A loop-mediated isothermal amplification assay for the diagnosis of pulmonary tuberculosis.

Quantitated Mycobacterium tuberculosis (M.tb) H37Rv DNA was used to analyse the sensitivity and the specificity was assessed using DNA isolated from the reference strain H37Rv, 12 nontuberculous mycobacterium (NTM) species and five nonmycobacterium species. Furthermore, performance of the assay was evaluated on the sputum samples and compared with smear microscopy, culture and PCR. mpt64 (also called mpb64 or Rv1980c) loop-mediated isothermal amplification (LAMP) successfully detected 1 pg DNA within 40 min and successfully rejected NTMs and other bacterial species tested. It specifically detected all the 119 confirmed TB cases and 100 of the 104 control cases. The resulting sensitivity and specificity of LAMP assay was found to be 100% (95% CI: 96·79-100%) and 96·15% (95% CI; 90·44-98·94%) respectively. SIGNIFICANCE AND IMPACT OF THE STUDY: Loop-mediated isothermal amplification (LAMP) is a technique for isothermal DNA amplification suitable for cost-limited settings as it prevents the use of sophisticated instruments. Using mpt64 antigenic protein gene, we developed a LAMP assay especially for organisms of the M. tuberculosis complex. mpt64 LAMP assay showed 100% sensitivity and detected all the bacteriologically and clinically positive TB cases not detected by smear, culture or PCR methods.

Sodium percarbonate as a novel intracoronal bleaching agent: assessment of the associated risk of cervical root resorption.

AIM: To evaluate the extraradicular peroxide release from sodium percarbonate compared to sodium perborate as an intracoronal bleaching agent. METHODOLOGY: Sixty mandibular single-rooted premolars with intact CEJ were selected. After root filling, gutta-percha was removed 4 mm apical to CEJ and 2 mm of GIC was condensed over the root filling. Intracoronal bleaching agents were placed into six groups of teeth (n = 10): sodium perborate with distilled water (SPW); sodium percarbonate with distilled water (SPCW); sodium perborate with 30% hydrogen peroxide (SPHP); sodium percarbonate with 30% hydrogen peroxide (SPCHP); 30% hydrogen peroxide as positive control (HP) and distilled water as negative control (CL). The teeth were then mounted in vials filled with distilled water, kept in an incubator and taken out at 1, 3 and 6 days for spectrophotometric analysis. Extraradicular peroxide release was quantified by the ferrothiocyanate method. Statistical analysis was undertaken with one-way anova and Scheffe post hoc tests. RESULTS: The greatest peroxide release occurred in the HP group, followed by the SPCHP and SPHP groups, and then by the SPCW and SPW groups. Intergroup comparison revealed that there was no significant difference in peroxide release among the groups SPCW and SPW on days 1, 3 and 6 (P > 0.05). Similarly, no significant difference was found between the SPCHP and SPHP treated groups on days 1, 3 and 6 (P > 0.05). CONCLUSION: Extraradicular peroxide release from sodium percarbonate was comparable to that of sodium perborate, as the differences were not significant.

Understanding the effect of irradiation temperature on microstructural evolution of 20MnMoNi55 steel.

In this study, the effect of irradiation temperature on microstructural evolution of Indian RPV steel is reported. This study, by virtue of helium ion irradiation at 77, 300 and 573 K, could bring out the effect of the irradiation induced defects on microstructural and mechanical property changes at different stages of their existence starting from the state of cascade damage till the point of their free migration. Irradiation experiments were performed with varying ion energies to achieve nearly uniform irradiation damage of 0.05, 0.2 and 3 dpa in a ~ 300 nm wide region. Irradiated samples were characterized using GIXRD, PAS, TEM and nanoindentation. Unirradiated samples showed predominant presence of a combination of di- and tri-vacancy type of defects. Most of the dislocations present in unirradiated samples were screw dislocations, while mixed type was noticed upon irradiation irrespective of the irradiation temperature. PAS study showed formation of distinct defect types at different irradiation temperatures. TEM study confirmed formation of dislocation loops and defect clusters on irradiation. Higher irradiation temperatures resulted in the extension of the width of the damage region owing to increased migration of defects.

