Purification and characterization of a high molecular weight serine protease from Microbacterium paraoxydans sp. SKS10.

Alkaline proteases from microbial sources have been found suitable for diverse industrial applications, with serine proteases being the most common enzymes used in the detergent industry. In the present study, we have purified and characterized an extracellular alkaline serine protease from Microbacterium paraoxydans sp. SKS10. The protease was purified using ammonium sulfate precipitation followed by different chromatography techniques (fold purification 6.919). Km and Vmax for the protease were determined to be 0.183 mg/mL and 4.904 U/mL, respectively. This enzyme is a thermostable high molecular weight (∼109.4 kDa) protease which has maximal activity at 60°C, and above pH 10. Inhibitor assays revealed the enzyme to be a serine protease whose activity increased by 2.5-fold in the presence of EDTA. This enzyme remained active in the presence of various metal salts and organic solvents and was compatible with commercially available laundry detergents highlighting its potential for use in the detergent industry.

Exploring the role of microbes for the management of persistent organic pollutants.

Persistent organic pollutants (POPs) are chemicals which have been persisting in the environment for many years due to their longer half-lives. POPs have gained attention over the last few decades due to the unsustainable management of chemicals which led to their widespread and massive contamination of biota from different strata and environments. Due to the widespread distribution, bio-accumulation and toxic behavior, POPs have become a risk for organisms and environment. Therefore, a focus is required to eliminate these chemicals from the environment or transform into non-toxic forms. Among the available techniques for the removal of POPs, most of them are inefficient or incur high operational costs. As an alternative to this, microbial bioremediation of POPs such as pesticides, polycyclic aromatic hydrocarbons, polychlorinated biphenyls, pharmaceuticals and personal care products is much more efficient and cost-effective. Additionally, bacteria play a vital role in the biotransformation and solubilization of POPs, which reduces their toxicity. This review specifies the Stockholm Convention that evaluates the risk profile for the management of existing as well as emerging POPs. The sources, types and persistence of POPs along with the comparison of conventional elimination and bioremediation methods of POPs are discussed comprehensively. This study demonstrates the existing bioremediation techniques of POPs and summaries the potential of microbes which serve as enhanced, cost-effective, and eco-friendly approach for POPs elimination.

A large-effect QTL introgressed from ricebean imparts resistance to Mungbean yellow mosaic India virus in blackgram (Vigna mungo (L.) Hepper).

KEY MESSAGE: Here, we report identification of a large effect QTL conferring Mungbean yellow mosaic India virus resistance introgressed from ricebean in blackgram variety Mash114. The tightly linked KASP markers would assist in marker-assisted-transfer of this region into Vigna species infected by MYMIV. Until recently, precise location of genes and marker-assisted selection was long thought in legumes such as blackgram due to lack of dense molecular maps. However, advances in next-generation sequencing based on high-throughput genotyping technologies such as QTL-seq have revolutionized trait mapping in marker-orphan crops. Using QTL-seq approach, we have identified a large-effect QTL for resistance to Mungbean yellow mosaic India virus (MYMIV) in blackgram variety Mash114. MYMIV is devastating disease responsible for huge yield losses in blackgram, greengram and other legumes. Mash114 showed consistent and high level of resistance to MYMIV since last nine years. Whole genome re-sequencing of MYMIV-resistant and susceptible bulks derived from RILs of cross KUG253 X Mash114 identified a large-effect QTL (qMYMIV6.1.1) spanning 3.4 Mb on chromosome 6 explaining 70% of total phenotypic variation. This region was further identified as an inter-specific introgression from ricebean. Linkage mapping using KASP markers developed from potent candidate genes involved in virus resistance identified the 500 kb genomic region equaling 1.9 cM on genetic map linked with MYMIV. The three KASP markers closely associated with MYMIV originated from serine threonine kinase, UBE2D2 and BAK1/BRI1-ASSOCIATED RECEPTOR KINASE genes. These KASPs can be used for marker-assisted transfer of introgressed segment into suitable backgrounds of Vigna species.

Developments and Prospects in Imperative Underexploited Vegetable Legumes Breeding: A Review.

