Potential Factors Related to Waist Circumference in Urban South Indian Children.

Objectives: To identify important factors (linked to lifestyle, eating and sedentary behaviors) relating to waist circumference among urban South Indian children aged 3 to 16 years. Design: Cross sectional. Setting: Urban schools of Bangalore, from August 2008 to January 2010. Participants: 8444 children; 4707 children aged 3-10 years and 3737 children aged 10-16 years. Methods: Data were collected on the frequency of consumption of certain foods, physical activity patterns, sedentary habits at home, sleep duration and behaviors such as habits of snacking, skipping breakfast, eating in front of television and frequency of eating out. Simple linear regression analysis of waist circumference on various food items, physical activity, behavior and parental BMI were performed. A path model was developed to R E S E A R C H P A P E R identify potential causal pathways to increase in waist circumference. Results: Increased consumption of bakery items, non vegetarian foods, increased television viewing, decreased sleep duration, eating while watching television, snacking between meals, family meals, skipping breakfast (in older children), and parental BMI were found to be related to waist circumference. Older children possibly underreported their intake of “unhealthy” foods, but not behaviors. Conclusions: This study identified potential behaviors related to waist circumference in urban school children in India. Longitudinal studies with better measures of morbidity and adiposity are warranted in order to derive casual relationships between various determinants and waist circumference.

Waist Circumference and Waist for Height Percentiles in Urban South Indian Children Aged 3-16 Years.

Objectives: To develop age and gender specific waist circumference references for urban Indian children aged 3 -16 years. Design: Cross-sectional study. Setting: Urban preschools and schools of Bangalore. Participants: 9060 children (5172 boys and 3888 girls) in the age group of 3-16 years. Methods: Weight, height, and waist circumference were measured using standard anthropometric methodology. Percentiles for waist circumference and Waist/height ratio (W/Ht) for each age and gender were constructed and smoothed using the LMS method. Results: Mean waist circumference increased with age for both girls and boys. The upper end of curve in boys continued to increase, whereas in the girls it tended to plateau at 14 years. The waist circumference of the Indian children from the present study was higher than age and sex matched European children. The proportion of children with W/Ht ratio greater than 0.5 decreased as their age increased. Conclusions: These curves represent the first waist and waist height ratio percentiles for Indian children and could be used as reference values for urban Indian children. We suggest that for a start, the 75th percentile of waist circumference from this study be used as an “action point” for Indian children to identify obesity (as a tautological argument), while retaining the cut-off of 0.5 for the W/Ht ratio; however this underlines the need to derive biologically rational cut-offs that would relate to different levels of risk for adult cardiovascular disease.

A comparative evaluation of the fracture resistance of endodontically treated teeth with compromised intra radicular tooth structure using three different post system.

Intra-radicular loss of tooth structure in endodontically treated teeth poses a challenge. Available methods for treatment are cast post-core, intra-radicular resin reinforcement using composite resin followed by placement of prefabricated metal/fibre post (glass or carbon).This study is an attempt to investigate the validity of treatment of such teeth using above methods and evaluate which post system is best suited for rehabilitation. Thirtysix endodontically treated anterior teeth were prepared by uniformly removing intra-radicular tooth structure from buccal, lingual, mesial & distal surfaces such that only 0.75mm-1mm remained. Twelve teeth were subsequently restored with cast metal post & core, 12 with intra-radicular resin reinforcement followed by prefabricated titanium post (Luminex post system) & 12 with intra-radicular resin reinforcement followed by glass fibre post (Luscent Anchor post system). Statistical analysis used was t-test. There was no statistically significant difference between the 3 post systems, but it was observed that cast post & cores caused more apical & oblique fractures, rendering the teeth unrestorable. Teeth restored with intra-radicular resin reinforcement & placement of titanium or glass fibre post failed with root fractures limited to the coronal aspect along with dislodgement of post. Intra-radicular resin reinforcement offers advantages like preventing the metal display of the post through the thin dentinal wall, reinforcement of the thin walled teeth & comparable fracture resistance to cast post and core.

Evaluation of suitable solvents for testing the anti-proliferative activity of triclosan - a hydrophobic drug in cell culture.

Triclosan, a broad spectrum antibiotic is currently being evaluated for its anti-cancer property. Though several solvents are available to dissolve lipophilic (hydrophobic) drugs, solubility and toxicity aspects pose a challenge, when combined with the cell culture medium. In this paper, we present a simple approach based on physico-chemical and biologic criteria to choose a suitable solubilizing agent to study the anti-proliferative property of triclosan in breast cancer cell line MCF-7. Triclosan was dissolved in five different solvents viz. DMSO, absolute ethanol, 1 N NaOH, 55% polyethylene glycol + 45% ethanol mixture (PEM) and acetone and diluted with the culture medium (1 mg/ml). Although triclosan dissolved completely in all five solvents, on dilution with culture medium, turbidity was observed in DMSO, 1 N NaOH and ethanol. Cell viability was 95.23% in 10 ml of acetone, when compared with 49.45% at the same volume of PEM. This non-toxic nature of acetone was supported by DNA fragmentation analysis and phase contrast microscopy. A significant decrease in cancer cell proliferation at 100 mg/ml of acetone-solubilized triclosan, compared with 100 mg/ml of PEM-solubilized triclosan (p<0.05) indicated stronger anti-proliferative effect and greater drug-sensitivity of triclosan when solubilized in acetone. Results showed that acetone-solubilized triclosan was suitable for anti-cancer investigations in cultured MCF-7 cells.

