The evolutionary genomics of adaptation to stress in wild rhizobium bacteria.

Microbiota comprise the bulk of life's diversity, yet we know little about how populations of microbes accumulate adaptive diversity across natural landscapes. Adaptation to stressful soil conditions in plants provides seminal examples of adaptation in response to natural selection via allelic substitution. For microbes symbiotic with plants however, horizontal gene transfer allows for adaptation via gene gain and loss, which could generate fundamentally different evolutionary dynamics. We use comparative genomics and genetics to elucidate the evolutionary mechanisms of adaptation to physiologically stressful serpentine soils in rhizobial bacteria in western North American grasslands. In vitro experiments demonstrate that the presence of a locus of major effect, the nre operon, is necessary and sufficient to confer adaptation to nickel, a heavy metal enriched to toxic levels in serpentine soil, and a major axis of environmental soil chemistry variation. We find discordance between inferred evolutionary histories of the core genome and nreAXY genes, which often reside in putative genomic islands. This suggests that the evolutionary history of this adaptive variant is marked by frequent losses, and/or gains via horizontal acquisition across divergent rhizobium clades. However, different nre alleles confer distinct levels of nickel resistance, suggesting allelic substitution could also play a role in rhizobium adaptation to serpentine soil. These results illustrate that the interplay between evolution via gene gain and loss and evolution via allelic substitution may underlie adaptation in wild soil microbiota. Both processes are important to consider for understanding adaptive diversity in microbes and improving stress-adapted microbial inocula for human use.

Hosts winnow symbionts with multiple layers of absolute and conditional discrimination mechanisms.

In mutualism, hosts select symbionts via partner choice and preferentially direct more resources to symbionts that provide greater benefits via sanctions. At the initiation of symbiosis, prior to resource exchange, it is not known how the presence of multiple symbiont options (i.e. the symbiont social environment) impacts partner choice outcomes. Furthermore, little research addresses whether hosts primarily discriminate among symbionts via sanctions, partner choice or a combination. We inoculated the legume, Acmispon wrangelianus, with 28 pairs of fluorescently labelled Mesorhizobium strains that vary continuously in quality as nitrogen-fixing symbionts. We find that hosts exert robust partner choice, which enhances their fitness. This partner choice is conditional such that a strain's success in initiating nodules is impacted by other strains in the social environment. This social genetic effect is as important as a strain's own genotype in determining nodulation and has both transitive (consistent) and intransitive (idiosyncratic) effects on the probability that a symbiont will form a nodule. Furthermore, both absolute and conditional partner choice act in concert with sanctions, among and within nodules. Thus, multiple forms of host discrimination act as a series of sieves that optimize host benefits and select for costly symbiont cooperation in mixed symbiont populations.

Evolution of manipulative microbial behaviors in the rhizosphere.

The rhizosphere has been called "one of the most complex ecosystems on earth" because it is a hotspot for interactions among millions of microbial cells. Many of these are microbes are also participating in a dynamic interplay with host plant tissues, signaling pathways, and metabolites. Historically, breeders have employed a plant-centric perspective when trying to harness the potential of microbiome-derived benefits to improve productivity and resilience of economically important plants. This is potentially problematic because: (i) the evolution of the microbes themselves is often ignored, and (ii) it assumes that the fitness of interacting plants and microbes is strictly aligned. In contrast, a microbe-centric perspective recognizes that putatively beneficial microbes are still under selection to increase their own fitness, even if there are costs to the host. This can lead to the evolution of sophisticated, potentially subtle, ways for microbes to manipulate the phenotype of their hosts, as well as other microbes in the rhizosphere. We illustrate this idea with a review of cases where rhizosphere microbes have been demonstrated to directly manipulate host root growth, architecture and exudation, host nutrient uptake systems, and host immunity and defense. We also discuss indirect effects, whereby fitness outcomes for the plant are a consequence of ecological interactions between rhizosphere microbes. If these consequences are positive for the plant, they can potentially be misconstrued as traits that have evolved to promote host growth, even if they are a result of selection for unrelated functions. The ubiquity of both direct microbial manipulation of hosts and context-dependent, variable indirect effects leads us to argue that an evolutionary perspective on rhizosphere microbial ecology will become increasingly important as we continue to engineer microbial communities for crop production.

