Metapopulation dominance and genomic-island acquisition of Bradyrhizobium with superior catabolic capabilities

Root nodule-forming rhizobia exhibit a bipartite lifestyle, replicating in soil and also within plant cells where they fix nitrogen for legume hosts. Host control models posit that legume hosts act as a predominant selective force on rhizobia, but few studies have examined rhizobial fitness in natural populations. Here, we genotyped and phenotyped Bradyrhizobium isolates across more than 800 km of the native Acmispon strigosus host range. We sequenced chromosomal genes expressed under free-living conditions and accessory symbiosis loci expressed in planta and encoded on an integrated ‘symbiosis island’ (SI). We uncovered a massive clonal expansion restricted to the Bradyrhizobium chromosome, with a single chromosomal haplotype dominating populations, ranging more than 700 km, and acquiring 42 divergent SI haplotypes, none of which were spatially widespread. For focal genotypes, we quantified utilization of 190 sole-carbon sources relevant to soil fitness. Chromosomal haplotypes that were both widespread and dominant exhibited superior growth on diverse carbon sources, whereas these patterns were not mirrored among SI haplotypes. Abundance, spatial range and catabolic superiority of chromosomal, but not symbiosis genotypes suggests that fitness in the soil environment, rather than symbiosis with hosts, might be the key driver of Bradyrhizobium dominance

Metapopulation dominance and genomic-island acquisition of Bradyrhizobium with superior catabolic capabilities

Root nodule-forming rhizobia exhibit a bipartite lifestyle, replicating in soil and also within plant cells where they fix nitrogen for legume hosts. Host control models posit that legume hosts act as a predominant selective force on rhizobia, but few studies have examined rhizobial fitness in natural populations. Here, we genotyped and phenotyped Bradyrhizobium isolates across more than 800 km of the native Acmispon strigosus host range. We sequenced chromosomal genes expressed under free-living conditions and accessory symbiosis loci expressed in planta and encoded on an integrated ‘symbiosis island’ (SI). We uncovered a massive clonal expansion restricted to the Bradyrhizobium chromosome, with a single chromosomal haplotype dominating populations, ranging more than 700 km, and acquiring 42 divergent SI haplotypes, none of which were spatially widespread. For focal genotypes, we quantified utilization of 190 sole-carbon sources relevant to soil fitness. Chromosomal haplotypes that were both widespread and dominant exhibited superior growth on diverse carbon sources, whereas these patterns were not mirrored among SI haplotypes. Abundance, spatial range and catabolic superiority of chromosomal, but not symbiosis genotypes suggests that fitness in the soil environment, rather than symbiosis with hosts, might be the key driver of Bradyrhizobium dominance

Host investment into symbiosis varies among genotypes of the legume Acmispon strigosus, but host sanctions are uniform.

Efficient host control predicts the extirpation of ineffective symbionts, but they are nonetheless widespread in nature. We tested three hypotheses for the maintenance of symbiotic variation in rhizobia that associate with a native legume: partner mismatch between host and symbiont, such that symbiont effectiveness varies with host genotype; resource satiation, whereby extrinsic sources of nutrients relax host control; and variation in host control among host genotypes. We inoculated Acmispon strigosus from six populations with three Bradyrhizobium strains that vary in symbiotic effectiveness on sympatric hosts. We measured proxies of host and symbiont fitness in single- and co-inoculations under fertilization treatments of zero added nitrogen (N) and near-growth-saturating N. We examined two components of host control: 'host investment' into nodule size during single- and co-inoculations, and 'host sanctions' against less effective strains during co-inoculations. The Bradyrhizobium strains displayed conserved growth effects on hosts, and host control did not decline under experimental fertilization. Host sanctions were robust in all hosts, but host lines from different populations varied significantly in measures of host investment in both single- and co-inoculation experiments. Variation in host investment could promote variation in symbiotic effectiveness and prevent the extinction of ineffective Bradyrhizobium from natural populations

Evolutionary Instability of Symbiotic Function in Bradyrhizobium japonicum

Bacterial mutualists are often acquired from the environment by eukaryotic hosts. However, both theory and empirical work suggest that this bacterial lifestyle is evolutionarily unstable. Bacterial evolution outside of the host is predicted to favor traits that promote an independent lifestyle in the environment at a cost to symbiotic function. Consistent with these predictions, environmentally-acquired bacterial mutualists often lose symbiotic function over evolutionary time. Here, we investigate the evolutionary erosion of symbiotic traits in Bradyrhizobium japonicum, a nodulating root symbiont of legumes. Building on a previous published phylogeny we infer loss events of nodulation capability in a natural population of Bradyrhizobium, potentially driven by mutation or deletion of symbiosis loci. Subsequently, we experimentally evolved representative strains from the symbiont population under host-free in vitro conditions to examine potential drivers of these loss events. Among Bradyrhizobium genotypes that evolved significant increases in fitness in vitro, two exhibited reduced symbiotic quality, but no experimentally evolved strain lost nodulation capability or evolved any fixed changes at six sequenced loci. Our results are consistent with trade-offs between symbiotic quality and fitness in a host free environment. However, the drivers of loss-of-nodulation events in natural Bradyrhizobium populations remain unknown

