Evaluation of burden of SCN1A pathogenicity in North Indian children with Dravet syndrome.

BACKGROUND: Dravet syndrome is an infantile-onset developmental and epileptic encephalopathy with limited data on the frequency of SCN1A in Indian children. The study aimed to identify and characterize the burden of SCN1A pathogenic variants associated with the Dravet syndrome phenotype through genetic testing in the North Indian population. METHOD: In this prospective, cross-sectional study from March 2015 to February 2019, we enrolled 52 children with Dravet syndrome phenotype who underwent genetic testing for SCN1A gene pathogenicity. We assessed variant effect using multiple algorithms, and genetic test results were reported based on recommendations from the American College of Medical Genetics and Genomics guidelines. Additionally, we performed multiplex-ligation dependent probe amplification (MLPA) to detect copy number variations of the SCN1A gene in children without identified genetic pathogenicity (n = 22) and analysed the results using Coffalyser.net. RESULTS: Of the 52 probands studied, pathogenic variants in the SCN1A gene were identified in 30 children. Among these variants, 11 truncating variants (3 frame-shift variants, 3 intronic variants in splice site regions, and 5 nonsense variants) in 12 unrelated probands, and 17 missense variants in 18 unrelated probands were found. The genetic yield of SCN1A pathogenicity in our cohort (n = 52) was 58 %. Additionally, two of the identified variants were novel. Furthermore, MLPA analysis of the SCN1A gene in 22 children without pathogenic variants yielded no results. CONCLUSION: This work represents a genetic analysis of a Dravet syndrome cohort, revealing a 58 % burden of SCN1A variants in children with the Dravet syndrome phenotype from the North Indian population.

Deciphering the biochemical and functional characterization of rice straw cultivars for industrial applications.

As an agricultural state, Haryana (India) produces about six million metric tons (mt) of rice straw every year from rice cultivation. Currently, rice straw is either burned or ploughed into the field without being turned into a functional product. Burning of paddy straw release green house gases and particulate matter (2.5 and 10 µm), which leads to air pollution and considerable loss of soil property viz. nutrients, organic matter, productivity and biodiversity, and on and off-farm humans and animals' health. The biochemically and functionally specified potential for optimal alternative use of the rice straw of 13 most widely produced rice varieties from Haryana's eastern and western agro-climate zones was undertaken. Pusa-1401 variety had the highest cellulose (46.55%) and silica content (13.70%), while Pusa-1718 had hemicellulose (28.25%) and lignin (11.60%), respectively. Maximum nitrogen (0.81%), phosphorus (0.32%) and potassium (2.78%) were found in rice variety Pusa-1509, Pusa-1401 and Rice-6129. The findings seemed to be statistically significant (p < 0.05). The biochemical profiles of rice straw cultivars were classified into distinct structural groups (C-H alkalanes, O-H alcohol, C[bond, double bond]O, C-H alkanes) based on the FTIR spectrum in order to find the best alternative possibilities for bioethanol and compost production. According to the study, these rice straw varieties could be used to make lucrative industrial products.

Strangles in equines: An overview.

Strangles, caused by Streptococcus equi subspecies equi, is a highly infectious respiratory disease affecting horses and other equines. The disease is economically important and compromises the productivity of equine farm significantly. The disease is characterized by pyrexia, mucopurulent nasal discharge, and abscess formation in the lymph nodes of the head and neck of horses. The disease transmission occurs either directly by coming in contact with infectious exudates or indirectly via fomite transmission. Besides this, carrier animals are the primary and most problematic source of disease infection. The organism not only initiates outbreaks but also makes the control and prevention of the disease difficult. The diagnosis of strangles is best done by isolating and characterizing the bacteria from nasal discharge, pus from abscesses, and lymphoid tissues or by using PCR. ELISA can also be used to detect serum protein M (SeM) antibodies for diagnosis. The most popular treatment for strangles is with penicillin; however, the treatment is affected by the stage, feature and severity of the disease. Prevention and control of strangles can be achieved through vaccination and good hygiene practices. Basically, this review describes the global prevalence of S. equi, as well as general aspects of the disease, like pathogenesis, diagnosis, treatment, prevention, control and management of the disease.

Understanding the killer-cell immunoglobulin like receptor polymorphism in retinoblastoma.

