Wednesday, August 08, 2012

Silly microbiologist, genomes are for mutualists

New paper in PLoS Genetics of possible interest: PLoS Genetics: Population Genomics of the Facultatively Mutualistic Bacteria Sinorhizobium meliloti and S. medica.

The abstract does an OK job with the technical details:
The symbiosis between rhizobial bacteria and legume plants has served as a model for investigating the genetics of nitrogen fixation and the evolution of facultative mutualism. We used deep sequence coverage (>100×) to characterize genomic diversity at the nucleotide level among 12 Sinorhizobium medicae and 32 S. meliloti strains. Although these species are closely related and share host plants, based on the ratio of shared polymorphisms to fixed differences we found that horizontal gene transfer (HGT) between these species was confined almost exclusively to plasmid genes. Three multi-genic regions that show the strongest evidence of HGT harbor genes directly involved in establishing or maintaining the mutualism with host plants. In both species, nucleotide diversity is 1.5–2.5 times greater on the plasmids than chromosomes. Interestingly, nucleotide diversity in S. meliloti but not S. medicae is highly structured along the chromosome – with mean diversity (θπ) on one half of the chromosome five times greater than mean diversity on the other half. Based on the ratio of plasmid to chromosome diversity, this appears to be due to severely reduced diversity on the chromosome half with less diversity, which is consistent with extensive hitchhiking along with a selective sweep. Frequency-spectrum based tests identified 82 genes with a signature of adaptive evolution in one species or another but none of the genes were identified in both species. Based upon available functional information, several genes identified as targets of selection are likely to alter the symbiosis with the host plant, making them attractive targets for further functional characterization.
I think the author summary is a bit more, well, friendly:
Facultative mutualisms are relationships between two species that can live independently, but derive benefits when living together with their mutualistic partners. The facultative mutualism between rhizobial bacteria and legume plants contributes approximately half of all biologically fixed nitrogen, an essential plant nutrient, and is an important source of nitrogen to both natural and agricultural ecosystems. We resequenced the genomes of 44 strains of two closely related species of the genus Sinorhizobium that form facultative mutualisms with the model legme Medicago truncatula. These data provide one of the most complete examinations of genomic diversity segregating within microbial species that are not causative agents of human illness. Our analyses reveal that horizontal gene transfer, a common source of new genes in microbial species, disproportionately affects genes with direct roles in the rhizobia-plant symbiosis. Analyses of nucleotide diversity segregating within each species suggests that strong selection, along with genetic hitchhiking has sharply reduced diversity along an entire chromosome half in S. meliloti. Despite the two species' ecological similarity, we did not find evidence for selection acting on the same genetic targets. In addition to providing insight into the evolutionary history of rhizobial, this study shows the feasibility and potential power of applying population genomic analyses to microbial species.
I have highlighted the section dissing pathogen studies ...

As with every good paper, it starts with a tree

Figure 1. Neighbor-joining trees showing relationships among 32 S. meliloti (blue squares) and 12 S. medicae (red circles). 
A) chromosomes, B) pSymA and pSMED02, and C) pSymB and pSMED01. Trees were constructed using sequences from coding regions only. The length of the branch separating S. medicae from S. meliloti strains is shown at a scale that is 5% of the true scale. The 24-strain S. meliloti group is marked by asterisks. All branches had 100% bootstrap support unless otherwise indicated. Branches with <80% bootstrap support were collapsed into polytomies. An identical tree with strain identifications is provided as Figure S2.

The tree lays out the phylogeny of the strains sequenced in this study.  And it provides the main framework for much of the rest of the paper.  

Some comments:
  • The genomes were sequenced to ~ 100x coverage with on an Illumina GAIIx.
  • Reads were then aligned to reference genomes of close relatives of the sequenced strains.
  • These alignments were then used for various comparative and population genetic analyses
  • As far as I can tell no de novo assemblies were done
  • I am quite confused about their methods for detecting putative regions that have undergone horizontal gene transfer:
    • In the methods: "We identified genes likely to have experienced recent horizontal gene transfer by comparing the ratio of polymorphisms that were shared between species to fixed differences between species. Based on the whole-genome distribution of this ratio (Figure S3) we identified putatively transferred genes as those with a ratio of shared polymorphisms to fixed differences >0.2."
    • Not sure how/why this should work.  Not saying it is a bad idea - I just don't really understand it.
  • They also examine various population genetic parameters including possible selection, SNPs, Tajima's D, and more. 
It is worth a read.  They summarize their various findings with:
Population genetic analyses of nucleotide diversity segregating within Sinorhizobium medicae and S. meliloti have provided unprecedented insight into the evolutionary history of these ecologically important facultative symbionts. While previous analyses have detected evidence for horizontal gene transfer between these species, our data reveal that gene transfer is restricted almost exclusively to plasmid genes and that the plasmid regions that show evidence of transfer have less interspecific divergence than other genomic regions. Interestingly, nucleotide variation segregating within a 24-strain subpopulation of S. meliloti is highly structured along the chromosome, with one half of the chromosome harboring approximately one-fifth as much diversity as the other. The causes of the difference between the two chromosome halves may be a selective sweep coupled with extensive hitchhiking, if this is correct it would suggest that bouts of strong selection may be important in driving the divergence of bacterial species. Finally, we've identified genes that bear a signature of having evolved in response to recent positive selection. Functional characterization of these genes will provide insight into the selective forces that drive rhizobial adaptation.


  1. Looks very interesting and will certainly read it. Only question that I have based on your blog post: why a Neighbor-Joining tree and why not Maximum Likelihood?

    1. What is so bad about NJ? (OK, we don't really use it anymore ... but I always liked it)

    2. Probably because a tree is an imperfect representation of genomic polymorphism data no matter how you construct it, so you may as well go with the simple, fast method.

    3. Well, trees are imperfect - sure - but they can be very useful and thus I would prefer closer to accurate vs. fast

  2. I would always prefer to use ML and if computational power is limiting an algorithm like Fasttree (approximate ML), which outperforms NJ. But very nice paper and I don't think using different phylogenetic algorithms would have changed any of the authors conclusions.