Vegetable legumes are an essential source of carbohydrates, vitamins, and minerals, along with health-promoting bioactive chemicals. The demand for the use of either fresh or processed vegetable legumes is continually expanding on account of the growing consumer awareness about their well-balanced diet. Therefore, sustaining optimum yields of vegetable legumes is extremely important. Here we seek to present d etails of prospects of underexploited vegetable legumes for food availability, accessibility, and improved livelihood utilization. So far research attention was mainly focused on pulse legumes' performance as compared to vegetable legumes. Wild and cultivated vegetable legumes vary morphologically across diverse habitats. This could make them less known, underutilized, and underexploited, and make them a promising potential nutritional source in developing nations where malnutrition still exists. Research efforts are required to promote underexploited vegetable legumes, for improving their use to feed the ever-increasing population in the future. In view of all the above points, here we have discussed underexploited vegetable legumes with tremendous potential; namely, vegetable pigeon pea (Cajanus cajan), cluster bean (Cyamopsis tetragonoloba), winged bean (Psophocarpus tetragonolobus), dolichos bean (Lablab purpureus), and cowpea (Vigna unguiculata), thereby covering the progress related to various aspects such as pre-breeding, molecular markers, quantitative trait locus (QTLs), genomics, and genetic engineering. Overall, this review has summarized the information related to advancements in the breeding of vegetable legumes which will ultimately help in ensuring food and nutritional security in developing nations.

Myxobacteria: biology and bioactive secondary metabolites.

Myxobacteria are Gram-negative eubacteria and they thrive in a variety of habitats including soil rich in organic matter, rotting wood, animal dung and marine environment. Myxobacteria are a promising source of new compounds associated with diverse bioactive spectrum and unique mode of action. The genome information of myxobacteria has revealed many orphan biosynthetic pathways indicating that these bacteria can be the source of several novel natural products. In this review, we highlight the biology of myxobacteria with emphasis on their habitat, life cycle, isolation methods and enlist all the bioactive secondary metabolites purified till date and their mode of action.

Genome-wide Association Study for Yield and Yield-Related Traits in Diverse Blackgram Panel (Vigna mungo L. Hepper) Reveals Novel Putative Alleles for Future Breeding Programs.

Blackgram (Vigna mungo L. Hepper) is an important tropical and sub-tropical short-duration legume that is rich in dietary protein and micronutrients. Producing high-yielding blackgram varieties is hampered by insufficient genetic variability, absence of suitable ideotypes, low harvest index and susceptibility to biotic-abiotic stresses. Seed yield, a complex trait resulting from the expression and interaction of multiple genes, necessitates the evaluation of diverse germplasm for the identification of novel yield contributing traits. Henceforth, a panel of 100 blackgram genotypes was evaluated at two locations (Ludhiana and Gurdaspur) across two seasons (Spring 2019 and Spring 2020) for 14 different yield related traits. A wide range of variability, high broad-sense heritability and a high correlation of grain yield were observed for 12 out of 14 traits studied among all environments. Investigation of population structure in the panel using a set of 4,623 filtered SNPs led to identification of four sub-populations based on ad-hoc delta K and Cross entropy value. Using Farm CPU model and Mixed Linear Model algorithms, a total of 49 significant SNP associations representing 42 QTLs were identified. Allelic effects were found to be statistically significant at 37 out of 42 QTLs and 50 known candidate genes were identified in 24 of QTLs.

SLIM: Simultaneous Logic-in-Memory Computing Exploiting Bilayer Analog OxRAM Devices.

