Characterizing clinical isolates of Acanthamoeba castellanii with high resistance to polyhexamethylene biguanide in Taiwan.

ABSTRACT

BACKGROUND/PURPOSE: Acanthamoeba keratitis (AK), a painful infectious corneal disease, is caused by the free-living pathogenic species Acanthamoeba. The symptoms include corneal infiltrate, epithelial, and stromal destruction, and loss of vision. Current treatment generally involves an hourly application of polyhexamethylene biguanide (PHMB) over a period of several days; however, even this is not entirely effective against all strains/isolates. The aims of this study were to confirm the existence of pathogenic strains in Taiwan which are highly resistant to drugs and to characterize the behavior of these strains. METHODS: An in vitro Acanthamoeba species culture platform was established to observe the effectiveness of treatment and chart the morphological changes that occur under the effects of drugs using a light microscope and time-lapse recording. Changes in gene expression were examined using reverse transcription polymerase chain reaction (RT-PCR) and real-time PCR. RESULTS: Over 90% of the standard strain cells (ATCC 30010) were lysed after being treated with PHMB for 1 hour; however, clinical isolates of Acanthamoeba castellanii that differed in their susceptibility to the treatment drug were only partly lysed. Following treatment with PHMB, National Cheng Kung University Hospital isolation B (NCKH_B) transformed into a pseudocyst under the effects of drug stress; however, National Cheng Kung University Hospital isolation D (NCKH_D), an isolate with higher tolerance for PHMB, did not transform. CONCLUSION: Our results confirm the existence of clinical isolates of A. castellanii with high resistance to PHMB in Taiwan and present the alternative drug tolerance of A. castellanii in addition to the transformation of pseudocyst/cyst.