative fluorescence polymerase chain reaction. The microarray was able to detect the pathogenic variants in 5.5% cases; the diagnostic yield in structurally abnormal foetuses was 8.8% and 4.7% in foetuses with a high aneuploidy risk. Balanced and unbalanced translocations, and low level mosaicism were detected. Reanalysis of variants of uncertain significance identified pathogenic variant. The study shows higher diagnostic yield in structurally abnormal cases, the importance of foetal karyotype and reanalysis in microarray. The acceptance rate of prenatal CMA increased five-fold over a period of five year

Molecular Testing of MECP2 Gene in Rett Syndrome Phenotypes in Indian Girls

Objective: To assess yield of MECP2 gene sequence variationsanalysis and large deletions in suspected cases of Rettsyndrome.Design: Descriptive study.Setting: Tertiary-care medical genetics center.Patients: Girls with neuroregression, postnatal microcephaly andsigns and symptoms suggestive of classical and atypical Rettsyndrome were classified into two groups. Group I consisted ofgirls with Classical and atypical Rett syndrome on basis on theRevised Rett Syndrome diagnostic criteria, 2010. Group II includedgirls with neuroregression and postnatal microcephaly and otherRett like features but not fulfilling the above criteria.Procedure: Sanger sequencing of coding regions and largedeletional analysis of MECP2 gene.Outcome measure: Identification of mutation in MECP2 gene.Result: Mutation in MECP2 gene was identified in 74% (14/19) ingroup I and none (0/17) in group II. The mutation detection ratewas 93% (13/14) in group I classical Rett syndrome girls (2 withlarge deletions identified with Multiplex ligation dependent probeamplification) and 20% (1/5) in group I atypical Rett syndromegirls. One novel MECP2 sequence variation was identified ingroup I classical Rett syndrome.Conclusion: The yield of the mutation detection in MECP2 ishigher in classical Rett syndrome. In girls with some Rett likefeatures, but not fulfilling revised Rett syndrome diagnosticcriteria, mutation testing for MECP2 gene has a low yield

Diagnosis and Management of Gaucher Disease in India – Consensus Guidelines of the Gaucher Disease Task Force of the Society for Indian Academy of Medical Genetics and the Indian Academy of Pediatrics

Justification: Gaucher disease (GD) is amongst the most frequently occurring lysosomal storage disorder in all ethnicities. The clinicalmanifestations and natural history of GD is highly heterogeneous with extreme geographic and ethnic variations. The literature on GD haspaucity of information and optimal management guidelines for Indian patients.Process: Gaucher Disease Task Force was formed under the auspices of the Society for Indian Academy of Medical Genetics. Invitedexperts from various specialties formulated guidelines for the management of patients with GD. A writing committee was formed andthe draft guidelines were circulated by email to all members for comments and inputs. The guidelines were finalized in December 2016at the annual meeting of the Indian Academy of Medical Genetics.Objectives: These guidelines are intended to serve as a standard framework for treating physicians and the health care systems foroptimal management of Gaucher disease in India and to define unique needs of this patient population.Recommendations: Manifestations of GD are protean and a high index of suspicion is essential for timely diagnosis. Patients frequentlyexperience diagnostic delays during which severe irreversible complications occur. Leucocyte acid ?-glucosidase activity ismandatory for establishing the diagnosis of Gaucher disease; molecular testing can help identify patients at risk of neuronopathicdisease. Enzyme replacement therapy for type 1 and type 3 Gaucher disease is the standard of care. Best outcomes are achieved byearly initiation of therapy before onset of irreversible complications. However, in setting of progressive neurological symptoms such asseizures and or/ neuroregression, ERT is not recommended, as it cannot cross the blood brain barrier. The recommendations herein arefor diagnosis, for initiation of therapy, therapeutic goals, monitoring and follow up of patients. We highlight that prevention of recurrenceof the disease through genetic counseling and prenatal diagnosis is essential in India, due to uniformly severe phenotypes encounteredin our population

Metatropic Dysplasia with a Novel Mutation in TRPV4.

Back ground: Metatropic dysplasia is a skeletal dysplasia characterized by rhizomelia, severe kyphoscoliosis and a coccygeal tail. Case characteristics: A 12 day-old male neonate had facial dysmorphism, short limbs and coccygeal tail and showed radiological features of metatropic dysplasia. Observation: A novel heterozygous variant was observed in TRPV4 gene. Message: We report a novel mutation in an Indian neonate with metatropic dysplasia.

