Figure 2. Effect of incubation in different media types on capsule growth and cell survival.

C. neoformans strains H99 and C. gattii strains WM161 and 409 were cultured overnight in YPD media, washed once in phosphate buffered saline (PBS), and then incubated for 3 d in PBS, filtered seawater (SW), or minimal media (MM). Capsules and cell bodies were measured, and capsule:body volume ratio was calculated. Results represent two independent experiments per strain and condition. A) Incubation in SW did not induce significant capsular growth in any strain (H99, P=0.1606; WM161, P=0.6451; 409, P>0.9999). Both C. gattii strains had larger average capsule:body volume ratios after PBS incubation compared to baseline (WM161, P<0.0001); 409, P=0.0005), while the average capsule:body volume ratio of strain H99 did not change in response to PBS incubation (P>0.9999). As expected, incubation in MM resulted in capsular growth in all strains (P<0.0001). B) Representative microscopy images with India Ink counterstaining showing relative differences in capsule size between H99 and 409 strains under four media conditions. Strain H99 showed no change in capsule size in response to incubation in PBS or SW, while strain 409 developed a larger capsule following PBS incubation. Strain WM161 was similar to strain 409 in appearance. Scale bar = 10 μm.