Background To date no standardized methods are used in order to assess the amoebicidal efficacy of commercial contact lens solutions for both trophozoites and cysts of Acanthamoeba species. Here we present two methods that are suitable for this purpose: The Spearman-Karber log reduction method and XTT colorimetric assay. Methods Acanthamoeba castellanii (ATCC 50370) and A. polyphaga (ATCC 30461) trophozoites were cultured in peptone-yeast extract-glucose medium. Cysts were developed in Neff’s encystement medium for 1 week. Spearman-Karber and XTT colorimetric assay were used to evaluate trophozoite and cystocidal efficacy of multi-purpose contact lens solutions (MPS). Results With trophozoites, the Spearman-Karber method gave a log reduction estimate of morphological kill between log 0,83 and log 3,61 of the various contact lens solutions, enabling the differentiation between efficacious and less efficacious solutions. With cysts the maximum log reduction of 2,17 was achieved for all 3 MPS solutions at 8 hours. The XTT colorimetric assay showed reduction in trophozoite metabolic rates between 50 and 100% as provided by an optical density signal. All lens fluid solution with a reduction rate > 90% showed residual growth of Acanthamoeba after one week of incubation on nutrient agar covered with Enterobacter aerogenes. Conclusion Both methods give reproducible estimates of amoebicidal efficacy of contact lens solutions, however, XTT colorimetric assay should be followed by an assay for residual growth to test for viable cysts.

Acanthamoeba keratitis is almost universally associated with contact lens (CL) use. Until today, however, CL solution manufacturing protocols lack testing of anti-amoebic activity. This study investigates the effectiveness of CL solutions available on the Dutch market against trophozoites and cysts of Acanthamoeba castellanii and Acanthamoeba polyphaga. Sixteen CL solutions were tested: 13 multiple purpose solutions (MPS), 2 hydrogen peroxidase solutions (HPS) and 1 povidone-iodine-based solution (PIS). The Spearman–Karber (SK) log reduction method and an XTT colorimetric assay were used to evaluate the effectiveness at the manufacturer’s minimum recommended disinfection time (MMRDT) and after eight hours. At the MMRDT, one MPS showed an SK mean log reduction (MLR) of >3.0 against A. castellanii trophozoites. Two additional MPS and both HPS reached this threshold after eight hours. The SK MLR values for A. polyphaga trophozoites were between 1 and 3 at all time points. Using the XTT colorimetric assay, only HPS 1 showed >99.9% reduction (equivalent to 3 log reduction) in metabolic activity of A. castellanii trophozoites after eight hours. For A. polyphaga, both HPS and PIS showed a metabolic reduction of >99.9% after eight hours. Cysts were resistant against all solutions. We conclude that following the manufacturer’s guidelines, few solutions provide sufficient effectiveness against Acanthamoeba trophozoites and none against cysts. The results underline the importance of adequate hygiene when handling CLs.