Pročišćavanje i karakterizacija ekstracelularne dekstran saharaze iz bakterije Pediococcus pentosaceus, izolirane iz tla sjeveroistočne Indije

The extracellular dextransucrase produced from Pediococcus pentosaceus, a new isolate from the soil in Assam, India, was purified and characterized. The enzyme activity of cell-free supernatant was 3.4 U/mL and specific activity was 0.6 U/mg. The crude enzyme was purified by a single-step fractionation using polyethylene glycols of different molecular mass. The specific activity achieved was 18 U/mg with 31-fold purification by PEG 400 and 26 U/mg with 45-fold purification by PEG 1500. The molecular mass of dextransucrase determined by non-denaturing SDS-PAGE was approx. 180 kDa. The dextran formation activity of the enzyme was confirmed by activity staining. Optimum conditions for dextransucrase activity were: pH=5.4, reaction temperature 30 °C, 5 % sucrose and 20 mM sodium acetate buffer. A concentration of 1 mM MgCl2 and 6 mM CaCl2 enhanced dextransucrase activity by 5 and 150 %, respectively. The chaotropic agent urea (7 M) and chelating agent EDTA (1 mM) resulted in the residual enzyme activity of 98 and 80 %, respectively. The organic solvents such as ethanol (50 %), DMSO (90 %), acetone (50 %) and acetonitrile (20 %) decreased the dextransucrase activity by 80, 91, 94 and 80 %, respectively.U radu je pročišćena i okarakterizirana dekstran saharaza iz bakterije Pediococcus pentosaceus, izolirane iz tla u gradu Assamu, Indija. Aktivnost je enzima u supernatantu bila 3,4 U/mL, a njegova je specifična aktivnost iznosila 0,6 U/mg. Sirovi je enzim pročišćen jednostupanjskim frakcioniranjem pomoću polietilen glikola različite molekularne mase. Utvrđena je specifična aktivnost enzima od 18 (pročišćenog 31 put pomoću PEG 400), odnosno 26 U/mg (pročišćenog 45 puta pomoću PEG 1500). Molekularna je masa dekstran saharaze određena pomoću SDS-PAGE, a iznosila je otprilike 180 kDa. Aktivnost je enzima potvrđena bojanjem nastalog dekstrana s Coomasie brilijant plavom bojom. Optimalni su uvjeti za aktivnost enzima bili: pH=5,4; temperatura reakcije od 30 °C; te dodatak 5 %-tne saharoze i acetatnog pufera (20 mM). Dodatak 1 mM MgCl2 i 6 mM CaCl2 povećali su aktivnost enzima za 5, odnosno 150 %. Inaktivirajući agensi, poput uree (7 M) i EDTA (1 mM) smanjili su aktivnost enzima na 98 odnosno 80 %. Organska su otapala također smanjila aktivnost enzima, i to: 50 %-tni etanol na 80 %, 90 %-tni DMSO na 91 %, 50 %-tni aceton na 94 % i 20 %-tni acetonitril na 80 %

Role of Artistic Copper-T Shiksha Model to sensitize women for the use of Copper–T as a safe contraceptive

The contraceptives are integral part of healthy sexual life. The choice of contraceptives depends on family planning and will of couple specially woman. Copper-T, an intrauterine device (IUD) is one of the most effective, cheapest and reversible contraceptive methods. It can be used for both spacing and birth control and it has minimum side effects as compared to other contraceptives. But it is highly neglected contraceptive though Copper- T has High pearl index and has less failure rate than other temporary contraceptive methods. As most of women are unaware of its potential significance & refuse it after misconception of perforation of uterus and expulsion. Copper-T Shiksha Model is innovative way of contraceptive counselling by artistic presentation to female and her family even in presence of children. It is based on the Arts Integration Approach and Neuro-linguistic programming. Copper-T is promoted as ornament of uterus instead of sharp object and displayed beautifully with a framed painting with significant meaning of contraception by famous artist Rahul Bhandare. Woman is also warned of side effects of Copper-T and conditions like excessive bleeding. It boosts up faith and confidence in contraceptive method. Both hand cards and clinic model is used in awareness. 400 women were selected randomly in group meets, door to door survey and individual counselling

Analysis of Reliability and Cost of Complex Systems with Metaheuristic Algorithms

