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Exposure to sublethal concentrations of chlorine enhances the cytotoxicity of Acanthamoeba castellanii

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Abstract

Free-living amoebae belonging to the genus Acanthamoeba are the causative agents of infections in humans and animals. Many studies are being conducted to find effective compounds against amoebae, but their sublethal concentration effects on surviving amoebae seem to have been overlooked. Chlorine is a common disinfection agent commonly added to public water facilities and supplies. In this study, the cytopathic and phagocytic properties of Acanthamoeba castellanii trophozoites following exposure to sublethal concentrations of chlorine were examined. Two hours of exposure to 5 ppm hypochlorite calcium was considered the sublethal concentration for A. castellanii trophozoites. To compare the pathogenic potential of treated and untreated Acanthamoeba trophozoites, cytotoxicity, adhesion assays in RAW 264.7 macrophages, osmo, and thermotolerance tests were carried out. Bacterial uptake was assessed in treated cells to evaluate their phagocytic characteristics. Oxidative stress biomarkers and antioxidant activities were compared in treated and untreated trophozoites. Finally, the mRNA expression of the mannose-binding protein (MBP), cysteine protease 3 (CP3), and serine endopeptidase (SEP) genes was determined in cells. In all the experiments, untreated trophozoites were considered the control. In comparison to untreated trophozoites, in chlorine-treated trophozoites, cytopathic effects were more extensive and resulted in the detachment of macrophage monolayers. Treated trophozoites could not grow at high temperatures (43 °C). Besides, they showed osmotolerance to 0.5 M D-mannitol but not to 1 M. Results demonstrated a higher bacterial uptake rate by chlorine-treated trophozoites than untreated cells. The treated and untreated cells had significantly different glutathione and glutathione/glutathione disulfide ratios. Antioxidant enzyme activities, total antioxidant capacity, and malondialdehyde levels were increased significantly in chlorine-treated cells. Quantifying mRNA expression in chlorine-treated trophozoites revealed that virulence genes were upregulated. Chlorine can form resistance and virulent amoebae if it is not used at a proper concentration and exposure time. Identification of stress responses, their mechanisms in Acanthamoeba, and their relation to amoeba virulence would give us a better perception of their pathophysiology.

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Acknowledgement

This study was supported by the research grant provided by the Shahid Chamran University of Ahvaz.

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Authors and Affiliations

  1. Department of Parasitology, Faculty of Veterinary Medicine, Shahid Chamran University of Ahvaz, Ahvaz, Iran

    Somayeh Bahrami, Zeinab Asadi & Hossein Hamidinejat

  2. Department of Food Hygiene, Faculty of Veterinary Medicine, Shahid Chamran University of Ahvaz, Ahvaz, Iran

    Mehdi Zarei

  3. Institute of Biomedical and Environmental Health Research, School of Health and Life Sciences, University of the West of Scotland (UWS), Paisley, PA1 2BE, Scotland, UK

    Fiona L. Henriquez

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  1. Somayeh Bahrami
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  2. Zeinab Asadi
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  3. Mehdi Zarei
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  5. Fiona L. Henriquez
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Contributions

S.B., M.Z., and F.L.H. designed the research; S.B., M.Z., and Z.A. performed the experiments. Z.A., S.B., and H.H. performed data analysis and prepared the figures; S.B. wrote the manuscript; F.L.H. revised the manuscript; S.B., Z.A., M.Z., H.H., and F.L.H. approved the version to be published.

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Correspondence to Somayeh Bahrami.

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Bahrami, S., Asadi, Z., Zarei, M. et al. Exposure to sublethal concentrations of chlorine enhances the cytotoxicity of Acanthamoeba castellanii. Parasitol Res 122, 1371–1380 (2023). https://doi.org/10.1007/s00436-023-07837-z

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