Precision diagnosis, personalized management and future directions of 21-hydroxylase deficiency

Genetic testing for 21-hydroxylase deficiency (21-OHD) is always challenging. The current approaches of short-read sequencing and multiplex ligation-dependent probe amplification (MLPA) are insufficient for the detection of chimeric genes or complicated variants from multiple copies. Recently developed long-read sequencing (LRS) can solve this problem. To investigate the clinical utility of LRS in precision diagnosis of 21-OHD. In the cohort of 832 patients with 21-OHD, the current approaches provided the precise molecular diagnosis for 81.7% (680/832) of cases. LRS was performed to solve the remaining 144 cases with complex chimeric variants and 8 cases with variants from multiple copies. Clinical manifestations in patients with continuous deletions of CYP21A2 extending to TNXB (namely CAH-X) were further evaluated. Using LRS in combination with previous genetic test results, a total of 16.9% (281/1664) CYP21A1P/CYP21A2 or TNXA/TNXB chimeric alleles were identified in 832 patients, with CYP21A1P/CYP21A2 accounting for 10.4% and TNXA/TNXB for 6.5%. The top 3 common chimeras were CYP21 CH-1, TNX CH-1, and TNX CH-2, accounting for 77.2% (217/281) of all chimeric alleles. The 8 patients with variants on multiple copies of CYP21A2 were accurately identified with LRS. The prevalence of CAH-X in our cohort was 12.1%, and a high frequency of connective tissue-related symptoms was observed in CAH-X patients. LRS can detect all types of CYP21A2 variants, including complex chimeras and pathogenic variants on multiple copies in patients with 21-OHD, which could be utilized as a first-tier routine test for the precision diagnosis and categorization of congenital adrenal hyperplasia.

Serum vitamin B 12 and methylmalonic acid determinations in the diagnosis of cobalt deficiency in pregnant ewes

Congenital adrenal hyperplasia

Antithrombin (AT), protein C (PC), and protein S (PS) are natural anticoagulant proteins that deficiency in each of them is associated with an increased risk of venous thromboembolism.The overlapping of plasma levels of AT, PC, and PS between healthy individuals and heterozygote carriers poses significant challenges in precise diagnosis. This study aimed to evaluate the effect of most influencing variables on plasma levels of these proteins and propose specific reference intervals to improve the interpretation of the laboratory results. This study was conducted on 1464 individuals who were referred to Massoud medical laboratory, Tehran, Iran, from 2019 to 2020. AT and PC were measured through chromogenic assay and PS plasma level with the clot-based assay. A multivariable linear regression model was performed to evaluate the effect of sex, age, oral contraceptive (OCP) intake, and menopause state. Normal deviate z value was used for different subgroups to justify the need for a separate reference interval. 1200 verified healthy individuals (434males and 766 females), aged between 18 and 69years were included in the study. The mean±SD age of the participants was 39.78±11.79years. The age-related effects for AT were found in men. In females, increasing age was associated with a rise in AT, PC, and PS plasma levels. No sex difference was found in AT plasma level. OCP-taking is associated with a decrease in AT and an increase in PC plasma levels. This is the largest study ever conducted on healthy individuals in the Iranian population, using specific reference interval results in accurate diagnosis of true AT, PC, and PS deficiency.

