Relevance of augmented kisspeptin signaling through H364 KISS1R in central precocious puberty.

Understanding the pathophysiology of idiopathic central precocious puberty (ICPP) is essential, in view of its consequences on reproductive health and metabolic disorders in later life. Towards this, estimation of circulating levels of the neuropeptides, viz; Kisspeptin (Kp-10), Neurokinin B (NKB) and Neuropeptide Y (NPY), acting upstream to Gonadotropin-Releasing Hormone (GnRH), has shown promise. Insights can also be gained from functional studies on genetic variations implicated in ICPP. This study investigated the pathophysiology of ICPP in a girl by exploring the therapeutic relevance of the circulating levels of Kp-10, NKB, NPY and characterizing the nonsynonymous KISS1R variant, L364H, that she harbours, in a homozygous condition. Plasma levels of Kp-10, NKB and NPY before and after GnRH analog (GnRHa) treatment, were determined by ELISA. It was observed that GnRHa treatment resulted in suppression of circulating levels of Kp-10, NKB and NPY. Further, the H364 variant in KISS1R was generated by site directed mutagenesis. Post transient transfection of either L364 or H364 KISS1R variant in CHO cells, receptor expression was ascertained by western blotting, indirect immunofluorescence and flow cytometry. Kp-10 stimulated signalling response was also determined by phospho-ERK and inositol phosphate production. Structure-function studies revealed that, although the receptor expression in H364 KISS1R was comparable to L364 KISS1R, there was an enhanced signalling response through this variant at high doses of Kp-10. Thus, elevated levels of Kp-10, acting through H364 KISS1R, contributed to the manifestation of ICPP, providing further evidence that dysregulation of Kp-10/KISS1R axis impacts the onset of puberty.

The dynamics of SARS-CoV-2 infection in unvaccinated and vaccinated populations in Mumbai, India, between 28 December 2020 and 30 August 2021.

The emergence and evolution of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) variants that could compromise vaccine efficacy (VE) with re-infections in immunized individuals have necessitated continuous surveillance of VE. Here, the occurrence and dynamics of SARS-CoV-2 infections in the context of vaccination during the second wave of infection in Mumbai were evaluated. RT-PCR cycle threshold (Ct) values of the open reading frame (ORF)/envelope (E)/nucleocapsid (N) genes obtained from a total of 42415 samples, comprising unvaccinated (96.88%) and vaccinated cases (3.12%) were analyzed between December 28, 2020, and August 30, 2021. A lower incidence of SARS-CoV-2 infection in fully vaccinated cases (5.07%) compared to partially vaccinated cases (6.5%) and unvaccinated cases (13.453%) was recorded. VE was significant after the first dose of vaccination (ORF gene p-value = 0.003429, and E/N gene p-value = 0.000866). Furthermore, VE was observed to be significant when the post-immunization (first dose) period was stratified to within 30 days (ORF gene p-value = 0.0094 and E/N gene p-value = 0.0023) and to 60 days following the second dose of vaccination (ORF gene p-value = 0.0238). Also, significantly higher efficacy was observed within individuals receiving two doses compared to a single dose (ORF gene p-value = 0.0132 and E/N gene p-value = 0.0387). The emergence of breakthrough infections was also evident (odds ratio= 0.34; 95% confidence interval= 0.27-0.43). Interestingly, viral loads trended towards being higher in some groups of partially vaccinated individuals compared to completely vaccinated and unvaccinated populations. Finally, our results delineated a significantly higher incidence of SARS-CoV-2 acquisition in males, asymptomatic individuals, individuals with comorbidities, and those who were unvaccinated.

Cytomegalovirus reactivation in a SARS-CoV-2 infected woman experiencing fetal demise in the first trimester with fetal trisomy 21: A case report.

