Molecular Studies of the Pathogenic Free-living Amoeba, Acanthamoeba

Ubiquitous amoebae from the genus Acanthamoeba are associated with two main serious infections: The more common eye disease acanthamoeba keratitis (AK), which can result in blindness, and the rare and often fatal disease affecting the central nervous system, granulomatous amoebic encephalitis (GAE). The traditional morphological taxonomic system for Acanthamoeba is based on cyst size and shape, and divides the amoebae into three groups (I, II and III). Since the discovery that cyst shape can be modified by culture conditions, the classic system has become largely redundant. A more robust system has been developed, based on the nucleotide sequence of the 18S rRNA (Rns) gene. It types Acanthamoeba into 15 T-groups, with most species including environmental and clinical clumped into three groups T3, T4 and T11, with little resolution between them. Although speciation does not help cure patients directly, it can provide valuable information regarding disease epidemiology and ultimately benefit patient prognosis. Here a system to better resolve strains has been developed, using the mitochondrial cytochrome oxidase subunit 1 and 2 (cox1/2) gene sequence. When used in conjunction with the T-group system, resolution between strains including those with a T3, T4 or T11 genotype is obtained. Additionally the combined approach identified a mixed infection in a patient suffering with AK, and the occurrence of Acanthamoeba strains with multiple alleles of 18S and cox1/2 genes. The combined use of both genotyping systems was used to investigate an unprecedented outbreak of GAE within a Swedish hospital. Results confirmed Acanthamoeba had infected several immunocompromised paediatrics from a single ICU, and the source was from within the unit’s water system. In vitro assays were used to test the strains pathogenic abilities and sensitivities to antimicrobial compounds, identifying if they are more virulent than typical strains of Acanthamoeba