von Neumann architecture based computers isolate computation and storage (i.e. data is shuttled between computation blocks (processor) and memory blocks). The to-and-fro movement of data leads to a fundamental limitation of modern computers, known as the Memory wall. Logic in-Memory (LIM)/In-Memory Computing (IMC) approaches aim to address this bottleneck by directly computing inside memory units thereby eliminating energy-intensive and time-consuming data movement. Several recent works in literature, propose realization of logic function(s) directly using arrays of emerging resistive memory devices (example- memristors, RRAM/ReRAM, PCM, CBRAM, OxRAM, STT-MRAM etc.), rather than using conventional transistors for computing. The logic/embedded-side of digital systems (like processors, micro-controllers) can greatly benefit from such LIM realizations. However, the pure storage-side of digital systems (example SSDs, enterprise storage etc.) will not benefit much from such LIM approaches as when memory arrays are used for logic they lose their core functionality of storage. Thus, there is the need for an approach complementary to existing LIM techniques, that's more beneficial for the storage-side of digital systems; one that gives compute capability to memory arrays not at the cost of their existing stored states. Fundamentally, this would require memory nanodevice arrays that are capable of storing and computing simultaneously. In this paper, we propose a novel 'Simultaneous Logic in-Memory' (SLIM) methodology which is complementary to existing LIM approaches in literature. Through extensive experiments we demonstrate novel SLIM bitcells (1T-1R/2T-1R) comprising non-filamentary bilayer analog OxRAM devices with NMOS transistors. Proposed bitcells are capable of implementing both Memory and Logic operations simultaneously. Detailed programming scheme, array level implementation, and controller architecture are also proposed. Furthermore, to study the impact of proposed SLIM approach for real-world implementations, we performed analysis for two applications: (i) Sobel Edge Detection, and (ii) Binary Neural Network- Multi layer Perceptron (BNN-MLP). By performing all computations in SLIM bitcell array, huge Energy Delay Product (EDP) savings of ≈75× for 1T-1R (≈40× for 2T-1R) SLIM bitcell were observed for edge-detection application while EDP savings of ≈3.5× for 1T-1R (≈1.6× for 2T-1R) SLIM bitcell were observed for BNN-MLP application respectively, in comparison to conventional computing. EDP savings owing to reduction in data transfer between CPU ↔ memory is observed to be ≈780× (for both SLIM bitcells).

Enzymatic Degradation of Biofilm by Metalloprotease From Microbacterium sp. SKS10.

Enzymes have replaced or decreased usage of toxic chemicals for industrial and medical applications leading toward sustainable chemistry. In this study, we report purification and characterization of a biofilm degrading protease secreted by Microbacterium sp. SKS10. The protease was identified as a metalloprotease, Peptidase M16 using mass spectrometry. It showed optimum activity at 60°C, pH 12 and retained its activity in the presence of various salts and organic solvents. The enzyme was able to degrade biofilms efficiently at enzyme concentration lower than other known enzymes such as papain, trypsin and α-amylase. The presence of this protease increased the accessibility of antibiotics inside the biofilm, and was found to be non-cytotoxic toward human epidermoid carcinoma cells (A431) at the effective concentration for biofilm degradation. Thus, this protease may serve as an effective tool for management of biofilms.

Anti-brain cancer activity of chloroform and hexane extracts of Tinospora cordifolia Miers: an in vitro perspective.

BACKGROUND: Plants have been suggested as safest source of therapeutic agents, with multi targeted mode of action and least side effects. Tinospora cordifolia, commonly known as Guduchi in India, is one of the most highly valued herbs in Ayurvedic medicine. It possesses potential anti-cancer, anti-inflammatory, hepatoprotective, anti-diabetic, immune-stimulatory and various other beneficial activities. PURPOSE: The present study was aimed to investigate the differentiation inducing potential of chloroform and hexane extracts of T. cordifolia using U87MG glioblastoma and IMR-32 neuroblastoma cell lines as model system. RESULTS: Chloroform (Chl-TCE) and hexane (Hex-TCE) extracts significantly reduced the rate of proliferation and induced cell differentiation as evidenced by MTT assay and immunostaining for GFAP and MAP-2 in glioblastoma and neuroblastoma, respectively. Further these extracts increased the expression of stress markers HSP70 and Mortalin and induced senescence. Chloroform and hexane extracts also inhibited the migration of U87MG glioblastoma and IMR-32 neuroblastoma as indicated by wound scratch assay and supported by reduced expression of NCAM. Furthermore these extracts are not toxic to normal cells as they showed no inhibitory effects on primary astrocytic and neuronal cultures. CONCLUSIONS: The present study suggests that chloroform and hexane extracts of T. cordifolia retard the rate of proliferation, induce differentiation and inhibit migration of human glioblastomas and neuroblastomas, thus may act as potential phytotherapeutic intervention in treatment of neural cancers.

Characterization of thermostable alkaline proteases from Bacillus infantis SKS1 isolated from garden soil.