Cytotoxic and genotoxic effects of mercury in house fly Musca domestica (Diptera: Muscidae).

Mercury, one of the most widely diffused and hazardous environmental contaminants, induces oxidative stress in organisms, which ultimately leads to genotoxicity and cytotoxicity. House fly Musca domestica L. was used as a model for assaying the genotoxic potential of mercury with the help of micronucleus assay, chromosomal aberration assay as end points and cytotoxicity by assaying the mitotic index and the extent of tissue damage by trypan blue dye exclusion. Late third instar larvae were exposed to different dietary concentrations of mercury (0.0001 mg/ml- 10 mg/ml) for various time intervals. A dose dependent increase in chromosomal aberrations, micronucleus frequency and mitotic index was observed. Micronucleus frequency increases with time while mitotic index decreases showing decreasing rate of cell proliferation due to an increase in cell death. Trypan blue staining gives the visual manifestation of cytotoxicity at higher concentrations of mercury (1 mg/ml- 10mg/ml). The present study suggests that the house fly model may be used to assay the genotoxicity and cytotoxicity of mercury as well as other environmental pollutants.

Seasonal genetic variation in house fly populations, Musca domestica (Diptera: Muscidae).

Seasonal genetic variation was assessed in the common house fly Musca domestica. Allozymes at six gene enzyme system viz., ACPH, EST, G6PD, ME, AO and XDH were analyzed. Fourteen loci with twenty seven alleles were unraveled. The genetic variations were found to be affected to a great extent by environmental influence .F statistics has been used to calculate genetic variation which revealed that very little genetic variation has occurred among the house fly populations analyzed in the present study. Further, except ACPH-2, G6PD-2 and XDH-1 all the other loci show inbreeding (Fis>Fst). Thus it appears that the house fly populations analyzed are characterized by high level of inbreeding. Nei's genetic identity (I) and distance (D) values reveal a close similarity between summer and rainy season collections.

Maximizing the native concentration and shelf life of protein: a multiobjective optimization to reduce aggregation.

A multiobjective optimization was performed to maximize native protein concentration and shelf life of ASD, using artificial neural network (ANN) and genetic algorithm (GA). Optimum pH, storage temperature, concentration of protein, and protein stabilizers (Glycerol, NaCl) were determined satisfying the twin objective: maximum relative area of the dimer peak (native state) after 48 h of storage, and maximum shelf life. The relative area of the dimer peak, obtained from size exclusion chromatography performed as per the central composite design (CCD), and shelf life (obtained as turbidity change) served as training targets for the ANN. The ANN was used to establish mathematical relationship between the inputs and targets (from CCD). GA was then used to optimize the above determinants of aggregation, maximizing the twin objectives of the network. An almost fourfold increase in shelf life (~196 h) was observed at the GA-predicted optimum (protein concentration: 6.49 mg/ml, storage temperature: 20.8 °C, Glycerol: 10.02%, NaCl: 51.65 mM and pH: 8.2). Since no aggregation was observed at the optimum till 48 h, all the protein was found at the dimer position with maximum relative area (64.49). Predictions of the finally adapted network also reveal that storage temperature and solvent glycerol concentration plays key role in deciding the degree of ASD aggregation. This multiobjective optimization strategy was also successfully applied in minimizing the batch culture period and determining optimum combination of medium components required for most economical production of actinomycin D.

ENPP1/PC-1 K121Q polymorphism and genetic susceptibility to type 2 diabetes in North Indians.