Cytomegalovirus (CMV) is the most common cause of congenital viral infections. Women seropositive for CMV prior to pregnancy can develop a non-primary CMV infection. Here, we present a case of first trimester pregnancy loss during active SARS-CoV-2 infection. There was no evidence of SARS-CoV-2 RNA in placenta and fetal tissue, but there was presence of congenital cytomegalovirus infection by nested PCR. To the best of our knowledge, this is the first report demonstrating association of early congenital CMV infection due to reactivation and fetal demise in a SARS-CoV-2 positive woman with fetal trisomy 21.

Evaluation of Single Exon Deletions in DMD/BMD: Technical and Analytical Concerns.

Background: Oftentimes, a variation at the multiplex ligation-dependent probe amplification (MLPA) probe binding site leads to improper hybridrization/ligation of the probe showing up as a deletion of an exon leading to false positive results for the detection of Duchenne muscular dystrophy (DMD)/Becker muscular dystrophy (BMD). Objective: Investigating cases with single exon deletion using an alternate method [polymerase chain reaction (PCR) or sequencing] for confirmation of the deletion. Methods: We evaluated males with single exon deletion (n = 49) in our study population (2015-2019). Forty-six were confirmed by an alternate method (conventional PCR/Sanger's sequencing) to confirm the deletion. Results: We observed 25.12% single exon deletions in our study cohort. Further evaluation determined a false positive rate of 6.12%. Three out of 49 single exon deletions had a point mutation near the probe-binding site, indicating a false positive result. Single exon deletions, thus, need to be evaluated with extreme caution, and point mutations, if any, need to be characterized to determine the nature of their pathogenicity.

Transformation to telephonic genetic counselling during SARS-CoV-2 pandemic: challenges and suggested protocol in Indian scenario.

Genetic counselling (GC) is a process of communicating and educating patients and/or their family members diagnosed with genetic abnormalities. Ideally, GC is offered in-person, physical presence of both the counselee and the counsellor. However, COVID-19 pandemic and new norms of social distancing precluded undertaking GCs. In the wake of this, Genetic Research Centre at ICMR-NIRRH, Mumbai, arranged virtual sessions for GC. Here, we describe our experience of initiating genetic counselling services on virtual platform. This report presents the challenges faced by the genetic counsellors as well as the counselees and suggests a protocol to be followed during tele-genetic counselling. It is based on the retrospective data of 65 cases that were counselled from July 2020 to September 2020 which was the period of lockdown and restriction. Although a few issues emerged during the process of GC, virtual tele-counselling was a preferred option due to social distancing, lack of public transport facilities and COVID-19 specific restrictions. Effective virtual follow ups saved time, energy and finances of providers as well as clients. This article presents providers' experience of the process and some recommendations in Indian scenario.

Detection of SARS-CoV-2 infection by RT-PCR test: factors influencing interpretation of results.

In this current pandemic of coronavirus disease 2019 (COVID-19), prompt interventions in terms of early detection and clinical management along with isolation of positive cases is of utmost importance. This helps to limit not only the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections but also the morbidity and mortality associated with it. Different strategies for screening of COVID-19 in containment zones and non-containment areas include testing of symptomatic patients and their contacts in fever clinics, hospital-based testing, testing on demand and population-based screening. The choice of tests like reverse-transcription polymerase chain reaction (RT-PCR), rapid antigen testing (RAT) or antibody test depends upon these strategies and also the turnaround time. Currently, RT-PCR is considered the gold standard for COVID-19 detection. This commentary provides the insights and experiences on COVID-19 diagnosis by RT-PCR. The utility of this test is limited by several false positive, false negative and inconclusive results at early stages of infection, scarcity of reagents and lack of well-equipped labs including trained staff. Moreover, appropriate sample collection and transport, standard laboratory protocols, stringent quality control norms, good quality RNA extraction kits, PCR kits with suitable primers can help in improving accuracy of the test results. A careful assessment of clinical, radiological and molecular findings is required for identifying potential cases of COVID-19.

A retrospective analysis of COVID-19 diagnosis results obtained by rapid antigen tests and RT-PCR: Implications for disease management.