Dear Sir, We read with great interest an article by Scarpa et al.1 on a patient with functional vitamin B12 deficiency despite serum levels of the vitamin being in the reference interval. We congratulate the authors for diagnosing the patient with their precise biochemical and clinical assessments and for their valuable comments. We would like to add a short note to help readers to interpret the results of the study. The patient was diagnosed as having megaloblastic anaemia despite serum vitamin B12 and folic acid levels being within normative ranges, at 910 pg/mL and 10.3 ng/mL, respectively. One of the possible explanations for this condition is low tissue levels or functional deficiencies of both vitamin B12 and folic acid. Laboratory errors are much less frequent explanations in well-managed laboratories. The analysis of plasma total homocysteine (tHcy) has been recommended in the diagnosis of folate and cobalamin deficiencies, because these vitamins are both cofactors of the methionine synthase enzyme which has central role in one carbon metabolism2. Elevated tHcy concentrations could demonstrate a functional deficiency of folic acid, directly. Additionally, vitamin B12 metabolism could be evaluated through the concentration of methylmalonic acid (MMA) in either the serum or urine, which is accepted as a better approach for the assessment of functional or tissue levels of vitamin B12 deficiency3. Indeed, as in the case described by Scarpa et al., normal serum levels of vitamin B12 do not exclude low tissue levels or a functional deficiency of the vitamin. MMA measurement could, therefore, be a complementary way to evaluate intracellular vitamin B12 metabolism, although there are some problems relative to the specificity, complexity and cost of this test. In conclusion, patients considered to have megaloblastic anemia could be further evaluated by measuring their tHcy and MMA concentrations which would help clinicians to make a precise diagnosis and exclude possible laboratory errors. The case report presented highlights the importance of careful interpretation of clinical laboratory results.

Diagnosis of 5α-Reductase Deficiency in a Teenage Turkish Girl

BackgroundAcid sphingomyelinase deficiency (ASMD) disorder, also known as Niemann–Pick disease (NPD) is a rare genetic disease caused by mutations in SMPD1 gene, which encodes sphingomyelin phosphodiesterase (ASM). Except for liver and spleen enlargement and lung disease, two subtypes (Type A and B) of NDP have different onset times, survival times, ASM activities, and neurological abnormalities. To comprehensively explore NPD’s genotype-phenotype association and pathophysiological characteristics, we collected 144 NPD cases with strict quality control through literature mining.ResultsThe difference in ASM activity can differentiate NPD type A from other subtypes, with the ratio of ASM activity to the reference values being lower in type A (threshold 0.045 (4.45%)). Severe variations, such as deletion and insertion, can cause complete loss of ASM function, leading to type A, whereas relatively mild missense mutations generally result in type B. Among reported mutations, the p.Arg3AlafsX76 mutation is highly prevalent in the Chinese population, and the p.R608del mutation is common in Mediterranean countries. The expression profiles of SMPD1 from GTEx and single-cell RNA sequencing data of multiple fetal tissues showed that high expressions of SMPD1 can be observed in the liver, spleen, and brain tissues of adults and hepatoblasts, hematopoietic stem cells, STC2_TLX1-positive cells, mesothelial cells of the spleen, vascular endothelial cells of the cerebellum and the cerebrum of fetuses, indicating that SMPD1 dysfunction is highly likely to have a significant effect on the function of those cell types during development and the clinicians need pay attention to these organs or tissues as well during diagnosis. In addition, we also predicted 21 new pathogenic mutations in the SMPD1 gene that potentially cause the NPD, signifying that more rare cases will be detected with those mutations in SMPD1. Finally, we also analysed the function of the NPD type A cells following the extracellular milieu.ConclusionsOur study is the first to elucidate the effects of SMPD1 mutation on cell types and at the tissue level, which provides new insights into the genotype-phenotype association and can help in the precise diagnosis of NPD.

Comparative Study of Limbal Stem Cell Deficiency Diagnosis Methods: Detection of MUC5AC mRNA and Goblet Cells in Corneal Epithelium

Challenges in implementation of ISTH diagnostic algorithm for diagnosis and classification of factor XIII deficiency in Iran