Genetic susceptibility may be responsible for high prevalence of type 2 diabetes worldwide. A common missense single nucleotide polymorphism, K121Q in the ectoenzyme nucleotide pyrophosphate phosphodiesterase (ENPP1) gene, has recently been associated with type 2 diabetes in Italian, South Indian, and American populations. The objective of this study was to investigate the possible role of K121Q polymorphism in ENPP1 gene with type 2 diabetes in North Indians. The genotype of the ENPP1/PC-1 K121Q polymorphism was determined using polymerase chain reaction-restriction fragment length polymorphism analysis for 328 T2DM patients and 326 non-diabetic participants. Anthropometric and clinical characteristics (Body mass index (BMI), glucose, total cholesterol, triglyceride, high-density lipoprotein cholesterol (HDL), Creatinine, HbA1c, and insulin levels) were measured using standard protocols. Their Chi-square analyses were used to test the significance differences in genotypic and allelic frequencies. Association studies were undertaken using the t test and logistic regression analyses. Our results revealed there was no significant difference in the genotypic distribution between T2DM patients and control subjects. The KK and KQ genotype frequencies were similar in T2DM cases and controls (60.7 and 39.3% in T2DM and 59.8 and 40.2% in controls). No subjects with the QQ genotype were found. Binary logistic regression analysis of data did not show any association of K121Q polymorphism with type 2 diabetes (OR; 0.97, 95% CI; 0.7-1.32, P = 0.82). No significant correlation among the BMI, WHR, BP, TG, TC, HDL-C, LDL-C, Glucose, HbA1c, Creatinine, and insulin indices (HOMA-IR) was observed in the individuals carrying KK and KQ genotypes. In conclusion, our results showed that ENPP1/PC-1 K121Q polymorphism is not associated with type 2 diabetes and related quantitative metabolic traits in North Indian Punjabi population.

Atypical Squamous Cells of Undetermined Significance : A Follow up Study.

BACKGROUND: Atypical squamous cells of undetermined significance (ASCUS) is the term in cervical cytology reporting where the changes are suggestive of a sqamous extraepithelial lesion but lack criteria for definition interpretation. Its clinical significance is variable. METHODS: A one year study in a zonal hospital evaluating all cervical smears received for cytology by two pathologists independently and subsequently reported as per the Bethesda guidelines. The patients were folloed up for six months. RESULT: A total of 24 (4%) smears were reported as ASCUS out of 604 examined. Of these, 15 were followed up for one year. Mean age was 39 years and 60% were premenopausal. After six months, 11 cases were reported negative for intraepithelial lesion or malignancy and four showed ASCUS on repeat pap smear. CONCLUSION: ASCUS shows a repression rate of 73%. Colpscopic evaluation should be done only if ASCUS persists. Diligent follow up and biopsy, when indicated is a must.

Improving patient care with shorter hospital admissions.

Prolonged hospital admissions frequently overshadow the experience of gynaecological cancer patients. In East Kent, we identified obstacles in the inpatient journey through the hospital by performing a patient pathway mapping audit of admissions in May/June 2006 and 2007. We achieved substantial reductions in the length of stay for major gynaecological oncology surgery. The number of the lead consultant's patients with a total duration of admission of no more than 5 days increased from one of 28 in Q1 2006 (3.6%), to 17 of 28 (60.7%) in Q4 2007. Identified problems prolonging the length of stay included admission the day before surgery, prolonged use of i.v. antibiotics and late referrals to social services. Early discharge planning involving staff and patients as active participants has the greatest impact on reduction of duration of hospital admission. A reduction in duration of hospital admissions is associated with an increase in the rate of readmissions.

Cloning, characterization and expression of the chitinase gene of Enterobacter sp. NRG4.

A chitinase producing bacterium Enterobacter sp. NRG4, previously isolated in our laboratory, has been reported to have a wide range of applications such as anti-fungal activity, generation of fungal protoplasts and production of chitobiose and N-acetyl D-glucosamine from swollen chitin. In this paper, the gene coding for Enterobacter chitinase has been cloned and expressed in Escherichia coli BL21(DE3). The structural portion of the chitinase gene comprised of 1686 bp. The deduced amino acid sequence of chitinase has high degree of homology (99.0%) with chitinase from Serratia marcescens. The recombinant chitinase was purified to near homogeneity using His-Tag affinity chromatography. The purified recombinant chitinase had a specific activity of 2041.6 U mg(-1). It exhibited similar properties pH and temperature optima of 5.5 and 45°C respectively as that of native chitinase. Using swollen chitin as a substrate, the K(m), k(cat) and catalytic efficiency (k(cat)/K(m)) values of recombinant chitinase were found to be 1.27 mg ml(-1), 0.69 s(-1) and 0.54 s(-1)M(-1) respectively. Like native chitinase, the recombinant chitinase produced medicinally important N-acetyl D-glucosamine and chitobiose from swollen chitin and also inhibited the growth of many fungi.

