IndiGenomes: a comprehensive resource of genetic variants from over 1000 Indian genomes.

With the advent of next-generation sequencing, large-scale initiatives for mining whole genomes and exomes have been employed to better understand global or population-level genetic architecture. India encompasses more than 17% of the world population with extensive genetic diversity, but is under-represented in the global sequencing datasets. This gave us the impetus to perform and analyze the whole genome sequencing of 1029 healthy Indian individuals under the pilot phase of the 'IndiGen' program. We generated a compendium of 55,898,122 single allelic genetic variants from geographically distinct Indian genomes and calculated the allele frequency, allele count, allele number, along with the number of heterozygous or homozygous individuals. In the present study, these variants were systematically annotated using publicly available population databases and can be accessed through a browsable online database named as 'IndiGenomes' http://clingen.igib.res.in/indigen/. The IndiGenomes database will help clinicians and researchers in exploring the genetic component underlying medical conditions. Till date, this is the most comprehensive genetic variant resource for the Indian population and is made freely available for academic utility. The resource has also been accessed extensively by the worldwide community since it's launch.

GnIH and GnRH expressions in the central nervous system and pituitary of Indian major carp, Labeo rohita during ontogeny: An immunocytochemical study.

Gonadotropin-releasing hormone (GnRH) is the major hypothalamic neuropeptide stimulating gonadotropin secretion in vertebrates. In 2000, gonadotropin-inhibitory hormone (GnIH) was discovered as a hypothalamic neuropeptide that inhibits gonadotropin secretion in birds. Subsequent studies have shown that GnIH is present in the brain of other vertebrates. We show for the first time GnIH immunoreactivity in the central nervous system and pituitary during development of Indian major carp, Labeo rohita and compare it with the localization of GnRH. GnIH and GnRH immunoreactivities were observed from the olfactory system to spinal cord throughout development. In the brain, both neuropeptides were localized in the telencephalon, diencephalon including the preoptic area and rhombencephalon. The localization of GnIH and GnRH in the pituitary suggests that these neuropeptides are involved in the regulation of pituitary hormones by an autocrine manner during development. In addition, the presence of GnIH and GnRH in several other brain regions including the olfactory system suggests their involvement in the regulation of other physiological functions.

Role of catecholamines and nitric oxide on pigment displacement of the chromatophores of freshwater snakehead teleost fish, Channa punctatus.

We are reporting for the first time that the catecholamines (adrenaline and noradrenaline) inhibit the effect of nitric oxide (NO) on melanosome dispersion in freshly isolated scales of the freshwater snakehead fish, Channa punctatus. We studied the effect of NO and catecholamines on the pigment displacement by observing the changes in the melanophore index. The scales when treated with solution containing NO donor sodium nitroprusside (SNP) showed dispersion of melanosomes, whereas NO synthase blocker N-omega-Nitro-L-arginine suppresses this action of SNP. Treatment with adrenaline and noradrenaline on the isolated scales caused aggregation of melanosomes. Scales treated with solution containing catecholamines and SNP resulted in aggregation of melanosomes suggesting that catecholamines mask the effect of SNP. These results suggest that the catecholamines are inhibiting the effect of NO and causing the aggregation of the melanosomes may be via surface receptors.

Nitric oxide inhibited the melanophore aggregation induced by extracellular calcium concentration in snakehead fish, Channa punctatus.

We studied the role of nitric oxide (NO) and extra-cellular Ca(2+) on the melanophores in Indian snakehead teleost, Channa punctatus. Increase of Ca(2+) level in the external medium causes pigment aggregation in melanophores. This pigment-aggregating effect was found to be inhibited when the external medium contained spontaneous NO donor, sodium nitro prusside (SNP) at all the levels of concentration tested. Furthermore, it has been observed that SNP keeps the pigment in dispersed state even after increasing the amount of Ca(2+). In order to test whether NO donor SNP causes dispersion of pigments or not is checked by adding the inhibitor of nitric oxide synthase, N-omega-Nitro-L-arginine (L-NNA) in the medium. It has been noted that the inhibitor L-NNA blocked the effect of NO donor SNP causing aggregation of pigments. In that way NO is inhibiting the effect of extracellular Ca(2+), keeping the pigment dispersed.

Pharmacogenomic landscape of COVID-19 therapies from Indian population genomes.

