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J. J. Hutchinson, K. Lathrop, P. Thompson, R. Shanks, R. Kowalski, F. Mah, P. Charukamnoetkanok; The Use of Video Modeling to Evaluate Susceptibility of Acanthamoeba spp. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4307.
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© ARVO (1962-2015); The Authors (2016-present)
To study the potential use of video modeling in evaluating Acanthamoeba spp. susceptibility to clinically utilized agents.
An Acanthamoeba culture taken from an infected eye was cultured in a liquid media mix of 10% Fetal Bovine Serum and 90% YPD Broth. The culture was incubated at 30 degrees C for an average of one week until optimal trophozoite mass were achieved. One mL of the flask culture was transferred to a 1.7mm thickness bioptics dish with one mL of the 10%/90% mixed media. The dish was incubated overnight and transferred to a video microscope for recording. Each dish was photographed at 20X power every 10 seconds for 10 minutes, following which 100 microns of the agent (70% Ethyl alcohol, Brolene 0.1%, Econopred 1%, Gatifloxacin, Chlorhexidine, PHMB) were pipetted into the dish. Photographs were taken every 10 seconds for 2 hours. Following each recording a sample from each dish was inoculated onto an Acanthamoeba culture. The plates were observed for 7 days.
The addition of most agents showed no effect either on the mobility or the morphology of the Acanthamoeba on video recording. Only 70% Ethyl alcohol demonstrated an immediate change. The organism lost its trophozoite appearance and ceased its movement. A few cysts and trophs were observed on the post-recording culture plate with no growth. Cystic looking structures with abnormal morphology and no growth were observed on the Brolene and PHMB plates. No organisms were observed on the Chlorhexidine plate. Trophs with normal multiplication were observed on the Gatifloxacin plate. Precipitate obscured the recording of the Econopred dish.
In this study the response of Acanthamoeba observed using the video model correlated well only with the post recording cultures after the addition of Ethyl alcohol and Gatifloxacin. The cultures performed after the addition of Chlorhexidine, Brolene, and PHMB showed equivocal results with either no organisms or cysts and trophs of abnormal morphology that did not correlate with the healthy and viable appearance of the organisms on the recording. This suggests that there may be a loss of viability of Acanthamoeba due to these agents that is difficult to discern using video modeling.
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