Investigating the evolutionary consequences of gaining a novel pSym on the rhizobia-legume symbiosis. 3 Rhizobium leguminosarum strains lacking native pSyms were given a new pSym confering symbiosis with a novel host and allowed to evolve in planta
In this project, we aimed to investigate the evolutionary process involved in a rhizobial host shift. For almost all rhizobia, genes encoding symbiosis, including host specificity, are located on mobile genetic elements. Thus, host shifts can, and do, occur 'instantly' through horizontal gene transfer. However, novel genes often require a process of network integration and refinement in order to function effectively and to remove any fitness costs stemming from regulatory interference, epistatic interactions or unnecessary biosynthetic burden. For this project the plasmid pJB5JI was introduced to 3 strains lacking their native pSyms. pJB5JI is a conjugative symbiosis plasmid ('pSym) which carries the genes required for symbiosis with pea. We used 3 different backgrounds to represent different levels of evolutionary transition; Strain 248 is the original host of pJB5JI and adapted to hosts such as pea, LPR5039 was previously associated with clover (symbiovar trifolii) - an alternative legume group, but one that forms nodules in a similar way to pea (indeterminate). In indeterminate nodules, the bacteria differentiate terminally into a symbiotic morphotype that cannot resume growth, while a population of vegetatively growing bacteria are maintained which can form the next generation. 8401 was previously associated with common bean (symbiovar phaseolii), which forms determinate nodules. In these hosts symbiotic bacteria are less differentiated and can resume growth and so these bacteria will contribute to the next generation. Strains were evolved in passage through novel (for 2 strains) host, Pea, for 3 experimental transfers of 7 weeks. For each transfer, bacterial cultures grown in the lab (transfer 1) or isolated from crushed nodules (transfers 2 and 3) were introduced to 1-week-old seedlings growing in axenic conditions. Plants were then grown for a further 7 weeks. Nodules were then harvested for the subsequent transfer or for evolved strain isolation. For this, a single clone was selected from each population. Evolved strains were then tested for symbiotic function and growth in the lab. Clones were sequenced via Illumina.
In this project, we aimed to investigate the evolutionary process involved in a rhizobial host shift. For almost all rhizobia, genes encoding symbiosis, including host specificity, are located on mobile genetic elements. Thus, host shifts can, and do, occur 'instantly' through horizontal gene transfer. However, novel genes often require a process of network integration and refinement in order to function effectively and to remove any fitness costs stemming from regulatory interference, epistatic interactions or unnecessary biosynthetic burden. For this project the plasmid pJB5JI was introduced to 3 strains lacking their native pSyms. pJB5JI is a conjugative symbiosis plasmid ('pSym) which carries the genes required for symbiosis with pea. We used 3 different backgrounds to represent different levels of evolutionary transition; Strain 248 is the original host of pJB5JI and adapted to hosts such as pea, LPR5039 was previously associated with clover (symbiovar trifolii) - an alternative legume group, but one that forms nodules in a similar way to pea (indeterminate). In indeterminate nodules, the bacteria differentiate terminally into a symbiotic morphotype that cannot resume growth, while a population of vegetatively growing bacteria are maintained which can form the next generation. 8401 was previously associated with common bean (symbiovar phaseolii), which forms determinate nodules. In these hosts symbiotic bacteria are less differentiated and can resume growth and so these bacteria will contribute to the next generation. Strains were evolved in passage through novel (for 2 strains) host, Pea, for 3 experimental transfers of 7 weeks. For each transfer, bacterial cultures grown in the lab (transfer 1) or isolated from crushed nodules (transfers 2 and 3) were introduced to 1-week-old seedlings growing in axenic conditions. Plants were then grown for a further 7 weeks. Nodules were then harvested for the subsequent transfer or for evolved strain isolation. For this, a single clone was selected from each population. Evolved strains were then tested for symbiotic function and growth in the lab. Clones were sequenced via Illumina.
P3 Institute