There is a need for understanding and establishment of the most appropriate testing algorithm for COVID-19 diagnosis in asymptomatic high-risk groups. Here, we present a retrospective analysis of RT-PCR results obtained from 412 cases tested negative for coronavirus disease 2019 (COVID-19) by rapid antigen testing method. Among 178 (43.2%) asymptomatic individuals, 44.9% of the high risk contacts, 12.2% of police custody individuals, 22.22% of the pregnant women and 33.33% of individuals hospitalised for preoperative or other medical conditions showed RT-PCR positivity. Our results suggest a need for focussed and intensive (multi-modality) testing in groups at high risk for SARS-CoV-2 infection.

A complex karyotype with t(11;12)(q23;p13) translocation with coexistent clones of deletion 5q and cryptic deletion 7q in acute myeloid leukemia.

We report a rare case of acute myeloid leukemia (AML) with a cytogenetically complex karyotype with coexistence of KMT2A/MLL Mixed Lineage Leukemia (11q23) rearrangement with 5q deletion and 7q deletion as unrelated clones along with evolution of a subclone with translocation between chromosomes 6 and 17. A novel MLL fusion partner region 12p13 was identified in a 52 year old woman who presented with pyrexia of unknown origin. Unraveling the complexity of genomic alterations occurring in AML patients will lead to better understanding of leukemic transformation and identification of subsets of patients that may respond differently to therapy.

A novel CFTR gene variant - p.Tyr517* associated with cystic fibrosis: a case report.

INTRODUCTION: Cystic fibrosis (CF) is a genetic disease usually diagnosed by clinical findings and abnormal sweat chloride testing. CASE REPORT: We report a case of an 18-month-old Indian female with clinical findings suggestive of CF referred for genetic confirmation. The CFTR gene was sequenced for 23 mutations as per American College of Medical Genetics (ACMG) guidelines for CF and showed presence of a known common heterozygous delF508 (c.1521_1523delCTT, p.Phe508 del) variant. In addition to delF508 variant, exon 10 of CFTR gene also showed a novel variant c.1551C > G, p.Tyr517*, which was classified as "likely pathogenic" based on recent ACMG variant classification guidelines. The presence of compound heterozygous pathogenic variants along with classical clinical findings, confirmed the diagnosis of CF in this patient. CONCLUSION: The novel pathogenic variants (missense/nonsense/deletion/duplication) in CFTR gene are often identified and are associated with CF, thus highlighting the need of comprehensive complete CFTR gene analysis.

A Study of Triplet-Primed PCR for Identification of CAG Repeat Expansion in the HTT Gene in a Cohort of 503 Indian Cases with Huntington's Disease Symptoms.

BACKGROUND: Huntington's disease (HD) is an autosomal-dominant neurodegenerative disorder with an average age at onset of 40 years. It is a polyglutamine (polyQ) disorder that is caused by an increase in the number of CAG repeats in the huntingtin (HTT) gene. Genetic tests that accurately determine the number of CAG repeats are performed for confirmation of diagnosis, predictive testing of persons at genetic risk for inheriting HD, and prenatal testing. The aim of our study was to evaluate efficacy of triplet-primed polymerase chain reaction (TP-PCR) for routine diagnosis of HD in suspected cases from India. METHODS: We evaluated a combination of CAG flanking PCR and triplet-primed PCR for estimation of CAG repeats in 503 cases with clinical suspicion of HD. RESULTS: There were 250 cases (49.7%) that showed the presence of expanded alleles, with 241 (47.9%) being fully penetrant alleles and nine (1.8%) in the reduced penetrance category. There were seven juvenile cases with an age of onset of < 20 years, with the longest allele comprising 106 CAG repeats found in an 8-year-old male patient. The results demonstrated an inverse (R = - 0.67) relationship between CAG length and age at clinical onset. CONCLUSION: Our study on pan-Indian cases is one of the largest studies reported so far in India and focuses on the most accurate and comprehensive molecular diagnostic evaluation of HD.