Acanthamoeba is a heterotrophic protist that can cause visually devastating Acanthamoeba keratitis (AK), often linked with contact lens wear or ocular trauma. Acanthamoeba feeds on different microbes but some microbes can exploit it as a host. These intracellular microbes may contribute to the severity of amoebal infection or be trained to invade other eukaryotic cells. A combination of PCR, next-generation sequencing (NGS) of the 16S V1-V3 rRNA gene, ITS rRNA sequencing, and fluorescence in situ hybridization (FISH) techniques were used to identify intracellular bacteria and fungi in Acanthamoeba isolates (n=22) recovered from culture-proven AK infected patients and environmental samples.
Among the 22 isolates, ten clinical isolates and one environmental (water) isolate harboured bacteria (50%, 11/22) and two (9.1%, 2/22) clinical isolates harboured fungi. The core bacterial microbiome included bacteria of Enterobacteriaceae family with the most common intracellular bacteria being Enterobacter spp. (56.9%) followed by Escherichia spp. (1.53%) and Micrococcus spp. (0.25%). Actinobacteria spp. (1.2%) was also detected in one clinical isolate of Acanthamoeba and Aquella oligotrophica (0.6%) was detected only in the environmental isolate. Malassezia restricta was identified as the intracellular fungus in the two clinical Acanthamoeba strains. Bacterial species richness and community diversity were not significantly different (p>0.05) by geography or source of Acanthamoeba isolates. Upon FISH staining, the intracellular bacteria and fungi were localized throughout the cytosol.
There is a limited range of bacterial microbiome in Acanthamoeba isolates from both clinical and environmental sources. Although infrequent, clinical or environmental Acanthamoeba strains can harbour Malassezia restricta. Future studies will examine whether these intracellular microbes are associated with worse clinical diseases and whether their life within the Acanthamoeba can train them to be infectious for mammalian cells.