DNA sequencing by Microseq kit targeting 16S rRNA gene for species level identification of mycobacteria.

BACKGROUND & OBJECTIVES: Identification of mycobacteria to the species level is of therapeutic significance. Conventional methods are laborious and time consuming so we did 16S rRNA sequencing using a commercial MicroSeq sequencing kit, which includes DNA sequencing with software package for identification and phylogenetic analysis of clinical mycobacterial isolates. METHODS: A total of 47 mycobacteria were tested by both conventional and genotypic method using commercially available MicroSeq 500 amplification kit assay. The identification was determined by comparing the 500 bp amplified product of 16S rDNA sequence to the MicroSeq database. RESULTS: The phenotypic identification was concordant with genotypic identification in 33 (70.2%) isolates of 14 Mycobacterium tuberculosis, 11 M. fortuitum, 7 M. abscessus and 1 M. duvalii. For the discrepant isolates, identification was possible only by DNA sequencing in 14 (29.7%) isolates. The 14 discrepant isolates were 5 M. farcinogenes, 3 M. genavense, 2 M. species. nov and 1 each of M. fortuitum, M. immuogenum, M. simiae and M. wolinskyi. Of these, five were uncommon species that were difficult to identify by phenotypic method. INTERPRETATION & CONCLUSION: The MicroSeq DNA sequencing is an excellent tool for species identification of mycobacteria, which reduces the turn around time, makes repeat analysis easy as compared to phenotypic identification specially for mycobacterial isolates with ambiguous biochemical profiles.

Association of mycobacteria with Eales' disease.

BACKGROUND & OBJECTIVES: Eales' disease is an idiopathic disease resulting in retinal neovascularization, recurrent haemorrhages, with or without retinal detachment predominantly affecting healthy young males (97.6%) in the Indian subcontinent. Inspite of several studies, the aetiology of Eales' disease is not clear. The isolation of Mycobacterium fortuitum from the aqueous humour of a patient with classical Eales' disease, led us to hypothesize that rapid growing nontuberculous mycobacteria (RGNTM), particularly M. fortuitum and M. chelonae could be associated with Eales' disease. We therefore undertook this study to detect DNA of these RGNTM and also of M. tuberculosis in vitreous fluids (VFs) from patients with Eales' disease and non-Eales' disease. METHODS: We developed and optimized seminested polymerase chain reactions (SnPCRs) to detect DNAs of M. fortuitum and M. chelonae on archival ERMs (33) and VFs (19) of Eales' and control patients along with conventional mycobacteriological investigations. RESULTS: In the retrospective study, 70 per cent ERM samples were positive for one or more Mycobacterium spp. tested by snPCR. M. fortuitum and M. chelonae were isolated from two VFs, which were also positive by sn PCR in the prospective study. Statistical evaluation of the results of both retrospective and prospective investigations showed a statistically significant association of Mycobacterium spp. with Eales' disease. INTERPRETATION & CONCLUSION: The results of the present study suggested the involvement of Mycobacterium spp. in the aetiopathogenesis of Eales' disease. Further studies on a larger sample will be required to confirm these findings.

In-vitro susceptibility testing by agar dilution method to determine the minimum inhibitory concentrations of amphotericin B, fluconazole and ketoconazole against ocular fungal isolates.

PURPOSE: To standardize in-vitro antifungal susceptibility testing by agar dilution method to find out the minimum inhibitory concentration (MIC) of amphotericin B, fluconazole and ketoconazole on ocular fungal isolates. METHODS: A total of 180 ocular fungal isolates (130 filamentous fungi and 50 yeasts) were included. The antifungal drugs such as amphotericin B (0.0625-8 microg/mL), fluconazole (0.2-819.6 microg/mL) and ketoconazole (0.025-6.4 microg/mL) were incorporated in doubling dilutions in the yeast nitrogen base medium. The MIC was determined as the lowest concentration of the antifungal drug preventing growth of macroscopically visible colonies on drug containing plates when there was visible growth on the drug-free control plates. RESULTS: All 50 ocular isolates of yeast were susceptible to amphotericin B, while two (4%) and five (10%) strains were resistant to fluconazole and ketoconazole respectively. Of the 130 filamentous fungi tested, six (4.6%) were resistant to amphotericin B, 49 (37.7%) and 10 (7.6%) were resistant to fluconazole and ketoconazole respectively. Percentile 50 (MIC 50) and Percentile 90 (MIC 90) for all the three antifungal agents were calculated. Aspergillus niger, Aspergillus terreus and Candida krusei were found to be resistant to fluconazole and ketoconazole. CONCLUSION: This technique was found to be reliable, cost effective and easy to perform with consistent results.