BACKGROUND: The KIR receptors present on the natural killer (NK) cells play a crucial role by exercising cytotoxicity to eliminate tumor cells. Both KIR and class-I HLA molecules exhibit extensive polymorphism. Although RB1 inactivation triggers the initiation of retinoblastoma; however additional immune alterations trigger tumor development. The aim was to explore the KIR/HLA polymorphism and its role in the pathogenesis of retinoblastoma. METHODS: Patients with unilateral, non-familial retinoblastoma were enrolled as cases. Healthy individuals matched for ethnicity were enrolled as controls. KIR genotyping was performed by sequence-specific primer assay. The investigated KIR genes included: inhibitory (2DL1, 2DL2, 2DL3, 2DL4, 2DL5A, 2DL5B), activating (2DS1, 2DS2, 2DS3, 2DS4*FUL, 2DS4*DEL, 2DS5, 3DL1, 3DL2, 3DL3, 3DS1) and pseudogenes (2DP1, 3DP1*FUL, 3DP1*DEL). In addition, HLA ligands were investigated by sequence-specific oligonucleotide assay for HLA-A, B, and C locus. RESULTS: KIR genotyping was performed in 48 cases and 107 controls. The mean age of cases was 2.9 ± 2.2 years (range: 0.25-10). Among the 19 KIR genes, the frequency of KIR2DS4*FUL (p = 0.0019) and 2DS5 (p = 0.0095) was increased among cases. HLA ligands were investigated in 25 cases and 50 controls. The frequency of HLA ligands (C1/C2, Bw4, A3/A11) was similar among cases and controls. However, the KIR/HLA combination frequency for KIR3DS1/HLA-Bw4 was decreased in cases (p = 0.006). CONCLUSION: It is the pioneer study to report the association of killer cell immunoglobulin-like receptors in retinoblastoma. KIR2DS4*FUL and KIR2DS5 had a susceptible, and KIR3DS1/HLA-BW4 had a protective role in retinoblastoma. The results will aid in exploring the therapeutic potential of NK cell-based therapy for retinoblastoma.

Genome-Wide DNA Methylation and Its Effect on Gene Expression During Subclinical Mastitis in Water Buffalo.

Subclinical mastitis (SCM) in buffalo is one of the most challenging paradoxes for the dairy sector with very significant milk production losses and poses an imminent danger to milch animal's milk-producing ability. We present here the genome-wide methylation specific to SCM in water buffalo and its consequential effect on the gene expression landscape for the first time. Whole-genome DNA methylation profiles from peripheral blood lymphocytes and gene expression profiles from milk somatic cells of healthy and SCM cases were catalogued from the MeDIP-Seq and RNA-Seq data. The average methylation in healthy buffaloes was found to be higher than that in the SCM-infected buffaloes. DNA methylation was abundant in the intergenic region followed by the intronic region in both healthy control and SCM groups. A total of 3,950 differentially methylated regions (DMRs) were identified and annotated to 370 differentially methylated genes (DMGs), most of which were enriched in the promoter region. Several important pathways were activated due to hypomethylation and belonged to the Staphylococcus aureus infection, Th17 cell differentiation, and antigen processing and presentation pathways along with others of defense responses. DNA methylome was compared with transcriptome to understand the regulatory role of DNA methylation on gene expression specific to SCM in buffaloes. A total of 4,778 significant differentially expressed genes (DEGs) were extracted in response to SCM, out of which 67 DMGs were also found to be differentially expressed, suggesting that during SCM, DNA methylation could be one of the epigenetic regulatory mechanisms of gene expression. Genes like CSF2RB, LOC102408349, C3 and PZP like, and CPAMD8 were found to be downregulated in our study, which are known to be involved in the immune response to SCM. Association of DNA methylation with transposable elements, miRNAs, and lncRNAs was also studied. The present study reports a buffalo SCM web resource (BSCM2TDb) available at http://webtom.cabgrid.res.in/BSCM2TDb that catalogues all the mastitis-related information of the analyses results of this study in a single place. This will be of immense use to buffalo researchers to understand the host-pathogen interaction involving SCM, which is required in endeavors of mastitis control and management.

Deciphering the TLR transcriptome and downstream signaling pathway in cerebrospinal fluid in pediatric meningitis.