Introduction. The assessment of the reliability and cost of complex systems, such as Complex Bridge Systems (CBS) and Life Support Systems in Space Capsules (LSSSC), is fascinating. To achieve the ideal system design through diverse constraints and increase overall system reliability, researchers have extensively explored system reliability and cost optimization problems. Hence, the significant advancement in metaheuristic methods is the primary source of further system reliability and cost optimization process refinement. Aim and tasks. This research attempts to enhance the reliability and cost of complex systems named CBC and LSSSC has been presented.   Results. The structure is based on few recent metaheuristic techniques, such as Moth Flame Optimization (MFO), Whale Optimization Algorithm (WOA), Gazelle Optimization Algorithm (GOA), Dragonfly Algorithm (DA), and Coati Optimization Algorithm (COA). Comparing the acquired findings to those found in other proposed techniques demonstrates the usefulness of a methodology based on COA. The proposed COA algorithm exhibits enhanced efficiency by offering superior solutions to reliability and cost-optimization problems. In addition, a non-parametric Friedman ranking was performed for validation. The results of this research are based on improving the reliability of the parameters and decreasing complex systems’ costs used by the five metaheuristic methods. Observing the convergence graph, Friedman ranking, statistical results test, and tables determined that COA is the most effective algorithm for a complex system’s cost and reliability parameters compared to other existing approaches, and also provided a faster solution. Conclusions. This study proposes unique ways to reduce costs while increasing parameter reliability in complex systems. After analysing the comparative solution, the authors found that when comparing these approaches (GOA, DA, MFO, WOA, and COA), the COA provided the best minimum solution for the cost and reliability of complex systems. Hence, the suggested COA procedure was more successful than that described in this study

Dextransucrase from the mutant of Pediococcus pentosaceus (PPm) is more stable than the wild type

A comparative study on both wild type and mutant of Pediococcus pentosaceus for dextransucrase activity, its stability, dextran synthesizing activity, antibiotic sensitivity and carbohydrate utilization was performed. The wild type P. pentosaceus had specific activity of 0.58 U/mg whereas the mutant showed that of 1.0 U/mg with 72% enhancement. The antibiogram of 27 antibiotics tested against mutant showed significant differences with 9 antibiotics when compared to wild type. In carbohydrate fermentation profile, trehalose, galactose, maltose, lactose and fructose are metabolized by both the strains, but weakly in case of mutant. Stabilization of purified dextransucrase from wild type and mutant with various stabilizers was studied at 30 and 4 °C. Both enzymes were more stable at 4 °C. Among various stabilizers such as dextran (100 kDa, 10 μg/ml), glycerol (0.5%, v/v), PEG 8000 (10 μg/ml) and Tween 80 (0.5%, v/v), Tween 80 provided maximum stabilization at 4 and 30 °C. The mutant showed better stabilization than that of the wild type at both 30 and 4 °C. The loss of activity at 30 °C after 24 h in wild type and mutant in the presence of Tween 80 was only 34 and 32%, respectively, whereas the loss of activity in control of wild type and mutant was 76 and 59%, respectively. After 15 days at 4 °C, the loss of activity in control of wild type and mutant in the presence of Tween 80 was only 15 and 8%, respectively, whereas at 30 °C, the loss of activity in control of wild type and mutant was 49 and 42% respectively. Half-life of the enzyme with Tween 80 was 28.5 and 33.5 h for wild type and mutant, respectively, at 30 °C and 52.1 and 106.6 days for wild type and mutant respectively, at 4 °C

Evaluation of various culture and staining techniques for the detection of extra pulmonary tuberculosis

Background: Though pulmonary tuberculosis form is the commonest presentation, the extra pulmonary tuberculosis (EPTB) is also an important emerging clinical problem. The objective of the current study was to compare two staining techniques, Ziehl-Neelsen (ZN) stain, fluorescent stain and two-culture medium, solid Löwenstein-Jensen (LJ) medium and liquid 7H9 Middle brook medium in MGIT (Mycobacterium Growth Indicator Tube) 320 system, for detection of Mycobacterium in clinically suspected patients of EPTB.Methods: A total of 100 clinically suspected cases of EPTB samples from various extrapulmonary sites had been collected. All the specimens were stained with ZN stain and fluorescent stain. The culture were processed after decontamination of specimens with NaOH-NALC method and thereafter inoculated on solid and liquid culture medium.Results: Out of the 100 EPTB specimens, 30 were found positive by any of the above techniques used. Out of 30 positive cases 18 showed positivity by ZN staining while 20 showed positivity by fluorescent staining technique. In two culture methods, 27 isolates were grown by any of the culture system. Out of 27, 22 and 26 specimens showed growth of MTB complex on LJ media and MGIT culture system respectively. In AFB smear positive specimens, the average turnaround time was found to be 8.45 days and 22.5 days in MGIT and LJ medium culture assay respectively. While the turnaround times in AFB smear negative cases, it was 16.5 days and 32.3 days in MGIT and LJ medium culture assay respectively.Conclusions: MGIT was a dependable, highly efficient system for recovery of MTB complex for EPTB specimens in combination with LJ media