Epileptic seizure anticipation and localisation of epileptogenic region using EEG signals.

Electric activity of brain gets disturbed prior to epileptic seizure onset. Early prediction of an upcoming seizure can help to increase effectiveness of antiepileptic drugs. The scalp electroencephalogram signals contain information about the dynamics of brain and have been used to predict an upcoming seizure and localise its zone. The objective of this paper is to localise the epileptogenic region and predict an upcoming seizure at the earliest. To localise epileptogenic region, Electroencephalogram signals are categorised into four regions of brain (Frontal, Temporal, Parietal and Central). For each signal seventy-two (72) parameters in frequency domain have been extracted by using ten minute non overlapping window. Four prominent ratio parameters, γ1/γ5, γ3/γ1, θ/γ2 and γ4/θ have been identified as best parameters based on relative fisher score. Zone 2 shows the highest change in all the parameters as compared to the other zones. So, temporal region is identified as the epileptogenic region in this work. For prediction of the epileptic seizure machine learning algorithm artificial neural network (ANN) is proposed. The proposed machine learning algorithm has an accuracy of 92.3%, sensitivity of 100% and specificity of 83.3%.

Phytochemical analysis and mode of action against Candida glabrata of Paeonia emodi extracts.

In the present study, we have evaluated the antifungal activity of the seed, root and leaf of Paeonia emodi (commonly known as Himalayan peony) in four common solvents (acetone, chloroform, methanol and water) against six fungal strains. The methanolic seed extract (MSE) showed promising antifungal activity against Candida albicans (6.25mg/mL), Candida glabrata (3.12mg/mL) and Candida parapsilosis (12.50mg/mL) among all the fungal strains tested. Combination of the MSE with the well-known commercial antifungal drugs amphotericin B (Amp B), nystatin (NYS) and fluconazole (FLC) resulted in the killing of C. glabrata at non-inhibitory concentrations, i.e., 0.35µg/mL for Amp B, 0.55µg/mL for NYS and 1.19µg/mL for FLC. Notably, MSE caused cell wall damage of C. glabrata cells, as confirmed by confocal microscopy, flowcytometry and scanning electron microscopy (SEM). The MSE was fractionated by thin layer chromatography (TLC). TLC-bioautography was used to determine the active compounds present in the MSE. Column chromatography was used to separate the potential active compounds from the MSE. Furthermore, gas chromatography-mass spectrometry (GC-MS) andfourier-transform infrared spectroscopy (FTIR) were used to identify the phytocomponents of the MSE. These experiments revealed 13-docosenamide/9-octadecenamide/trans-13-docosenamide (89.70%) as being the predominant compound using a chloroform/methanol solvent system for the separation. Interestingly, the MSE also exhibited less significant cytotoxicity at the minimum inhibitory concentration (MIC) against mammalian cells (HeLa and HEK293). This study suggests that the MSE of P. emodi can be used for the treatment of C. glabrata infection.

Synthesis and characterization of monodispersed water dispersible Fe3O4 nanoparticles and in vitro studies on human breast carcinoma cell line under hyperthermia condition.

Monodispersed Fe3O4 magnetic nanoparticles (MNPs) having size of 7 nm have been prepared from iron oleate and made water dispersible by functionalization for biomedical applications. Three different reactions employing thioglycolic acid, aspartic acid and aminophosphonate were performed on oleic acid coated Fe3O4. In order to achieve a control on particle size, the pristine nanoparticles were heated in presence of ferric oleate which led to increase in size from 7 to 11 nm. Reaction parameters such as rate of heating, reaction temperature and duration of heating have been studied. Shape of particles was found to change from spherical to cuboid. The cuboid shape in turn enhances magneto-crystalline anisotropy (Ku). Heating efficacy of these nanoparticles for hyperthermia was also evaluated for different shapes and sizes. We demonstrate heat generation from these MNPs for hyperthermia application under alternating current (AC) magnetic field and optimized heating efficiency by controlling morphology of particles. We have also studied intra-cellular uptake and localization of nanoparticles and cytotoxicity under AC magnetic field in human breast carcinoma cell line.

Warfarin related acute kidney injury: A case report.

