Figure 4. Volatile organic compounds released by S. venezuelae raise the medium pH and induce exploratory growth in physically separated Streptomyces.

(A) Effect of S. venezuelae explorer cells on pH of physically separated medium. Each compartment is separated by a polystyrene barrier. S. venezuelae and S. cerevisiae were grown in the left compartment of one plate (left), while S. venezuelae alone was grown in the left compartment of the other plate (right). After 10 days, bromothymol blue pH indicator dye was spread on the agar in the right compartment of each plate. Blue indicates VOC-induced alkalinity. (B) S. venezuelae was grown alone on YP (G- agar) in the left compartment, while the right compartment contained uninoculated YP (G-) agar. After seven days, the same pH indicator dye as in Figure 4A was spread over the agar in the right compartment. Blue represents a rise in pH above 7.6. (C) Left: S. venezuelae alone was inoculated in each compartment. Right: S. venezuelae was grown beside S. cerevisiae in the left compartment, and S. venezuelae alone was grown in the right compartment. All strains were grown on YPD (G+) agar medium for 10 days. (D) Top left: Wild Streptomyces isolate WAC0566 was grown alone in each compartment. Top right: WAC0566 was grown beside S. cerevisiae in the left compartment, and grown alone in the right compartment. Bottom left: S. venezuelae was grown beside S. cerevisiae in the left compartment, and WAC0566 was grown alone in the right compartment. Bottom right: WAC0566 was grown beside S. cerevisiae in the left compartment, while S. venezuelae was grown alone in the right compartment. All strains were cultured on YPD (G+) agar medium for 10 days. (E) Schematic of the plate-based assay used to assess the effects of volatile-emitting solutions (and controls) on nearby Streptomyces colonies. H₂O, TMA, or ammonia solutions were placed in a blue plastic dish, and S. venezuelae was spotted around each dish on YPD medium. Plates were incubated at room temperature for seven days. (F) Surface area and pH of S. venezuelae colonies grown on YPD medium around small dishes containing H₂O or TMA solutions, as shown in Figure 4E. S. venezuelae was grown at room temperature for seven days on either unbuffered YPD medium or YPD medium buffered to pH 7.0 using MOPS. All values represent the mean ± standard error for four replicates.

DOI: http://dx.doi.org/10.7554/eLife.21738.015

Figure 4—figure supplement 1. The S. venezuelae cydCD mutant strain can explore in response to volatile signals produced by neighbouring explorer cells.

Left: Wild type (WT) S. venezuelae was grown beside S. cerevisiae in the left compartment, and the S. venezuelae cydCD mutant strain was grown alone in the right compartment. Middle: Wild type S. venezuelae was grown beside S. cerevisia in the left compartment, and wild type S. venezuelae was grown alone in the right compartment. Right: The cydCD mutant was grown adjacent to S. cerevisiae in the left compartment (where it sporulated but did not spread), while wild type S. venezuelae was grown alone in the right compartment. All strains were grown on YPD (G+) agar medium for 10 days.

DOI: http://dx.doi.org/10.7554/eLife.21738.016

Figure 4—figure supplement 2. Wild explorer Streptomyces species promote exploration in S. venezuelae using volatile signals.

Using our two-quadrant assay, 13 independent wild Streptomyces isolates (WAC strains) were inoculated on G- agar, adjacent to S. venezuelae inoculated on G+ agar medium (where no exploration was observed on its own). Each WAC strain was able to promote S. venezuelae exploration through the release of a volatile compound.

DOI: http://dx.doi.org/10.7554/eLife.21738.017

Figure 4—figure supplement 3. The VOC produced by S. venezuelae explorer cells can be produced by liquid-grown (G-) S. venezuelae and WAC0566 cultures.

All strains were grown in 48-well plates, with (l) indicating liquid cultures (top rows), and (s) indicating solid YPD agar (bottom rows). Liquid cultures were either G+ (glucose-containing) or G- (no glucose), while all solid medium was G+ (exploration repressive condition). Plates were grown shaking for three days. For all plates, we monitored exploration by strains growing on G+ agar (bottom rows), in response to VOCs produced by the liquid-grown cultures. The top panel shows S. venezuelae (left) and the wild Streptomyces strain WAC0566 (right) grown in G+ liquid (a condition where the VOC of interest is not expected to be produced). The middle panel shows the same strains, only grown in G- liquid (where the VOC was predicted to be produced). The bottom panel shows the test for interspecies VOC production/response, with S. venezuelae and WAC0566 grown in G- liquid, opposite WAC0566 and S. venezuelae, respectively.

DOI: http://dx.doi.org/10.7554/eLife.21738.018