Proteases are one of the largest groups of hydrolytic enzymes constituting about 60% of total worldwide sales of industrial enzymes due to their wide applications in detergent, leather, textile, food and pharmaceutical industry. Microbial proteases have been preferred over animal and plant proteases because of their fundamental features and ease in production. Bacillus infantis SKS1, an alkaline protease producing bacteria has been isolated from garden soil of north India and identified using morphological, biochemical and molecular methods. 16S rDNA sequence amplified using universal primers has 99% sequence identity with corresponding gene sequence of Bacillus infantis strain FM 34 and Bacillus sp. Beige. The bacterial culture and its 16S rDNA gene sequence have been deposited to Microbial Culture Collection (Pune, India) with accession number MCC 3035 and GenBank with accession number KR092197 respectively. The partially purified extract of Bacillus infantis SKS1 was thermostable and active in presence of Mg2+, acetyl acetone and laundry detergents implicating its application in industry. Production of these enzymes using this strain was maximized by optimization of various parameters including temperature, pH, media components and other growth conditions. Our results show that fructose and dextrose serve as the best carbon sources for production of these enzymes, highlighting the use of this strain for enzyme production utilizing relatively inexpensive substrates like beet molasses and corn steep liquor. Additionally, this strain showed maximum production of enzymes at 40°C similar to bacterial species used for commercial production of alkaline proteases. Characterization of alkaline proteases from this strain of Bacillus infantis and optimization of parameters for its production would help in understanding its industrial application and large-scale production.

Indoxacarb interaction alters immunotoxic and genotoxic potential of endotoxin.

Indoxacarb is commonly used to effectively control pests, cockroaches, termites, fleas, and houseflies. Although the toxicological profile of indoxacarb had already been well characterized, we examined the possible toxicological interaction with indoxacarb and endotoxin. Male Swiss albino mice aged 8-10 weeks were orally administered indoxacarb dissolved in groundnut oil at 4 mg/kg/day and 2 mg/kg/day for 90 days. On day 91, five animals from each group were challenged with lipopolysaccharides (LPS) at 80 µg/mouse, administered intranasally. Indoxacarb at 4 mg/kg significantly decreased Total leukocyte count, lymphocytopenia, and neutrophilia. Both doses of indoxacarb combined with LPS resulted in significant lymphocytopenia. Indoxacarb did not produce DNA damage in comet assay, but when combined with LPS, it resulted in a significant increase in tail length, tail moment, and olive moment. The data indicate that indoxacarb at 4 mg/kg administered orally for 90 days induced immune-response change. Further, both doses of indoxacarb, when combined with LPS, accelerate immunotoxicity and endotoxin-induced DNA damage.

A new simplified beading and boxing procedure for elastic impression.

Beading and Boxing of impression is taught in most dental colleges. The boxing procedure is crucial step to preserve the details of the final impression especially of the vestibular area. This article describes an alternative beading-boxing procedure that is compatible with all impression materials, is efficient, simple, inexpensive, and practicable. Use of commercially available instant adhesive around the border to act as a joining agent between elastic impressions and beading wax or bead made up of base plate wax is advocated in this technique.

Obesity among pre-adolescent and adolescents of a developing country (India).

There are very few reports from the developing world on the prevalence of obesity among children even though in developed countries it has reached epidemic proportions. The objective of this study was to determine the prevalence of obesity in pre-adolescent and adolescent children in a developing country (India) using WHO guidelines for defining obesity and overweight. This cross-sectional study was carried out on 2008 school-children aged 9-15 years. Approximately half the subjects belonged to a school attended by children of well to do families while the rest belonged to two schools from middle and lower socio-economic background. Weight and height were taken for all children and the body mass index (BMI) calculated. Children whose BMI was >85th percentile for age and sex were defined as overweight. Triceps skin fold thickness (TSFT) was measured for all overweight children and those with TSFT >90th percentile for age and sex were defined as obese. The overall prevalence of obesity and overweight was 11.1% and 14.2% respectively. The prevalence of obesity as well as overweight was higher in boys as compared to girls (12.4% vs 9.9%, 15.7% vs 12.9%). Prevalence of obesity decreased significantly with age, from 18.5% at 9 years to 7.6% at 14 years, rising at 15 years to 12.1%. Significantly more children from higher socio-economic status were obese and overweight than those from lower socio-economic status groups. No significant gender difference for obesity prevalence was seen among children from a less privileged background, however, amongst children from affluent families, significantly more boys were obese as compared to girls. Pediatric obesity is an emerging problem in developing countries, especially among higher socio-economic status groups. Significant gender disparity is seen, with boys of affluent background having a higher prevalence.