In-vitro study of the effect of Centella asiatica on cholera toxin production and the gene expression level of ctxA gene in Vibrio cholerae isolates.

ETHNOPHARMACOLOGICAL RELEVANCE: Centella asiatica (L.) Urb or Indian pennywort is a plant of ethnopharmacological relevance, commonly called as Brahmi in South India known for its antimicrobial property in gut and for the treatment of other gut ailments. Natural anti-virulence drugs that disarm pathogens by directly targeting virulence factors or the cell viability and are thus preferred over antibiotics as these drugs impose limited selection pressure for resistance development. In this regard, an in-vitro experimental study was conducted to know the effect of extract of Centella asiatica(L.) Urb. on cholera toxin, gene expression and its vibriocidal effect on five standard strains of Vibrio cholerae; IDH03097 (El Tor variant), N16961 (El Tor), O395 (Classical) as well as five clinical strains (Haitian variant). AIM OF THE STUDY: To study the effect of extract of Centella asiatica on Vibrio cholerae. MATERIALS AND METHODS: Crude extract was prepared from the leaves and stem part of the plant. The vibriocidal concentration was tested at different concentrations of the extract. The amount of cholera toxin released from the strains before and after exposure to the extract of Centella asiatica to Vibrio cholerae was measured using Bead ELISA. ctxA gene expression in the strains before and after exposure to extract of Centella asiatica was measured using quantitative real time PCR. All the above assays were performed with commercially obtained asiaticoside as well. RESULTS: The vibriocidal activity was tested at the different concentration of the extract, where 1g/mL of crude extract and 12.5mg/mL of asiaticoside was found to be vibriocidal. The amount of cholera toxin released before and after the exposure to extract of C. asiatica was measured using Bead ELISA, showing a reduction of 70%, 89% and 93% toxin produced by classical, El Tor and variant respectively. ctxA gene expression before and after exposure to extract of Centella asiatica as well as asiaticoside was measured using qRT-PCR. We found a decrease in expression of ctxA gene transcription by 6.19 fold in classical strain, 4.29 fold in El Tor, 1.133 fold in variant strains and about 10.13-10.20 fold for the clinical strains of V. cholerae using the extract of C.asiatica while, the reduction with the exposure to the asiaticoside were 2.762 fold in classical strain, 4.809 in El Tor, 24.1 in variant strain and 34.77 - 34.8 for the clinical strains. CONCLUSION: Centella asiatica extract inhibited the CT production in Vibrio cholerae as well as decreased the transcription of ctxA gene expression.

Antibiotic resistance pattern among common bacterial uropathogens with a special reference to ciprofloxacin resistant Escherichia coli.

BACKGROUND & OBJECTIVES: The resistance of bacteria causing urinary tract infection (UTI) to commonly prescribed antibiotics is increasing both in developing as well as in developed countries. Resistance has emerged even to more potent antimicrobial agents. The present study was undertaken to report the current antibiotic resistance pattern among common bacterial uropathogens isolated in a tertiary care hospital in south India, with a special reference to ciprofloxacin. METHODS: A total of 19,050 consecutive urine samples were cultured and pathogens isolated were identified by standard methods. Antibiotic susceptibility was done by Kirby Bauer disk diffusion method. The clinical and demographic profile of the patients was noted. RESULTS: Of the 19,050 samples, 62 per cent were sterile, 26.01 per cent showed significant growth, 2.3 per cent showed insignificant growth and 9.6 per cent were found contaminated. Significant association (P<0.001) of prior use of antibiotics in males, UTI in adults, gynaecological surgery in females, obstructive uropathy in males and complicated UTI in females with the occurrence of UTI with ciprofloxacin resistant Escherichia coli was noted. Significant association was noted in females with prior antibiotics, with prior urological surgery and in males with prior complicated UTI. There was no significant association with diabetes mellitus with the occurrence of UTI with ciprofloxacin resistant E. coli. Fluoroquinolone resistance was found to increase with age. INTERPRETATIONS & CONCLUSIONS: Ciprofloxacin resistance has emerged due to its frequent use. This resistance was seen more in the in-patients, elderly males and females. Also the resistance to other antibiotics was also high. Increasing antibiotic resistance trends indicate that it is imperative to rationalize the use of antimicrobials in the community and also use these conservatively.