Genes in the Ureteric Budding Pathway: Association Study on Vesico-Ureteral Reflux Patients

Vesico-ureteral reflux (VUR) is the retrograde passage of urine from the bladder to the urinary tract and causes 8.5% of end-stage renal disease in children. It is a complex genetic developmental disorder, in which ectopic embryonal ureteric budding is implicated in the pathogenesis. VUR is part of the spectrum of Congenital Anomalies of the Kidney and Urinary Tract (CAKUT). We performed an extensive association study for primary VUR using a two-stage, case-control design, investigating 44 candidate genes in the ureteric budding pathway in 409 Dutch VUR patients. The 44 genes were selected from the literature and a set of 567 single nucleotide polymorphisms (SNPs) capturing their genetic variation was genotyped in 207 cases and 554 controls. The 14 SNPs with p<0.005 were included in a follow-up study in 202 cases and 892 controls. Of the total cohort, ∼50% showed a clear-cut primary VUR phenotype and ∼25% had both a duplex collecting system and VUR. We also looked for association in these two extreme phenotype groups. None of the SNPs reached a significant p-value. Common genetic variants in four genes (GREM1, EYA1, ROBO2 and UPK3A) show a trend towards association with the development of primary VUR (GREM1, EYA1, ROBO2) or duplex collecting system (EYA1 and UPK3A). SNPs in three genes (TGFB1, GNB3 and VEGFA) have been shown to be associated with VUR in other populations. Only the result of rs1800469 in TGFB1 hinted at association in our study. This is the first extensive study of common variants in the genes of the ureteric budding pathway and the genetic susceptibility to primary VUR

Evolutionary instability of symbiotic function in Bradyrhizobium japonicum.

Bacterial mutualists are often acquired from the environment by eukaryotic hosts. However, both theory and empirical work suggest that this bacterial lifestyle is evolutionarily unstable. Bacterial evolution outside of the host is predicted to favor traits that promote an independent lifestyle in the environment at a cost to symbiotic function. Consistent with these predictions, environmentally-acquired bacterial mutualists often lose symbiotic function over evolutionary time. Here, we investigate the evolutionary erosion of symbiotic traits in Bradyrhizobium japonicum, a nodulating root symbiont of legumes. Building on a previous published phylogeny we infer loss events of nodulation capability in a natural population of Bradyrhizobium, potentially driven by mutation or deletion of symbiosis loci. Subsequently, we experimentally evolved representative strains from the symbiont population under host-free in vitro conditions to examine potential drivers of these loss events. Among Bradyrhizobium genotypes that evolved significant increases in fitness in vitro, two exhibited reduced symbiotic quality, but no experimentally evolved strain lost nodulation capability or evolved any fixed changes at six sequenced loci. Our results are consistent with trade-offs between symbiotic quality and fitness in a host free environment. However, the drivers of loss-of-nodulation events in natural Bradyrhizobium populations remain unknown

Efficiency of partner choice and sanctions in Lotus is not altered by nitrogen fertilization.

Eukaryotic hosts must exhibit control mechanisms to select against ineffective bacterial symbionts. Hosts can minimize infection by less-effective symbionts (partner choice) and can divest of uncooperative bacteria after infection (sanctions). Yet, such host-control traits are predicted to be context dependent, especially if they are costly for hosts to express or maintain. Legumes form symbiosis with rhizobia that vary in symbiotic effectiveness (nitrogen fixation) and can enforce partner choice as well as sanctions. In nature, legumes acquire fixed nitrogen from both rhizobia and soils, and nitrogen deposition is rapidly enriching soils globally. If soil nitrogen is abundant, we predict host control to be downregulated, potentially allowing invasion of ineffective symbionts. We experimentally manipulated soil nitrogen to examine context dependence in host control. We co-inoculated Lotus strigosus from nitrogen depauperate soils with pairs of Bradyrhizobium strains that vary in symbiotic effectiveness and fertilized plants with either zero nitrogen or growth maximizing nitrogen. We found efficient partner choice and sanctions regardless of nitrogen fertilization, symbiotic partner combination or growth season. Strikingly, host control was efficient even when L. strigosus gained no significant benefit from rhizobial infection, suggesting that these traits are resilient to short-term changes in extrinsic nitrogen, whether natural or anthropogenic