Adult zebrafish immunized with membrane vesicles and subsequently challenged with Piscirickettsia salmonis. (A) Cumulative survival (%) of adult zebrafish immunized with 20 μg of MVs isolated from LF-89 or injected with phosphate buffer before challenged with P. salmonis 1 × 10⁸ CFU (n = 65). (B) Histological sections from non-infected fish (I-II), fish injected with phosphate buffer (III-IV) and fish immunized with MVs (V-VI) at 3 and 7 days' post challenge (dpc), 10× magnifications, hematoxylin and Periodic acid Schiff's staining. Int: intestines, L. liver, arrows indicate the formation of granuloma-like structures.

Indirect Fluorescent Antibody Test of histological sections from adult zebrafish immunized with membrane vesicles and subsequently challenged with Piscirickettsia salmonis. (A) Identification of P. salmonis by Indirect Fluorescent Antibody Test (IFAT; green) at 3 (I-II) and 7 dpc (III-IV). Bacterial cells positive for IFAT staining are marked with arrowhead, 100× magnifications. (B) Image analysis using Image J automatic particle count of histological sections stained with IFAT at 3 and 7 dpc for quantification of the bacterial load. Results are presented as mean+/-SD. Asterisk indicate significant difference in particle count p < 0.05, two tailed unpaired Student's t-test (n = 2).

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