Isolation and Molecular Characterisation of Alkaline Protease Producing Bacillus thuringiensis

Proteases are of particular interest because of their action on insoluble keratin substrates and generally on a broad range of protein substrates. Proteases are one of the most important groups of industrial enzymes used in detergent, protein, brewing, meat, photographic, leather, dairy, pharmaceutical and food industry. In the present study, the organism isolated from the protein rich soil sample was identified by biochemical and molecular characterisation as Bacillus thuringiensis and further optimum conditions for alkaline protease synthesis were determined. The growth conditions for B. thuringiensis was optimised by inoculating into yeast extract casein medium at different pH and incubating at different temperatures. The maximum protease production occurred at pH 8 and at 37 °C. B. thuringiensis showed proteolytic activity at various culture conditions. Optimum conditions for the protease activity were found to be 47 °C and pH 8. In the later stage, the blood removing action of crude and partially purified protease was found to be effective within 25 min in the presence of commercial detergents indicating the possible use of this enzyme in detergent industry. Enzyme also showed good activity against hair substrate keratin and can be used for dehairing.     

Immunotherapy of prostate cancer: identification of new treatments and targets for therapy, and role of WAP domain-containing proteins

Prostate adenocarcinoma is present in over 80% of men over the age of 80 and is by far the most common cancer of men. Although radical prostatectomy is curative in early disease, the risks of incontinence and impotence can affect the quality of life of patients. Early intervention with localized immunotherapy represents a potential solution as lymphocyte infiltration does occur in prostate cancer lesions, and immunotherapy with dendritic cell vaccines can significantly increase survival in late stage disease. However, lymphocytic infiltrates in the cancerous prostates have an anergic character arising from the suppressive effects of the microenvironment resulting from a conversion of effector cells into regulatory T-cells. Although TGFβ (transforming growth factor β) and IL-10 (interleukin-10) are known to be strong suppressor molecules associated with prostate cancer, they are among many possible suppressive factors. We discuss the possible role of alternative suppressor molecules, including the WAP (whey acidic protein) homologue ps20 that is expressed on prostate stroma and other WAP domain-containing proteins in the immunosuppressive prostate cancer milieu and discuss novel immunotherapeutic strategies to combat this disease.     

Discovery of pyrazine carboxamide CB1 antagonists: the introduction of a hydroxyl group improves the pharmaceutical properties and in vivo efficacy of the series

Structure-activity relationships for a series of pyrazine carboxamide CB1 antagonists are reported. Pharmaceutical properties of the series are improved via inclusion of hydroxyl-containing sidechains. This structural modification sufficiently improved ADME properties of an orally inactive series such that food intake reduction was achieved in rat feeding models. Compound 35 elicits a 46% reduction in food intake in ad libidum fed rats 4-h post-dose.     

MED12 somatic mutations encompassing exon 2 associated with benign breast fibroadenomas and not breast carcinoma in Indian women

Fibroadenoma is the most common type of benign breast tumor, accounting for 90% of benign lesions in India. Somatic mutations in the mediator complex subunit 12 (MED12) gene play a critical role in fibroepithelial tumorigenesis. The current study evaluated the hotspot region encompassing exon 2 of the MED12 gene, in benign and malignant breast tumor tissue from women who presented for breast lump evaluation. A total of 100 (80 fibroadenoma and 20 breast cancer) samples were analyzed by polymerase chain reaction-Sanger sequencing. Sequence variant analysis showed that 68.75% of nucleotide changes were found in exon 2 and the remaining in the adjacent intron 1. Codon 44 was implicated as a hotspot mutation in benign tumors, and 86.36% of the identified mutations involved this codon. An in silico functional analysis of missense mutations using consensus scoring sorting intolerant from tolerant (SIFT), SIFT seq, Polyphen2, Mutation Assessor, SIFT transFIC, Polyphen2 transFIC, Mutation Assesor transFIC, I-Mutant, DUET, PON-PS, SNAP2, and protein variation effect analyzer] revealed that apart from variants involving codon 44 (G44S; G44H), others like V41A and E55D were also predicted to be deleterious. Most of the missense mutations appeared in the loop region of the MED12 protein, which is expected to affect its functional interaction with cyclin C–CDK8/CDK19, causing loss of mediator-associated cyclin depended kinase (CDK) activity. These results suggest a key role of MED12 somatic variations in the pathogenesis of fibroadenoma. For the first time, it was demonstrated that MED12 sequence variations are present in benign breast tumors in the south Indian population.     

