Fig. S3

Consequence of Bd infection on zebrafish 44 larvae host tissue (Related to Figure 3).

(a) Zebrafish larvae bath water was inoculated with with mTGhL plate washings (control) or low (< 200 zsp/μl) dose Bd zoospores and incubated for 72 hpi, then labelled with calcoluor white (CFW; for chitin, green) and evans blue (EB; for tissue damage, red), and visualised by confocal microscopy. Images taken at 63X, maximum intensity projection of Z-stack shown here. Cartoon depicts imaged region. Representative images with insets highlight colocalisation of CFW-labelled punctae with EB positive tissue damage. Scale bars = 50 μm. (b) Zebrafish larvae bath water was inoculated with high (> 200 zsp/μl) dose Bd zoospores, then labelled and imaged as in (a). Scale bar = 50 μm. (c) Zebrafish larvae bath water was inoculated with mTGhL (control) or low dose Bd zoospores and incubated for 72 hpi, then fixed, labelled with TUNEL (for apoptotic cells; red) and imaged by fluorescent stereomicroscopy. Representative images with dotted outline of larvae tail fin showing an increase in apoptotic cells on infected larvae. (d) Zebrafish larvae bath water was inoculated with control and incubated for 72 hpi, then fixed for electron microscopy (EM). Images show healthy larvae skin. (e) Diagram showing phenotypes used to categorise severity of muscle degeneration in zebrafish larvae. Left column shows cartoons to illustrate the appearance of healthy muscle; right column shows cartoons for eroded muscle, including loose packing, thinning, crumpling and deep tissue degeneration of muscle fibres.