Worsening of Callus Hyperplasia after Bisphosphonate Treatment in Type V Osteogenesis Imperfecta.

Background: Type V osteogenesis imperfecta is characterized by hyperplastic callus formation and interosseus membrane calcification. Case characteristics: A 16-year-old boy who presented with history of recurrent fractures, had hard persistent swellings at fracture sites, and had radiographic features of hyperplastic callus and interosseus membrane calcification. Outcome: Sequence analysis of the IFITM5 gene revealed the c.-14 C>T mutation. The patient had significant exacerbation of callus hyperplasia after initiation of bisphosphonate therapy, which reversed following cessation of the treatment. Message: Bisphosphonates may exacerbate callus hyperplasia, and may therefore have to be used with caution in patients with type V osteogenesis imperfecta.

Hunter Syndrome in Northern India: Clinical features and Mutation Spectrum.

Objective: To study the clinical profile and mutation spectrum of Hunter syndrome. Methods: Evaluation of 18 cases of Hunter syndrome from 17 families was done. Mutation analysis of Iduronate sulfatase (IDS) gene was done in 9 families, and mothers of four affected children with no family history. Results: Joint contracture, hepatomegaly and radiological changes were present in all children. 6 (33%) children had normal cognitive function at presentation. Point mutations were identified in all the 9 families for whom mutation analysis was done. Among 4 mothers tested from families without any family history, 2 (50%) were found to be carriers. Conclusion: Accurate etiological diagnosis by mutation analysis of IDS gene is important in Hunter syndrome.

Recurrent benign copy number variants & issues in interpretation of variants of unknown significance identified by cytogenetic microarray in Indian patients with intellectual disability.

Background & objectives: Cytogenetic microarray (CMA) is now recommended as a first-tier clinical diagnostic test in cases with idiopathic intellectual disability and/or developmental delay (ID/DD). Along with clinically relevant variants, CMA platforms also identify variants of unknown significance (VUS). This study was done to look for utility and various issues in interpretation of copy number variants (CNVs) in Indian patients with ID/DD. Methods: The CMA was performed in 86 Indian patients with idiopathic ID/DD with or without dysmorphic features. CNV was reported if copy number gain was >400 kb in size and copy number loss was > 200 kb in size. Results: Pathogenic CNVs were found in 18 of 86 (20.9%) patients. One large (14 Mb size) de novo heterozygous copy number gain was found in one patient. VUS (total 31) were present in 17 of 86 (19.7%) patients. Five novel recurrent benign CNVs were also present in our patients. Interpretation & conclusions: Our findings highlight the difficulties in interpretation of CNVs identified by CMA. More Indian data on VUS and recurrent benign CNVs will be helpful in the interpretation of CMA in patients with ID/DD.

Novel mutations of the arylsulphatase B (ARSB) gene in Indian patients with mucopolysaccharidosis type VI.

Background & objectives: Mucopolysaccharidosis type VI (MPS VI) is a rare, autosomal recessive lysosomal storage disorder caused by deficient enzymatic activity of N-acetyl galactosamine-4-sulphatase resulting from mutations in the arylsulphatase B (ARSB) gene. The ARSB gene is located on chromosome 5q11-q13 and is composed of eight exons. More than hundred ARSB mutations have been reported so far, but the mutation spectrum of MPS VI in India is still unknown. Hence, the aim of the present study was to identify the mutational spectrum in patients with MPS VI in India and to study the genotype-phenotype association and functional outcomes of these mutations. Methods: Molecular characterization of the ARSB gene by Sanger sequencing was done for 15 patients (aged 15 months to 11 yr) who were enzymatically confirmed to have MPS VI. Age of onset, clinical progression and enzyme activity levels in each patient were studied to look for genotype-phenotype association. Haplotype analysis performed for unrelated patients with the recurring mutation W450C, was suggestive of a founder effect. Sequence and structural analyses of the ARSB protein using standard software were carried out to determine the impact of detected mutations on the function of the ARSB protein. Results: A total of 12 mutations were identified, of which nine were novel mutations namely, p.D53N, p.L98R, p.Y103SfsX9, p.W353X, p.H393R, p.F166fsX18, p.I220fsX5, p.W450L, and p.W450C, and three were known mutations (p.D54N, p.A237D and p.S320R). The nine novel sequence variants were confirmed not to be polymorphic variants by performing sequencing in 50 unaffected individuals from the same ethnic population. Interpretation & conclusions: Nine novel mutations were identified in MPS VI cases from India in the present study. The study also provides some insights into the genotype-phenotype association in MPS VI.

