Sensing and adhesion are adaptive functions in the plant pathogenic xanthomonads

Nadia Mhedbi-Hajri 1 Armelle Darrasse 1 Sandrine Pign 1 Karine Durand 1 Stphanie Fouteau Valrie Barbe Charles Manceau 1 Christophe Lemaire 0 Marie-Agns Jacques 1 0 UMR077 PaVe, Universite d'Angers , 42, rue Georges Morel, F-49071 Beaucouze , France 1 UMR077 PaVe, INRA , 42, rue Georges Morel, F-49071 Beaucouze , France Background: Bacterial plant pathogens belonging to the Xanthomonas genus are tightly adapted to their host plants and are not known to colonise other environments. The host range of each strain is usually restricted to a few host plant species. Bacterial strains responsible for the same type of symptoms on the same host range cluster in a pathovar. The phyllosphere is a highly stressful environment, but it provides a selective habitat and a source of substrates for these bacteria. Xanthomonads colonise host phylloplane before entering leaf tissues and engaging in an invasive pathogenic phase. Hence, these bacteria are likely to have evolved strategies to adapt to life in this environment. We hypothesised that determinants responsible for bacterial host adaptation are expressed starting from the establishment of chemotactic attraction and adhesion on host tissue. Results: We established the distribution of 70 genes coding sensors and adhesins in a large collection of xanthomonad strains. These 173 strains belong to different pathovars of Xanthomonas spp and display different host ranges. Candidate genes are involved in chemotactic attraction (25 genes), chemical environment sensing (35 genes), and adhesion (10 genes). Our study revealed that candidate gene repertoires comprised core and variable gene suites that likely have distinct roles in host adaptation. Most pathovars were characterized by unique repertoires of candidate genes, highlighting a correspondence between pathovar clustering and repertoires of sensors and adhesins. To further challenge our hypothesis, we tested for molecular signatures of selection on candidate genes extracted from sequenced genomes of strains belonging to different pathovars. We found strong evidence of adaptive divergence acting on most candidate genes. Conclusions: These data provide insight into the potential role played by sensors and adhesins in the adaptation of xanthomonads to their host plants. The correspondence between repertoires of sensor and adhesin genes and pathovars and the rapid evolution of sensors and adhesins shows that, for plant pathogenic xanthomonads, events leading to host specificity may occur as early as chemotactic attraction by host and adhesion to tissues. - Background Deciphering how bacteria adapt to their hosts helps explain how they spread. Host specificity can be established by determining the genes coding virulence factors that are not conserved among strains, which differ in their host range [1]. Virulence-associated genes are expressed during initial host colonisation, multiplication, development of symptoms, and dispersal. Sarkar and colleagues [1] and Hajri and associates [2] demonstrated that canonical virulence factors such as type III effectors (T3Es) play a critical role in host specificity. T3Es, however, are injected into plant host cells once bacteria have already penetrated into host tissues [3]. Thus, phases preceding infection could also be involved in host specificity and therefore be under selective pressures. For bacteria to adapt specifically to their hosts, they sense favourable environmental stimuli and then they move toward favourable conditions [4,5]. Bacteria have evolved receptors and sensors in their cell walls to detect chemical and environmental signals such as the presence of chemoattractants, chemorepellents, and oxygen. They thereby integrate information on their biotic and abiotic environment [6]. Studies on Rhizobia revealed the importance of sensors in the perception of specific host signals early during symbiotic interaction with legumes [7,8]. Similarly, Agrobacterium tumefaciens and Ralstonia solanacearum specifically detect various components from root exudates that attract them toward their hosts [9,10]. Environmental signals are mainly detected by Methylaccepting Chemotaxis Proteins (MCPs) and Sensors of Two-Component Regulatory System (STCRS). MCPs are the principal components of the chemotaxis system [4]. Detection of signals by these transmembrane chemoreceptors directs cell locomotion by regulating the histidine kinase CheA, which in turn communicates the information to the flagellar motor by phosphorylating its cognate response regulator CheY [4,5]. Changes in the direction or the speed of flagellar rotation modify swimming behaviour, resulting in movement towards higher gradients of attractants and away from high concentrations of repellents [11,12]. In Escherichia coli, chemotaxis proteins cluster in membrane-associated patches [13,14]. Interactions within patches contribute to the notable features of this signalling system: high sensitivity, wide dynamic range, signal integration, memory, and adaptation [15]. Besides MCPs, bacteria sense their nutritional environment through TonBDependent Transporters (TBDTs) [16]. A large proportion of TBDT genes are related to plant scavenging and carbohydrate utilisation. TBDTs are over-represented in various bacteria interacting with plants such as Xanthomonas spp. [17]. Adhesion to a surface is a prerequisite for aggregation in a biofilm, which enhances the resistance of bacteria to various biotic and abiotic stresses, favours the coordination of adapted responses to environmental changes, and allows multiplication [18,19]. Sensing and adhesion mechanisms are interconnected since biofilm formation is regulated by a chemosensory system [20]. The adhesion step involves surface structures in a broad group of fimbrial and nonfimbrial adhesins. The fimbrial proteins include type IV pili (Tfp), which are polymeric assemblies of the protein pilin [21,22]. The nonfimbrial adhesins belong to the autotransporter family (e.g. XadA and YadA proteins) [23,24] and to the two-partner secretion system (e.g. FhaB and YapH proteins) [25]. Each plant pathogenic bacterium belonging to the Xanthomonas genus is able to colonise a restricted variety of plant hosts and microniches. Xanthomonas are exclusively plant-associated bacteria, mainly phyllosphere colonisers, and are not encountered in other environments [26]. Globally, they infect a huge range of economically important plants such as rice, banana, citrus, bean, tomato, pepper, sugarcane, and wheat [26]. The large host range of the genus strikingly contrasts with the typically narrow host range of individual strains restricted to one or several species of a botanical family [27]. Besides their very homogeneous phenotype, xanthomonads differ mainly by their host specificity. This is illustrated in the pathovar subspecific division, which clusters bacterial strains causing similar symptoms on a same host range [28]. A few pathovars are represented by polyphyle (...truncated)