Stages of fruiting body formation in wild-type M. xanthus. (A) Initial organization of cells after being spotted on an agarose surface (0 h). (B) Rearrangement of cells into a matte (3 h). (C) Towers of cells only a few cell layers thick (8 h). (D) Aggregation centers are larger towers of cells composed of many cell layers (10 h). (E) Mature fruiting body (48 h). Bar, 0.1 mm. Pictures are derived from two separate movies and are not spatially aligned. This figure contains some frames from Movie S1 in the supplemental material.

Tier formation during pilA mutant fruiting body development. Two successive tiers form in the same fruiting body (min 620 to 710 in Movie S2 in supplemental material, 10 min between frames). Pictures are spatially aligned. Bar, 0.1 mm.

Fruiting body formation in the csgA mutant. (A) Initial organization of cells after being spotted on agarose (0 h). (B) Instead of cells arranging into a stable matte, tower formation is observed (3 h). (C) Despite the absence of an initial quiescent phase, aggregates are formed at approximately the same time as the wild type (6 h). (D) Unusually large aggregates are formed (16 h). (E) Cells swarm away from large aggregates, often to form new aggregates which also dissolve (17 h). Arrows indicate large flares of cells swarming out of the aggregation center shown in panel D. (F) Constant swarming and reforming of aggregates disrupt fruiting body structure and maturation (45 h). Pictures are derived from the same movie, but with the exception of panels D and E, the pictures are not spatially aligned. Bars, 0.1 mm. The original movie on which this figure is based is available as Movie S3 in the supplemental material.

Stages of fruiting body formation in the pilA mutant. (A) Initial organization of cells after being spotted on agarose (0 h). (B) Rearrangement of cells into a matte (6 h). (C) Towers only three to four cell layers thick (9 h). (D) pilA mutant aggregation centers display tiers for a longer period of time than wild-type cells before becoming obscured by ECM material. Note that an aggregation center has two origins for a new tier, which is not often observed (12 h). (E) Mature fruiting body. pilA mutant fruiting bodies are often less compact and hemispherical than wild-type fruiting bodies (48 h). Bar, 0.1 mm. Pictures are derived from the same movie but not spatially aligned. The original movie on which this figure is based is available as Movie S2 in the supplemental material.

Model for tier formation in M. xanthus. The panels represent a sequence of events visualized from left to right. The top portion of each panel displays a top-down view of aggregates, while the bottom portion of the panel shows a side view of the aggregate. The blocks in the side view represent cell swarms traveling along the agar surface. In the first panel, colliding swarms moving in opposite directions intersect and cause one of the swarms to orient at a slight vertical pitch to cross over the other swarm (left arrow), generating an inclined plane. As the swarms continue to converge, cells stream along this vertical alignment and then realign horizontally with the cells below them, thereby forming the first tier (second panel from left). More cells stream into the first tier, increasing its surface area. In addition, some cells continue the vertical ascension through the first tier to initiate the second tier (third panel). This process can continue to form multiple tiers (fourth panel).