DEXAMETHASONE TREATMENT OF CONGENITAL ADRENAL HYPERPLASIA IN UTERO: AN EXPERIMENTAL THERAPY OF UNPROVEN SAFETY

Neonatal encephalopathy with seizures is a presentation in which rapid whole-genome sequencing (rWGS) has shown clinical utility and improved outcomes. We report a neonate who presented on the third day of life with seizures refractory to antiepileptic medications and neurologic and computerized tomographic findings consistent with severe generalized brain swelling. rWGS revealed compound heterozygous variants in the molybdenum cofactor synthesis gene, type 1A (MOCS1 c.*7 + 5G > A and c.377G > A); a provisional diagnosis of molybdenum cofactor deficiency on day of life 4. An emergency investigational new drug application for intravenous replacement of the MOCS1 product, cyclic pyranopterin monophosphate, was considered, but felt unsuitable in light of the severity of disease and delay in the start of treatment. The patient died on day of life 9 despite having a precise molecular diagnosis within the first week of life. This case illustrates that an rWGS-based molecular diagnosis within the first week of life may be insufficient to improve outcomes. However, it did inform clinical decision-making with regard to resuscitation and predicted long-term outcome. We suggest that to achieve optimal reductions in morbidity and mortality, rWGS must be implemented within a comprehensive rapid precision medicine system (CRPM). Akin to newborn screening (NBS), CRPM will have onboarding, diagnosis, and precision medicine implementation components developed in response to patient and parental needs. Education of health-care providers in a learning model in which ongoing data analyses informs system improvement will be essential for optimal effectiveness of CRPM.

We aimed to develop policy in relation to three areas: (i) the diagnosis of iron deficiency; (ii) maternal-infant issues and the prevention of iron deficiency; and (iii) the treatment of iron deficiency. Within each of these topic areas we completed a literature review and developed recommendations to help direct activities of the Royal Australasian College of Physicians, update paediatricians and guide clinical practice. Iron deficiency can be defined using cut-off values for laboratory measures of iron status or, if an intercurrent infection is not present, by demonstrating a response to a therapeutic trial of iron. The appropriate measures of iron status vary depending upon the presence of intercurrent infection. Full-term babies are born with iron stores sufficient to meet their needs to age 4-6 months but premature infants are not. After age 6 months infants are dependent upon dietary iron from complementary foods even with continued breastfeeding. Infants <33 weeks gestation or <1800 g birthweight should receive iron from 4 weeks of age. In most settings recommended treatment of iron deficiency is with oral ferrous sulphate as a single or twice daily dose of between 3 and 6 mg/kg/day. Iron deficiency is prevalent and an important determinant of child health. Precise and accurate diagnosis remains challenging. Iron supplementation is required for premature and low-birthweight infants. Oral iron salts remain the recommended treatment of choice in most instances.

Hearing deficiency is the world’s most common sensation of impairment and impedes human communication and learning. Early and precise hearing diagnosis using electroencephalogram (EEG) is referred to as the optimum strategy to deal with this issue. Among a wide range of EEG control signals, the most relevant modality for hearing loss diagnosis is auditory evoked potential (AEP) which is produced in the brain’s cortex area through an auditory stimulus. This study aims to develop a robust intelligent auditory sensation system utilizing a pre-train deep learning framework by analyzing and evaluating the functional reliability of the hearing based on the AEP response. First, the raw AEP data is transformed into time-frequency images through the wavelet transformation. Then, lower-level functionality is eliminated using a pre-trained network. Here, an improved-VGG16 architecture has been designed based on removing some convolutional layers and adding new layers in the fully connected block. Subsequently, the higher levels of the neural network architecture are fine-tuned using the labelled time-frequency images. Finally, the proposed method’s performance has been validated by a reputed publicly available AEP dataset, recorded from sixteen subjects when they have heard specific auditory stimuli in the left or right ear. The proposed method outperforms the state-of-art studies by improving the classification accuracy to 96.87% (from 57.375%), which indicates that the proposed improved-VGG16 architecture can significantly deal with AEP response in early hearing loss diagnosis.

An Overview of Molecular Diagnosis of Steroid 21-Hydroxylase Deficiency

Problems in diagnosis and management of congenital adrenal hyperplasia due to 21-hydroxylase deficiency