Serial In Vivo Passage of Acanthamoeba lugdunensis Is Associated with Altered Transcription and Increased Blood-Brain Barrier Permeability in Mice.

We examined passage-associated phenotypic and transcriptional changes in the clinical Acanthamoeba lugdunensis keratitis isolate KA/E2 using preplanned serial in vivo passage in C57BL/6 mice. Amoebae were recovered from lung but not brain cultures during the first cycle. Brain cultures yielded lineages P2 and P3 during the second and third cycles, respectively, with significantly greater recovery in the third than the second cycle on day 4 after plating. In a separate experiment, P3 infection was associated with greater blood-brain barrier (BBB) permeability and earlier body-weight loss than E2 infection; body weight differed significantly between infected groups only on day 8. Cldn5 and Ocln expression was lower in P3-infected mice than in controls; Tjp1 showed no significant pairwise differences. RNA sequencing identified 794 differentially expressed genes between parental E2 and P3. Upregulated genes were enriched for carbohydrate metabolism and hydrolase annotations and included transcripts annotated as a putative IgA-specific serine endopeptidase-like protein and cysteine proteinase 5. Phosphorylation-related genes were enriched among downregulated genes. These differences were detected after five axenic subcultures of P3, although their stability and heritability remain unknown. Total gelatinolytic activity was higher in P3 excretory-secretory products than in E2. Phenylmethylsulfonyl fluoride reduced activity in both isolates, with no significant difference between isolates after treatment; E-64 produced no significant change in total activity. Serial in vivo passage was associated with brain culture recovery, transcriptional differences, and greater BBB permeability, without establishing causality.