Aim: Numerous drugs are being widely prescribed for COVID-19 treatment without any direct evidence for the drug safety/efficacy in patients across diverse ethnic populations. Materials & methods: We analyzed whole genomes of 1029 Indian individuals (IndiGen) to understand the extent of drug-gene (pharmacogenetic), drug-drug and drug-drug-gene interactions associated with COVID-19 therapy in the Indian population. Results: We identified 30 clinically significant pharmacogenetic variants and 73 predicted deleterious pharmacogenetic variants. COVID-19-associated pharmacogenes were substantially overlapped with those of metabolic disorder therapeutics. CYP3A4, ABCB1 and ALB are the most shared pharmacogenes. Fifteen COVID-19 therapeutics were predicted as likely drug-drug interaction candidates when used with four CYP inhibitor drugs. Conclusion: Our findings provide actionable insights for future validation studies and improved clinical decisions for COVID-19 therapy in Indians.

Genetic epidemiology of autoinflammatory disease variants in Indian population from 1029 whole genomes.

BACKGROUND: Autoinflammatory disorders are the group of inherited inflammatory disorders caused due to the genetic defect in the genes that regulates innate immune systems. These have been clinically characterized based on the duration and occurrence of unprovoked fever, skin rash, and patient's ancestry. There are several autoinflammatory disorders that are found to be prevalent in a specific population and whose disease genetic epidemiology within the population has been well understood. However, India has a limited number of genetic studies reported for autoinflammatory disorders till date. The whole genome sequencing and analysis of 1029 Indian individuals performed under the IndiGen project persuaded us to perform the genetic epidemiology of the autoinflammatory disorders in India. RESULTS: We have systematically annotated the genetic variants of 56 genes implicated in autoinflammatory disorder. These genetic variants were reclassified into five categories (i.e., pathogenic, likely pathogenic, benign, likely benign, and variant of uncertain significance (VUS)) according to the American College of Medical Genetics and Association of Molecular pathology (ACMG-AMP) guidelines. Our analysis revealed 20 pathogenic and likely pathogenic variants with significant differences in the allele frequency compared with the global population. We also found six causal founder variants in the IndiGen dataset belonging to different ancestry. We have performed haplotype prediction analysis for founder mutations haplotype that reveals the admixture of the South Asian population with other populations. The cumulative carrier frequency of the autoinflammatory disorder in India was found to be 3.5% which is much higher than reported. CONCLUSION: With such frequency in the Indian population, there is a great need for awareness among clinicians as well as the general public regarding the autoinflammatory disorder. To the best of our knowledge, this is the first and most comprehensive population scale genetic epidemiological study being reported from India.

Neuroanatomical demonstration of calbindin 2a- and calbindin 2b-like calcium binding proteins in the early embryonic development of zebrafish: mRNA study.

Certain calcium binding proteins (CaBPs) are essential for metabolic processes but the role of these proteins in the development is not well known. We have investigated the mRNA expression of CaBPs, calbindin 2a (Calb2a) and calbindin 2b (Calb2b) in the zebrafish embryos 24, 36, 48 and 72h post fertilization (hpf). We have seen very high Calb2a mRNA expression in the tegmentum (Tg), midbrain-hindbrain boundary (Mhb), hindbrain (Hb), spinal cord (Sc), retina and cranial ganglion (Crg). Also very high Calb2b mRNA expression was noted in olfactory cells, cerebellum, Tg, Mhb, Hb, optic tectum, retina, retinal ganglion cell layer, retinal inner nuclear layer, Sc, Neural crest, infraorbital neuromasts, pharyngeal arch 3-7 skeleton and mandibular neuromasts. It is known that many factors are involved in the differentiation of Mhb. Here we are reporting for the first time the mRNA expression of CaBPs (Calb2a and Calb2b) in the Mhb indicating their role in the differentiation of Mhb and development of the brain, eyes and other tissues in the zebrafish. We suggest that Calb2a and Calb2b play an important role in the regulation of zebrafish early embryonic development.

Short-term exposure to L-type calcium channel blocker, verapamil, alters the expression pattern of calcium-binding proteins in the brain of goldfish, Carassius auratus.