Warfarin is an oral anticoagulant used extensively in clinical practice; However, its side-effect of causing renal damage has been recently detected. The mechanism leading to renal damage is glomerular hemorrhage and red blood cell tubular casts prothrombin time. Recently, it was found that warfarin causes renal damage in patients with chronic kidney disease and is also associated with progression of renal disease. Warfarin causing acute kidney injury in patients with normal renal function is a rare manifestation. It is important to be aware of this condition as its innocuous presence can lead to chronic kidney disease if not corrected in time. Further studies have also found that novel oral anticoagulants such as dabigatran also cause a similar syndrome and hence a new term called anticoagulant-related nephropathy is now in vogue.

How Difficult Is It To Implement A Competency-Based Medical Education In India: The Next Steps?

Background:Medical Council of India is in the revamping mode to align the medical curriculum to the emerging health care needs of the society, by introducing Competency-Based Medical Education (CBME). This new medical curriculum intends toimpartthecompetencies that are needed to produce medical graduates who would function efficiently as Physicians of first contact or basic doctorsin the community. The ECE program would enhance the knowledge, skills, and attitude of the medical graduates if it is implemented. We conducted this study to assess the challengescurrently facedby faculties for the implementation of this new curriculum.Material and Methodology:It is a cross-sectional study conducted at NC Medical College, Israna during the study period was July 2021 to November 2021. The study team regularly scrutinized the data collection process and met periodically to review the study conduct and computing of data. At the end of the study period, the consolidated data were analyzedusing IBM SPSS version 22.Result:A total of 50 teaching faculties, we acknowledged 48questionnairesthat were complete. 96% faculties showed positive and opportunistic be haviourtowards the new curriculum of CBME with constructive ideas but 4% faculties declined.Conclusion:CBME is theparadigm shift in the teaching and learning approach of medical education. Mostof the medical facultymembersare aware of the need and have attained a positive attitude for theapplication of CBME. However significant barriers do exist in the form of manpower, new time mapping and resources which need to be addressed.

Liquid like nucleation in free-standing nanoscale films.

The concept of a critical nucleus size (r*) is of pivotal importance in phase transformations involving nucleation and growth. The current investigation pertains to crystallization in nanoscale thin films and study of the same using high resolution lattice fringe imaging (HRLFI) and finite element simulations. Using the CuZrAl bulk metallic glass system as a model system for this study, we demonstrate a liquid like nucleation behaviour in nanoscale free-standing films upon heating. The r* for the formation of the Cu10Zr7 phase in thin films (of decreasing thickness) approaches that of the r* for the formation of the crystal from a liquid (i.e.). Working in the nucleation dominant regime, we introduce the concept of 'depth sensitive lattice fringe imaging'. The thickness of the film is determined by electron energy loss spectroscopy and the strain energy of the system is computed using finite element computations.

Suppressive effect of 3-methylcholanthrene on phagocytic activity of mouse peritoneal macrophages for Torulopsis glabrata.

The effect of 3-methylcholanthrene (MCA) on the phagocytic activity of mouse peritoneal macrophages for Torulopsis glabrata was investigated. Macrophages were maintained in glass scintillation vials or on cover slips in Leighton tubes with the use of Hanks' balanced salt solution plus 30% horse serum. Graded amounts of MCA were incorporated into the medium and the macrophages were parasitized with viable cells of T. glabrata. Macrophages from C3H mice, a strain highly susceptible to MCA carcinogenesis, were more prone to the suppressive effect of MCA than were the macrophages from CFW mice, a relatively resistant strain. Significant suppressive effect on phagocytosis of macrophages from C3H mice was observed with 5 micrograms MCA/ml, whereas up to 50 micrograms MCA/ml did not alter the phagocytic activity of CFW macrophages. However, 100 micrograms MCA/ml also suppressed the phagocytosis of CFW macrophages. Suppression in phagocytosis of C3H macrophages was observed after 6 hours' exposure to MCA, whereas a similar effect on CFW macrophages was seen after 12 hours. Treatment with 100 micrograms MCA/ml imparied the fungicidal activity of both C3H and CFW macrophages. These results indicate a correlation between the suppressive effect of MCA on macrophage activity and the strain susceptibility of mice to chemical carcinogenesis.