OBJECTIVE/DESIGN: Pediatric meningitis is characterized by a colossal inflammatory response to the pathogen in the central nervous system (CNS). This unabated inflammatory response persists even after the removal of the pathogen by antibiotics/steroids causing collateral damage to CNS tissue. Toll-like receptors (TLRs) are the key players in the recognition and elicitation of innate-immune response against bacterial/viral components in cerebrospinal fluid (CSF). Till date, the precise understanding of TLR-triggered inflammatory response in pediatric meningitis is lacking. The present study was designed to delineate the role of TLR transcriptome and downstream signaling pathways in CSF of pediatric meningitis. METHODS: Children in the age group of > 3 months to 12 years with pediatric meningitis were included. A total of 249 cases of pediatric meningitis (bacterial = 89, viral = 160) were included. In addition, 71 children who tested negative to the pathogen in CSF tap and did not have signs of infection clinically constituted the controls. RNA was extracted from the CSF samples of both cases and controls. The relative gene expression profile of 42 TLR signaling pathway genes was performed. For the analysis of secretory cytokines and chemokines in CSF, Luminex assay was performed. RESULTS: We report global upregulation of TLR genes in patients with acute bacterial meningitis (ABM). The downstream signaling molecules were upregulated as well. The CSF of pediatric ABM patients revealed a predominant pro-inflammatory milieu marked by increased levels of pro-inflammatory cytokines. A significant correlation between poor clinical outcomes of patients and an increased expression of TLR/pro-inflammatory cytokine genes was observed. CONCLUSION: Our findings provide support for future studies exploring TLR-based adjunct therapy to limit the neurological sequelae, owing to persistent inflammation in pediatric ABM patients.

Understanding HLA-G driven journey from HPV infection to cancer cervix: Adding missing pieces to the jigsaw puzzle.

Human Papillomavirus (HPV) is a vital risk-factor for cancer cervix. However, persistent HPV infection results in cervical cancer in only a minority. Probably, HPV subdues the host immune response for persistence, which includes augmentation of HLA-G and plausibly aids in progression to cervical cancer. HLA-G, which comprises of membrane and soluble form, downregulates the host's immune response and generate tolerance. The current study aimed to analyze both forms of HLA-G in fresh tissue and plasma of women with HPV-infected and uninfected cervix and cancer cervix using Western blot and ELISA. The study cohort included 30 women with cervical carcinoma and equal number with normal cervix and 6 with HPV infected cervix. We observed a significant upregulation of membranous HLA-G expression in HPV infected cervix and cervical carcinoma (P < 0.001). Interestingly, the pairwise comparison of HLA-G tissue protein expression of the normal cervix and cervical carcinoma, as well as the normal cervix with HPV infected cervix, was significant (P < 0.001). Levels of soluble HLA-G were significantly raised in carcinoma cervix. We observed a progressive increase in HLA-G protein expression in HPV infected cervix and cervical carcinoma. These findings compel us to hypothesize that the upregulation of HLA-G expression favors the persistence of HPV in a microenvironment of a submissive host response. This progressive upregulation further leads to cervical cancer. Thus elimination of HPV infection seems to be a desirable proposition to prevent cervical cancer. In the absence of antiviral therapy for HPV, exploration of HLA-G antibody-based therapeutic strategies appear promising.

Gold nanoparticles modulate the steroidogenic and apoptotic pathway in a buffalo granulosa cell model.

OBJECTIVES: Granulosa cells are associated with steroidogenesis and ovarian function in females. Aims of the study are to understand the effects of gold nanoparticles (AuNP) on steroidogenesis and apoptotic pathway associated genes in buffalo granulosa cells. RESULTS: The AuNP were prepared chemically and thereby characterized by transmission electron microscope (TEM) imaging, absorbance and dynamic light scattering (DLS) measurements for hydrodynamic diameter and zeta potential. The cultured buffalo granulosa cells (BGC) were co-incubated with AuNP in two concentrations (2 × 109 and 2 × 1010 AuNP/ml) for 24 h. Treatment of BGC with AuNP significantly modulated the steroidogenesis associated genes (3ß-Hsd and Cyp19A1) expression and progesterone accumulation in the culture fluid. AuNP affected the apoptotic pathway in BGC by affecting the gene expression of Caspase-3, Bad and Bax. The AuNP did not exert oxidative stress through anti-oxidant induction & lipid peroxidation in the buffalo GC. CONCLUSIONS: AuNP may modulate the endocrine system by having impact on the steroidogenesis pathway and also have the potential to affect apoptotic pathway in a buffalo granulosa cell model.