Genetic diversity of the Pseudomonas population
Representative Neighbour-Joining (NJ) trees are shown in Figure 2 and Figure 3A. The Pseudomonas population was clearly separated into two major clusters according to origin of isolation, i.e., Oxford or Auckland (ANOSIM in PRIMER, r = 0.477, P < 0.001 (Fig. 3B); Libshuff in Mothur, dCXYScore 0.0254, P < 0.0001). The Oxford cluster contains the reference strain, P. fluorescen s SBW25, which was separately isolated from the phyllosphere of sugar beet grown in the same field. Interestingly, the Oxford cluster contains few isolates from Auckland, and vice versa (highlighted by asterisk in Fig. 2).
A total of 44 and 37 unique STs (81 in total) were detected in the Oxford and Auckland populations, respectively (Table 3). The two populations had no shared STs. However, when the closely related STs were grouped into OTUs at the distance level of 0.06, four shared OTUs were identified for the 11 and 15 OTUs present in the population of Oxford and Auckland, respectively (Table 3). The Oxford population displayed lower levels of nucleotide sequence polymorphism (Fig. 3C), and lower diversity in terms of the Simpson’s index on the basis of both unique STs and OTUs (Fig. 3D). Single likelihood ancestor counting (SLAC) analysis indicated no sites under positive or negative selection for these housekeeping genes.
Recombination, as mentioned above, was detected in a single gene (gapA ) but not in the concatenated sequence by the likelihood permutation test. However, when the analysis was performed with data from Oxford and Auckland separately, significant levels of recombination were noted in the concatenated sequence, particularly for the Oxford population; these were further supported by single breakpoint recombination analysis using the 10340 model (Table 3). Further GARD analysis revealed two breakpoints with significant topological incongruence, indicating recombination breakpoints between the MLSA genes (Fig. 4A).
Recombination events can be utilized to separate the isolates into populations on the basis of ancestry, i.e. the presence of linked alleles in multiple isolates mandates that they have shared ancestry to some degree. Obviously, multiple sources of ancestry can be present in any one isolate. Ancestral analysis performed in STRUCTURE found evidence of six ancestral populations, and their distributions at the level of the population and isolate are presented in Figure 4B and 4C, respectively. Few isolates were “highly recombinant” without any predominant source of their ancestry (8% in total, mostly from Oxford).