Evaluation and comparison of transverse and impact strength of different high strength denture base resins.

AIM: The present study was undertaken to evaluate and compare the impact strength and transverse strength of the high-impact denture base materials. A conventional heat polymerized acrylic resin was used as a control. MATERIALS AND METHODS: The entire experiment was divided into four main groups with twenty specimens each according to denture base material selected Trevalon, Trevalon Hi, DPI Tuff and Metrocryl Hi. These groups were further subgrouped into the two parameters selected, impact strength and flexural strength with ten specimens each. These specimens were then subjected to transverse bend tests with the help of Lloyds instrument using a three point bend principle. Impact tests were undertaken using an Izod-Charpy digital impact tester. RESULTS: This study was analyzed with one-way analysis of variance using Fisher f-test and Bonferroni t-test. There was a significant improvement in the impact strength of high-impact denture base resins as compared to control (Trevalon). However, in terms of transverse bend tests, only DPI Tuff showed higher transverse strength in comparison to control. Trevalon Hi and Metrocryl Hi showed a decrease in transverse strength. CONCLUSIONS: Within the limits of this in vitro study, (1) There is a definite increase in impact strength due to the incorporation of butadiene styrene rubber in this high strength denture base materials as compared to Trevalon used as a control. (2) Further investigations are required to prevent the unduly decrease of transverse strength. (3) It was the limitation of the study that the exact composition of the high-impact resins was not disclosed by the manufacturer that would have helped in better understanding of their behavior.

Long-term visual outcomes in intraocular retinoblastoma with eye preservation.

PURPOSE: Inconsistent data exist on long-term visual outcomes in survivors of retinoblastoma. No studies have been reported on role of ocular coherence tomography (OCT) in predicting visual acuity. We assessed visual acuity in patients with retinoblastoma treated at our center in whom affected eyes were preserved. METHODS: Patients who had completed a 2-year follow-up and were more than 5 years of age at assessment were included. Clinical data were obtained from database and factors predicting visual acuity were analyzed. OCT was performed in these patients to assess central macular thickness (CMT). RESULTS: Visual outcomes were assessed in 45 eyes of 43 patients, of which 38 (88 %) had bilateral retinoblastoma. The median age at diagnosis was 12 months. Sixty percent (27/45) had International classification of retinoblastoma group C or D disease with 40 % eyes showing macular lesions. The far visual acuity was better than 6/12 in 53 % (24/45), 6/12 to 6/60 in 40 % (18/45) and 6/60 in 7 % (6/60). Macular location and International classification of retinoblastoma predicted poor vision (p = 0.06 and 0.07, respectively). CMT was less than 200 µm in 3 of 36 eyes (8 %) and 1 eye showed epiretinal membrane. Radiotherapy was associated with foveal thinning (p = 0.003). Two of 3 eyes with foveal thinning had a vision of 6/60. CONCLUSIONS: Good visual outcomes were observed in half of retinoblastoma patients treated with eye preservation. Macular location and International classification of retinoblastoma group C and D predicted poor visual acuity, while previous radiotherapy predicted foveal thinning, which was associated with poor visual acuity.

Self-assembling N-(9-Fluorenylmethoxycarbonyl)-l-Phenylalanine hydrogel as novel drug carrier.

Supramolecular hydrogel as a novel drug carrier was prepared from N-(9-Fluorenylmethoxycarbonyl) (Fmoc) modified l-phenylalanine. Its different properties like stability at different pH, temperature and rheology were evaluated in reference to salicylic acid (SA) as a model drug, entrapped in the supramolecular hydrogel network. The release behaviour of SA drug in supramolecular hydrogel was investigated by UV-vis spectroscopy. The influence of hydrogelator, pH values of the accepting media, temperature and concentration of SA drug on the release behaviour was investigated under static conditions. The results indicated that the release rate of SA in the supramolecular hydrogels was slightly retarded with an increase of the hydrogelator concentration. Also, the release rates of SA increased with an increase of temperature and its concentration. Furthermore, the release behaviour of SA was found to be different at various pH values in buffers. The study of the release kinetics indicated that the release behaviour of SA from the carrier was in accord with the Peppas model and the diffusion controlled mechanism involved in the Fickian model.