Comparative analysis of injection clonidine and injection dexmedetomidine added to injection bupivacaine for spinal anaesthesia in lower abdominal surgeries

Background:Efficacy of sub-arachnoid block can be improved by addition of various adjuvants to local anesthetics. Intrathecal administration of clonidine or dexmedetomidine has improved the quality of spinal anesthesia in terms of longer duration of post-operative analgesia with comparatively lesser side effects. In present study we compared the onset and duration of motor and sensory block, hemodynamic effects, post-operative analgesia and adverse effects of clonidine and dexmedetomidine used intrathecally with bupivacaine.Methods: Present study was conducted in 150 patients (ASA class I and II) undergoing lower abdominal surgeries. Patients were randomly divided into three group’s viz. B, C and D. Group B received bupivacaine (12.5 mg), group C received clonidine (30 µg) with bupivacaine and group D received dexmedetomidine (5 µg) with bupivacaine. Volume of administered drug was set at 3ml in all the groups. The onset time to reach peak sensory and motor block level, regression time to sensory and motor block, hemodynamic changes and side effects if any were assessed and recorded.Results: In our study we observed that there was no significant difference in patient demography and duration of surgical procedure. The time to onset of sensory blockage was similar in all the three groups but time to onset of motor block was shorter in group C and D compared to group B. Total duration of sensory and motor block was significantly higher in group D compared to group C and B. The duration of sensory block in group D was 139.58+14.49, in group C it was 122.46+18.55 and in group B it was 100+13.43 minutes. The duration of motor block in group D was 250.40+27.33, in group C it was 229.28+23.68 and in group B it was 175.64+17.41 minutes.Conclusions: It was concluded that though both clonidine and dexmedetomidine prolonged duration of sensory and motor block of Bupivacaine, Dexmedetomidine is better in terms of longer duration of action.

Targeting NSP-13 protein of SARS CoV-2 with selected natural compounds: An in-silico approach

SARS-CoV-2 swiftly spread in Wuhan, China, leading to a pandemic crisis worldwide. Genome sequence analysis of this virus revealed a close analogy with its closely related strains, SARS-COV and MERS-COV. In the case of SARS-CoV-2, Nonstructural protein 13 (NSP13), also known as helicase, has been identified as a target for reducing the severity of infection due to its high sequence conservation and essential role in viral replication. NSP13 helicase structure in SARS-CoV-2 differs only by one amino acid from the SARS-CoV helicase structure. Targeting NSP13 with natural compounds holds significant potential for developing safe and effective antiviral therapies utilizing advanced computational approaches. The properties of 8 different natural compounds, i.e. Imidazole, Pyrrole, Tropolone, Benzotriazole, Imidazodiazepine, Phenothiazine, Acridone and Bananin were screened by applying Lipinski’s rule of five, ADME (absorption, distribution, metabolism, and excretion) properties, and Radar plots to discover their drug efficacy at a target site, safety, and absorption. Docking studies confirmed Bananin with a binding affinity of -7 kcal/mol as a potential inhibitor of NSP13 of SARS-CoV-2 with better pharmacokinetics, drug likeliness, and oral bioavailability. Based on the in silico study, it is suggested that Bananin shows promising effects against NSP13 protein, forming a maximum number of hydrogen bonds exhibiting higher binding affinity. This stronger affinity indicates a stronger interaction between the compound and its target, potentially leading to enhanced biological activity and therapeutic efficacy. This novel study has unlocked the door for a prospective SARS-CoV-2 inhibition strategy and developing antiviral interventions targeting NSP13 based on molecular docking.