The influx of calcium ions (Ca(2+)) is responsible for various physiological events including neurotransmitter release and synaptic modulation. The L-type voltage dependent calcium channels (L-type VDCCs) transport Ca(2+) across the membrane. Calcium-binding proteins (CaBPs) bind free cytosolic Ca(2+) and prevent excitotoxicity caused by sudden increase in cytoplasmic Ca(2+). The present study was aimed to understand the regulation of expression of neuronal CaBPs, namely, calretinin (CR) and parvalbumin (PV) following blockade of L-type VDCCs in the CNS of Carassius auratus. Verapamil (VRP), a potent L-type VDCC blocker, selectively blocks Ca(2+) entry at the plasma membrane level. VRP present in the aquatic environment at a very low residual concentration has shown ecotoxicological effects on aquatic animals. Following acute exposure for 96h, median lethal concentration (LC50) for VRP was found to be 1.22mg/L for goldfish. At various doses of VRP, the behavioral alterations were observed in the form of respiratory difficulty and loss of body balance confirming the cardiovascular toxicity caused by VRP at higher doses. In addition to affecting the cardiovascular system, VRP also showed effects on the nervous system in the form of altered expression of PV. When compared with controls, the pattern of CR expression did not show any variations, while PV expression showed significant alterations in few neuronal populations such as the pretectal nucleus, inferior lobes, and the rostral corpus cerebellum. Our result suggests possible regulatory effect of calcium channel blockers on the expression of PV.

Neuroanatomical localization of nitric oxide synthase (nNOS) in the central nervous system of carp, Labeo rohita during post-embryonic development.

Nitric oxide (NO) is a chemically diffusible molecular messenger playing various roles in both vertebrates and invertebrates. Nitric oxide synthase (NOS) is the key enzyme in synthesis of NO. The neuroanatomical distribution pattern of neuronal nitric oxide synthase (nNOS) was studied and developing stages of Labeo rohita such as hatchlings (10-15mm), frys (15-35mm), semi-fingerlings (35-65mm), fingerlings (65-100mm) and adults (350-370mm) were used. In the telencephalon, nitrergic cells were observed in both pallial and subpallial regions along with entopeduncular nucleus suggesting the involvement of NO in the control of sensory functions throughout the development. In the diencephalon, nNOS positive neurons were localized in the nucleus preopticus periventricularis and preopticus parvocellularis throughout development while nucleus preopticus magnocellularis was found immunopositive only in adult specimens who suggest the involvement of NO in the hormonal regulation. nNOS immunoreaction was also noted in suprachaismatic nucleus, habenula, lateral tuberal nucleus, paraventricular organ and anterior division of preglomerular nucleus throughout development. In the mesencephalic region, nNOS immunoreactivity was seen in the optic tectum, torus longitudinalis, nucleus of median longitudinal fascicle and occulomotor nucleus indicate the role of NO in integration of visual inputs and modulates motor control of the eyes and movements. Caudally, in the rhombencephalon, the cerebellum, the nucleus reticularis, the octaval nucleus and the motor nucleus of vagal nerve were nNOS positive during development. nNOS reactive cells and fibers were noted in the spinal motor column, thus suggesting a role of NO in gestation and startle response from early development.

The ontogeny of Mauthner cells in the brain of Labeo rohita as revealed by NADPH-d and nNOS immunohistochemistry.

Nitric oxide (NO) is well demonstrated to act as a neuronal messenger in neurotransmission in vertebrate animals. We are for the first time reporting nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and neuronal nitric oxide synthase (nNOS) in the Mauthner cells (MC) of the carp, Labeo rohita brain during post-embryonic development. The NADPH-d and nNOS-positive MC were seen intensely labeled in all young stages on either side of the ventricle in the hindbrain. In adult, these cells were moderately stained with NADPH-d but not with nNOS. Further, the morphometeric analysis of the staining intensities showed the decrease in the optical densities of the MC as the development progresses. The gradual increase in cell diameter was evident from hatchling to adult (18-22 µm to 98-106 µm), while the nuclear diameter remained nearly unchanged (10-12 µm to 18-20 µm). In fishes, MC are known to be involved in the control of startle response. The NADPH-d enzymatic activity seen in the MC of early development may play an important role to activate the startle response to fast escape from the predators and unfavorable environmental conditions to survive during early phase of life. As nNOS is absent in the adult MC, we assume that the adult MC express a different splice variant of nNOS which is detected by NADPH-d, but not by the currently used antibody. Hence, the presence of nNOS (enzyme required for the synthesis of NO) in the MC suggests the involvement of NO in neurotransmission to generate the startle response rapidly. Furthermore, we also suggest that NO may be involved in other aspects such as pathway finding, neuronal cell migration, signaling and refinement during the development of the brain of L. rohita.