Novel mutations in PRG4 gene in two Indian families with camptodactyly-arthropathy- coxa vara- pericarditis (CACP) syndrome.

Background & objectives: Camptodactyly – arthropathy- coxa vara- pericarditis (CACP) syndrome is an autosomal recessive disorder caused by mutations in the PRG4 (proteoglycan 4) gene. Hallmarks of the syndrome include congenital or early-onset camptodactyly and arthropathy with synovial hyperplasia, progressive coxa vara deformity and non-inflammatory pericardial effusions. Till date only around 25 pathogenic mutations have been reported in this gene and none have been reported from India. We report here the mutations in the PRG4 gene in three patients of CACP from two unrelated families from India. Methods: Molecular genetic studies were done for the three patients with the CACP syndrome, from two unrelated Indian families, through sequence analysis of all coding exons and the exon-intron boundaries of the PRG4 gene. Results: Two novel frame-shift deletion mutations leading to premature protein termination were found. One patient was identified to be homozygous for a 2 base pair deletion in exon 6 (c.2645_2646delGA) and the two affected siblings from the other family were found to be homozygous for a 4 base pair deletion in exon 6 (c.2883_2886delAAGA). Conclusions: This is perhaps the first report of PRG4 mutations from India. Further mutation studies in Indian CACP cases will help to determine the mutation spectrum of the PRG4 gene in the Indian population and also help to further elucidate the molecular pathology and the genotype-phenotype correlation of this rare disease.

Williams Syndrome: A Case Series.

Pediatricians’ awareness about malformation syndromes can help in their timely diagnosis. Williams syndrome is a microdeletion syndrome associated with characteristic facial features and behavioral phenotype. Diagnosis can be confirmed by fluorescence-in-situ hybridization or multiplex ligation probe amplification. Correct diagnosis can help in diagnosing hypercalcemia and cardiac defects, and providing genetic counseling to the family.

Clinical utility of multiplex ligation-dependent probe amplification technique in identification of aetiology of unexplained mental retardation: A study in 203 Indian patients.

Background & objectives: Developmental delay (DD)/mental retardation also described as intellectual disability (ID), is seen in 1-3 per cent of general population. Diagnosis continues to be a challenge at clinical level. With the advancement of new molecular cytogenetic techniques such as cytogenetic microarray (CMA), multiplex ligation-dependent probe amplification (MLPA) techniques, many microdeletion/microduplication syndromes with DD/ID are now delineated. MLPA technique can probe 40-50 genomic regions in a single reaction and is being used for evaluation of cases with DD/ID. In this study we evaluated the clinical utility of MLPA techniques with different probe sets to identify the aetiology of unexplained mental retardation in patients with ID/DD. Methods: A total of 203 randomly selected DD/ID cases with/without malformations were studied. MLPA probe sets for subtelomeric regions (P070/P036) and common microdeletions/microduplications (P245-A2) and X-chromosome (P106) were used. Positive cases with MLPA technique were confirmed using either fl uorescence in situ hybridization (FISH) or follow up confirmatory MLPA probe sets. Results: The overall detection rate was found to be 9.3 per cent (19 out of 203). The detection rates were 6.9 and 7.4 per cent for common microdeletion/microduplication and subtelomeric probe sets, respectively. No abnormality was detected with probe set for X-linked ID. The subtelomeric abnormalities detected included deletions of 1p36.33, 4p, 5p, 9p, 9q, 13q telomeric regions and duplication of 9pter. The deletions/duplications detected in non telomeric regions include regions for Prader Willi/Angelman regions, Williams syndrome, Smith Magenis syndrome and Velocardiofacial syndrome. Interpretation & conclusions: Our results show that the use of P245-A2 and P070/P036-E1 probes gives good diagnostic yield. Though MLPA cannot probe the whole genome like cytogenetic microarray, due to its ease and relative low cost it is an important technique for evaluation of cases with DD/ID.