Colistin, the last resort antibiotic: challenges in the implementation of its routine susceptibility testing

Background: colistin has become a critical antibiotic for lifethreatening multidrug resistance Gram-negative infections, particularly carbapenemase-producing bacteria. Detecting colistin resistance in routine microbiology laboratories is crucial for combating these fatal infections poses a challenge. Especially in developing countries, there is a need for a cost-effective, rapid, and user-friendly diagnostic method. Objective: implementing the various available methods for colistin testing is a significant challenge in resource-limited settings due to logistic difficulties and the need for technical expertise. Materials and Methods: this study shares experiences and insights gained while implementing in-vitro colistin susceptibility testing in a high-load bacteriology laboratory of a tertiary care center in Delhi, India. The following test methods for colistin susceptibility testing were incorporated in the routine antimicrobial susceptibility testing of our laboratory: Colistin Agar Test, Colistin Broth Disk Elution Test, Broth Microdilution susceptibility testing. Results: inconsistent growth patterns were observed in the colistin agar dilution Minimum Inhibitory Concentration (MIC) method, which could be resolved only after the preparation of fresh plates containing that specific concentration of colistin. The contamination issue of plates on use over a few days was addressed by pouring agar containing various concentrations of colistin in cottonplugged glass tubes. In the colistin broth disk elution test, due to the non-availability of screw-capped 10 mL glass tubes, MacCornety bottles (30 mL) were used. Subcultures were performed from the turbid wells to rule out the growth of contaminants when encountering discordant MIC values or skipped wells on the colistin broth microdilution test. Conclusions: despite several technical issues in in-vitro colistin susceptibility testing, we have successfully implemented it in our laboratory. Our experiences can offer guidance to laboratories that are still in the process of implementing it

Impact of patient positioning on radiotherapy dose distribution: An assessment in parotid tumor

Purpose: We intended to study the impact of patient positioning on the dose distribution within target volume and organs at risk in patients with parotid malignancies treated with 3D conformal radiotherapy (3D-CRT) with photon wedge pair (WP) or intensity modulated radiotherapy (IMRT).Methods: Three patients with a non-Hodgkin’s lymphoma of the right parotid gland were consecutively immobilized using thermoplastic cast in 2 positions: supine with head in neutral position (HN) and with head turned 90° to the left side (HT). Images for treatment planning purpose were acquired in both positions. For both positions, photon WP plans and 5 field IMRT plans were generated, after contouring clinical target volume (CTV), planning target volume (PTV= CTV + 5 mm margin) and organs at risk (OAR). All plans were evaluated for target coverage and dose to OARs.Results: Both CTV and PTV were apparently larger in HN compared with HT (31.76±8.89 cc, 30.31±7.83 cc and 62.49±19.01 cc, 58.89±15.33 cc) respectively. The CI value for PTV was slightly better for HT compared to HN position in both the WP and IMRT plans. The homogeneity was comparable in both the head positions in case of WP plan. The mean HI of PTV was increased in case of IMRT plan at HT versus HN position (1.108 vs. 1.097). A change in head position from HN to HT with wedge pair plan resulted in a reduction of brainstem Dmax and Dmean. Lesser dose was observed in HN position for contralateral parotid. A difference of 0.9 Gy in the average Dmax to spinal cord was seen. The values of Dmean to mandible, oral cavity, ipsilateral and contralateral cochlea were higher in the HT position. A change in head position from HN to HT with IMRT plan resulted in a dose reduction in average Dmax to brainstem. The spinal cord Dmax increased at the HT position by 1.2 Gy. The dose to contralateral parotid and cochlea was comparable in both the positions. However, the Dmean to oral cavity was reduced at HT position. Whereas for IMRT versus wedge pair plan at head neutral position average Dmean to the contralateral parotid was reduced with the IMRT plan. A considerable reduction in Dmax to spinal cord and Dmean to ipsilateral cochlea was observed. A slight increase in average Dmax to brainstem and was observed with the IMRT plan. The doses to the remaining OARs were lesser in case of IMRT plan. For IMRT versus wedge pair plan at head tilt position slight increase in average Dmax to brainstem was observed in case of IMRT plan. A considerable reduction in Dmax to spinal cord and Dmean to ipsilateral cochlea was observed. The doses to the remaining OARs were reduced with IMRT plan.Conclusion: Change in head position from neutral to 90° contralateral tilt for wedge pair plan in parotid tumor may considerably reduce dose to the brainstem and spinal cord with a modest increase in dose to mandible, oral cavity, contralateral parotid, and bilateral cochlea. The alteration in head position has minimal impact on IMRT planning