Insights into the intracellular mechanisms of citronellal in Candida albicans: implications for reactive oxygen species-mediated necrosis, mitochondrial dysfunction, and DNA damage

Abstract INTRODUCTION Citronellal (Cit) possesses antifungal activity and has possible implications for reactive oxygen species (ROS) generation in Candida albicans. In this study, the effects of Cit on ROS generation and the mechanisms by which Cit exerts anti-Candida effects were examined. METHODS A 2′,7′-dichlorodihydrofluorescein diacetate assay was used to assess oxidative damage. Cell necrosis was determined by flow cytometry after FITC-Annexin V staining. Mitochondrial function was studied based on mitochondrial potential, metabolic activity (MTT assay), and phenotypic susceptibility on a non-fermentable carbon source. Membrane intactness and DNA damage were estimated by a propidium iodide (PI) uptake assay and 4',6-diamidino-2-phenylindole (DAPI) staining. RESULTS ROS generation was enhanced in response to Cit, leading to necrosis (2%). Additional hallmarks of cell death in response to Cit, such as mitochondrial membrane depolarization and DNA damage, were also observed. Cit treatment resulted in dysfunctional mitochondria, as evidenced by poor labeling with the mitochondrial membrane potential-sensitive probe rhodamine B, reduced metabolic activity (61.5%), and inhibited growth on a non-fermentable carbon source. Furthermore, Cit induced DNA damage based on DAPI staining. These phenotypes were reinforced by RT-PCR showing differences in gene expression (30-60%) between control and Cit-treated cells. Finally, PI uptake in the presence of sodium azide confirmed non-intact membranes and suggested that Cit activity is independent of the energy status of the cell. CONCLUSIONS Cit possesses dual anticandidal mechanisms, including membrane-disruptive and oxidative damage. Taken together, our data demonstrated that cit could be used as a prominent antifungal drug.

Clinical and biochemical parameters in relation to serum leptin levels in south indian children and adolescents.

Objective. To investigate the relationship between serum leptin and lipid profile in South Indian School children and adolescents, and to evaluate the role of serum leptin in obese, overweight and congenital heart diseased children and adolescents; in South Indian population and its correlation with anthropometric and biochemical parameters. Methods. The study included 185 school going children and adolescents. (52 obese, 49 overweight, 25 congenital heart disease children and adolescents, were compared with 59 normal controls, aged between 10-17 years). Anthropometric variables, lipid profile, fasting serum glucose were analyzed by autoanalyser and serum leptin by ELISA. Results. Serum leptin levels were significantly elevated in obese and overweight children than in control children (36.88±18.60ng/mL, 20.64±11.18ng/mL vs 7.97±2.79ng/mL; p value <0.001), and decreased in congenital heart diseased children than in control children (6.20±4.23 ng/mL vs 7.97±2.79ng/mL; p value <0.005). Conclusions. This study provides a good relationship between serum leptin levels and anthropometric and biochemical parameters, such as total cholesterol, triglycerides and LDL-cholesterol. We observed negative correlation between serum leptin and fasting glucose levels and HDL-cholesterol levels were found to be non-significant among the groups. Further studies with large sample size are needed to ascertain the relationship between serum leptin and lipid profile in different groups of children and adolescents.

Increased yield of smear positive pulmonary TB cases by screening patients with > or =2 weeks cough, compared to > or =3 weeks and adequacy of 2 sputum smear examinations for diagnosis.

BACKGROUND: RNTCP recommends examining three sputum smears for AFB from Chest Symptomatics (CSs) with cough of > or =3 weeks for diagnosis of Pulmonary TB (PTB). A previous multi-centric study from Tuberculosis Research centre (TRC) has shown that the yield of sputum positive cases can be increased if duration of cough for screening was reduced to > or =2 weeks. Other studies have shown that two smear examinations are adequate for diagnosis of smear positive PTB . To validate the above findings, a cross sectional multi-centric study was repeated in different settings in five geographical areas in India. METHODS: Three primary and secondary level health facilities with high out-patient attendance were selected from two Tuberculosis Units (TU) in each of the 15 selected districts to screen about 10,000 new adult outpatients from each state. For patients who did not volunteer history of cough, symptoms were elicited using a structured simple questionnaire. All the CSs were referred for sputum examination. RESULTS: A total of 96,787 out-patients were registered. Among them 69,209 (72%) were new adult out-patients. Using > or =2 weeks of cough instead of 3 weeks as the criterion for screening, there was an overall increase of 58% in CS and 23% increase in the detection of smear-positive cases. Among 211 patients, 210 were positive at least by one smear from the initial two specimens. Increase in the work-load if 2 smears were done for patients with cough of > or =2 weeks cough were 2 specimens (i.e. 13 to 15) per day for an adult OPD of 150. CONCLUSION: The yield of sputum positive PTB cases can be improved by screening patients with > or =2 weeks cough and two specimens are adequate for diagnosis.