ACC deaminase positive Enterobacter-mediated mitigation of salinity stress,and plant growth promotion of Cajanus cajan: a lab to field study

Salinity is a major abiotic stress that negatively impacts plant health and soil microbiota. ACC (1-aminocyclopropane carboxylic acid) deaminase producing microorganisms act as natural stress busters that protect plants from different kinds of stresses. The study focused on the isolation of potent, indigenous, multi-trait ACC deaminase producers. The shortlisted ACC deaminase producers were checked for their ability to promote growth of Cajanus cajan, and mitigate stress under laboratory conditions followed by validation of their potency in naturally saline field conditions. Physiological stress markers were assessed to evaluate the impact of salinity in plants treated with ACC deaminase producer, compared to controls. Further, the contribution of ACC deaminase in stress mitigation was demonstrated by using a chemical inhibitor for ethylene biosynthesis. This study presents a polyphasic approach, transitioning from the rhizospheric soil to the laboratory to validation in the field, and puts forth a promising eco-friendly alternative for sustainable agriculture.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12298-021-01031-0.     

Microbial biosynthesis and applications of gentamicin: a critical appraisal

Gentamicin is an aminoglycoside antibiotic produced by various species of the genus Micromonospora and has received much attention in the recent years as a broad-spectrum antibiotic for treatment of various infections. It exists as a complex of closely related aminoglycoside structures and the clinically significant one is the gentamicin C complex. This review article focuses attention on the present status of knowledge and the main advancements achieved in the last few decades on the subject of gentamicin with regard to its production, biosynthetic pathway, mode of action, and uses. The various nutritional and environmental parameters affecting gentamicin production and the factors affecting the release of bound gentamicin are discussed. Further, strain improvement using UV and/or chemical mutagenesis can be applied to augment the efficiency of the producer strain and a number of case studies are presented. Different detection and quantitative methods for gentamicin estimation and the mode of action of gentamicin are discussed in detail. This antibiotic finds extensive use in combination chemotherapy and as a drug for different delivery agents for treatment of osteomyelitis and other recent applications in gene therapy.     

Development of new genomic microsatellite markers from robusta coffee (<Emphasis Type="Italic">Coffea canephora</Emphasis>Pierre ex A. Froehner) showing broad cross-species transferability and utility in genetic studies

Background  

Species-specific microsatellite markers are desirable for genetic studies and to harness the potential of MAS-based breeding for genetic improvement. Limited availability of such markers for coffee, one of the most important beverage tree crops, warrants newer efforts to develop additional microsatellite markers that can be effectively deployed in genetic analysis and coffee improvement programs. The present study aimed to develop new coffee-specific SSR markers and validate their utility in analysis of genetic diversity, individualization, linkage mapping, and transferability for use in other related taxa.     

Effects of female sex steroids on concanavalin A-mediated agglutination of hepatocytes from nonregenerating and regenerating rat liver and hepatic tumor marker enzymes

Effect of treatment of female rats with an oral contraceptive agent (OCA), Ovulen-50, for 7 weeks on agglutination of hepatocytes with concanavalin A (con A) and activities of certain tumor marker enzymes were examined to find out if OCA treatment is related to preneoplastic or neoplastic processes. Hepatocytes from regenerating and nonregenerating livers of control female rats showed negligible agglutination with Con A, whereas hepatocytes from non regenerating but not from the regenerating livers of female rats treated with a combination of 5 micrograms ethinyl estradiol and 100 micrograms ethynodiol diacetate showed agglutination. Of the tumor marker enzymes such as hepatic glucose 6-phosphatase, gamma-glutamyl transpeptidase (gamma-GT), and arginase examined in the liver, only gamma-glutamyl transpeptidase showed a significant increase in activity in the steroid-treated rats. Plasma alkaline phosphatase activity was also higher in the treated animals. However, the magnitude of the changes observed was relatively small and perhaps unrelated to the neoplastic process.     

Next‐generation sequencing for identification of candidate genes for Fusarium wilt and sterility mosaic disease in pigeonpea (Cajanus cajan)

To map resistance genes for Fusarium wilt (FW) and sterility mosaic disease (SMD) in pigeonpea, sequencing‐based bulked segregant analysis (Seq‐BSA) was used. Resistant (R) and susceptible (S) bulks from the extreme recombinant inbred lines of ICPL 20096 × ICPL 332 were sequenced. Subsequently, SNP index was calculated between R‐ and S‐bulks with the help of draft genome sequence and reference‐guided assembly of ICPL 20096 (resistant parent). Seq‐BSA has provided seven candidate SNPs for FW and SMD resistance in pigeonpea. In parallel, four additional genotypes were re‐sequenced and their combined analysis with R‐ and S‐bulks has provided a total of 8362 nonsynonymous (ns) SNPs. Of 8362 nsSNPs, 60 were found within the 2‐Mb flanking regions of seven candidate SNPs identified through Seq‐BSA. Haplotype analysis narrowed down to eight nsSNPs in seven genes. These eight nsSNPs were further validated by re‐sequencing 11 genotypes that are resistant and susceptible to FW and SMD. This analysis revealed association of four candidate nsSNPs in four genes with FW resistance and four candidate nsSNPs in three genes with SMD resistance. Further, In silico protein analysis and expression profiling identified two most promising candidate genes namely C.cajan_01839 for SMD resistance and C.cajan_03203 for FW resistance. Identified candidate genomic regions/SNPs will be useful for genomics‐assisted breeding in pigeonpea.