A mechanistic explanation linking adaptive mutation, niche change, and fitness advantage for the wrinkly spreader

Abstract

Experimental evolution studies have investigated adaptive radiation in static liquid microcosms using the environmental bacterium Pseudomonas fluorescens SBW25. In evolving populations a novel adaptive mutant known as the Wrinkly Spreader arises within days having significant fitness advantage over the ancestral strain. A molecular investigation of the Wrinkly Spreader has provided a mechanistic explanation linking mutation with fitness improvement through the production of a cellulose-based biofilm at the air-liquid interface. Colonisation of this niche provides greater access to oxygen, allowing faster growth than that possible for non-biofilm-forming competitors located in the lower anoxic region of the microcosm. Cellulose is probably normally used for attachment to plant and soil aggregate surfaces and to provide protection in dehydrating conditions. However, the evolutionary innovation of the Wrinkly Spreader in static microcosms is the use of cellulose as the matrix of a robust biofilm, and is achieved through mutations that deregulate multiple diguanylate cyclases leading to the over-production of cyclic-di-GMP and the stimulation of cellulose expression. The mechanistic explanation of the Wrinkly Spreader success is an exemplar of the modern evolutionary synthesis, linking molecular biology with evolutionary ecology, and provides an insight into the phenomenal ability of bacteria to adapt to novel environments.

Figure 1

Adaptive radiation in static microcosms gives rise to the Wrinkly Spreader with a new niche preference. Shown on the left is a King's B agar plate, incubated for three days at 28°C, spread with a sample taken from a diversified P. fluorescens SBW25 population where both Smooth morphs and Wrinkly Spreader colonies are evident (this plate has seven Smooth colonies each with a rounded circumference and a smooth convex surface, with one positioned at the top of the plate; all of the rest are Wrinkly Spreader colonies which have irregular, multilobed circumferences and a flattened and wrinkled surface). On the right are two static King's B microcosms which were incubated for three days at 28°C. The left microcosm was inoculated with the wild-type or ancestral SBW25 which grows throughout the liquid column, and the right microcosm with the Wrinkly Spreader which colonises the A-L interface through the formation of a robust biofilm.

Figure 2

Linking the timeline of adaptive radiation to events and consequences in the rise of the Wrinkly Spreader. The adaptive radiation of P. fluorescens SBW25 in static microcosms can be mapped to a timeline and epochs during which the ancestral strain diversifies into new genotypes with altered niche preferences. The events and consequences of mutation are outlined for the Wrinkly Spreader along the bottom panel of the figure. Although not shown here, further diversification will occur amongst the Smooth morphs and Fuzzy Spreaders.

Figure 3

The Wsp system is responsible for the synthesis of cyclic-di-GMP and the activation of the Wrinkly Spreader (WS) phenotype. The functioning of the P. fluorescens SBW25 Wsp system has been modelled on the Che chemosensory system of E. coli and provides a mechanistic explanation linking adaptive mutations to the WS phenotype. The methyl-accepting chemotaxis protein (WspA), scaffold proteins (WspB and WspD), and histidine kinase (WspE) form a membrane-associated receptor-signaling complex. In the absence of an appropriate environmental signal, the complex is silent. Upon activation by phosphorylation (indicated by the black circles), the diguanylate cyclase (DGC) response regulator (WspR) synthesizes cyclic-di-GMP from GTP. The system is controlled by the opposing activities of a methyltransferase (WspC) and methylesterase (WspF), which add and remove, respectively, methyl (CH₃) groups on the signalling domain of WspA (circles). In wild-type SBW25, the activities of the two are balanced, preventing the activation of WspR and allowing the Wsp complex to oscillate between active and inactive states. Mutations inhibiting WspF function or activating WspE kinase activity result in the activation of WspR and the production of cyclic-di-GMP. Increased levels of cyclic-di-GMP then lead to the expression of the WS phenotype. The Wsp system is shown as a schematic only; the three-dimensional structure of the proteins, their relative placement, numbers, and the positioning of the complex in the inner membrane have not yet been determined.