Polyclonal antibody-mediated mitotic inhibition in Chinese hamster ovary cells.

The nucleus of a mammalian cell undergoes profound reorganization when the cell enters mitosis and a number of proteins involved at various levels of the cell cycle have been characterized. The presence of mitotic-specific proteins has been reported and their roles are important in understanding the mechanics of cell division. The ability of antibodies to recognize mitotic protein antigens and further inhibit mitosis is potentially valuable in their role as therapeutic and diagnostic agents in cancer therapy. In this study, we have aimed to analyze proteins isolated from mitotic cells of Chinese hamster ovary (CHO) cells and their significant role in inhibiting mitosis. The proteins extracted from mitotic cells were processed and antibodies produced. It was observed that the secondary response that yielded an antiserum of 1:8 titer was predominantly IgG. The antiserum was effective in inhibiting mitosis in CHO cells in culture in a dose-dependent manner. Although inhibition of mitosis was apparent by cell proliferation studies, there was no apparent effect of the antiserum on other cell morphology and culture characteristics. The unique molecular structure of the antibody by which it bivalently binds to a broad array of antigenic epitopes serves as the foundation of its utility. These antibodies, being polyclonal in nature, are targeted against a whole range of proteins; and their multiple epitopes involved in process of cell division might hence mediate recognition or inhibition of function of such proteins in a wholesome manner and thus accomplish inhibition of mitotic progression.

Detection and characterization of mutations in Rifampicin resistant Mycobacterium tuberculosis clinical isolates by DNA sequencing.

Background: Multiple Drug Resistant Tuberculosis (MDR-TB) is increasing because of widespread application and results in selection of mutants resistant to other components of short course chemotherapy. Resistance to Rifampicin can be considered as a surrogate marker for MDR-TB and the target gene for detection of rifampicin resistance is the rpo gene. Aims: To detect and characterize mutations in the rpo B region of Rifampicin resistant isolates of Mycobacterium tuberculosis by automated DNA sequencing. Methods: Absolute concentration method was used to determine the MIC of Rifampicin for 44 M. tuberculosis isolates (21 respiratory, 3 ocular, 3 cerebrospinal fluid and 17 biopsies). Automated DNA sequencing was performed in the ABI 310 Genetic Analyser. Results: Five isolates (2 sputa and one each from bronchoalveolar lavage, lymph node and endometrial biopsies) were rifampicin resistant with MIC greater than 128 mg/ml. Three of the five isolates showed mutations. Two of the isolates had the common missense mutation at codon 531(Ser®Leu), the other isolate showed three insertions and two of them did not show any mutation in the sequenced rpo B region. Conclusions: DNA sequencing technique is a rapid, conclusive and more advantageous technique than the conventional susceptibility testing for detection of rifampicin resistance in terms of the risk involved and time consumption.

Application of PCR-based restriction fragment length polymorphism for the identification of mycobacterial isolates.

BACKGROUND AND OBJECTIVE: Conventional identification of mycobacteria is achieved by standard biochemical tests that are time consuming, laborious and is not always conclusive. This study was thus undertaken to standardize a simple, rapid and cost-effective polymerase chain reaction based restriction fragment length polymorphism (PCR-RFLP) using primers coding for the 16S - 23S rRNA spacer region to identify the mycobacterial isolates to the species level. METHODS: The PCR with primers targeting the 16S-23S rRNA spacer region was standardized using the standard mycobacterial strains and applied on 51 clinical isolates. The PCR amplified products were subjected to RFLP using the restriction enzymes, Hae III, MspI and BstXI. The results obtained were compared with those of conventional biochemical tests. RESULTS: PCR was sensitive to detect 2.5 pg of H37Rv DNA (370 bp for slow grower mycobacteria) and 1.5 pg of M. fortuitum DNA (450 bp for rapid grower mycobacteria). Based on the PCRRFLP products obtained the 51 mycobacterial isolates were classified into 41 slow growers and 10 rapid growers. Among the 41 slow growers, 40 were identified as M. tuberculosis, one as M. xenopi and 10 rapid growers as M. fortuitum. INTERPRETATION AND CONCLUSION: PCR using primers targeting the 16S-23S rRNA spacer region was a reliable tool for rapid identification of mycobacterial isolates into slow and rapid growers within 4 h of isolation and further speciation by PCR-RFLP within 6-8 h.