Transgenic Soybeans Expressing Phosphatidylinositol-3-Phosphate-Binding Proteins Show Enhanced Resistance Against the Oomycete Pathogen Phytophthora sojae.

Oomycete and fungal pathogens cause billions of dollars of damage to crops worldwide annually. Therefore, there remains a need for broad-spectrum resistance genes, especially ones that target pathogens but do not interfere with colonization by beneficial microbes. Motivated by evidence suggesting that phosphatidylinositol-3-phosphate (PI3P) may be involved in the delivery of some oomycete and fungal virulence effector proteins, we created stable transgenic soybean plants that express and secrete two different PI3P-binding proteins, GmPH1 and VAM7, in an effort to interfere with effector delivery and confer resistance. Soybean plants expressing the two PI3P-binding proteins exhibited reduced infection by the oomycete pathogen Phytophthora sojae compared to control lines. Measurements of nodulation by nitrogen-fixing mutualistic bacterium Bradyrhizobium japonicum, which does not produce PI3P, revealed that the two lines with the highest levels of GmPH1 transcripts exhibited reductions in nodulation and in benefits from nodulation. Transcriptome and plant hormone measurements were made of soybean lines with the highest transcript levels of GmPH1 and VAM7, as well as controls, following P. sojae- or mock-inoculation. The results revealed increased levels of infection-associated transcripts in the transgenic lines, compared to controls, even prior to P. sojae infection, suggesting that the plants were primed for increased defense. The lines with reduced nodulation exhibited elevated levels of jasmonate-isoleucine and of transcripts of a JAR1 ortholog encoding jasmonate-isoleucine synthetase. However, lines expressing VAM7 transgenes exhibited normal nodulation and no increases in jasmonate-isoleucine. Overall, together with previously published data from cacao and from P. sojae transformants, the data suggest that secretion of PI3P-binding proteins may confer disease resistance through a variety of mechanisms.

Negotiating mutualism: A locus for exploitation by rhizobia has a broad effect size distribution and context-dependent effects on legume hosts.

In mutualisms, variation at genes determining partner fitness provides the raw material upon which coevolutionary selection acts, setting the dynamics and pace of coevolution. However, we know little about variation in the effects of genes that underlie symbiotic fitness in natural mutualist populations. In some species of legumes that form root nodule symbioses with nitrogen-fixing rhizobial bacteria, hosts secrete nodule-specific cysteine-rich (NCR) peptides that cause rhizobia to differentiate in the nodule environment. However, rhizobia can cleave NCR peptides through the expression of genes like the plasmid-borne Host range restriction peptidase (hrrP), whose product degrades specific NCR peptides. Although hrrP activity can confer host exploitation by depressing host fitness and enhancing symbiont fitness, the effects of hrrP on symbiosis phenotypes depend strongly on the genotypes of the interacting partners. However, the effects of hrrP have yet to be characterised in a natural population context, so its contribution to variation in wild mutualist populations is unknown. To understand the distribution of effects of hrrP in wild rhizobia, we measured mutualism phenotypes conferred by hrrP in 12 wild Ensifer medicae strains. To evaluate context dependency of hrrP effects, we compared hrrP effects across two Medicago polymorpha host genotypes and across two experimental years for five E. medicae strains. We show for the first time in a natural population context that hrrP has a wide distribution of effect sizes for many mutualism traits, ranging from strongly positive to strongly negative. Furthermore, we show that hrrP effect size varies across host genotypes and experiment years, suggesting that researchers should be cautious about extrapolating the role of genes in natural populations from controlled laboratory studies of single genetic variants.

Evolution of specialization in a plant-microbial mutualism is explained by the oscillation theory of speciation.

Specialization in mutualisms is thought to be a major driver of diversification, but few studies have explored how novel specialization evolves, or its relation to the evolution of other niche axes. A fundamental question is whether generalist interactions evolve to become more specialized (i.e., oscillation hypothesis) or if partner switches evolve without any change in niche breadth (i.e., musical chairs hypothesis). We examined alternative models for the evolution of specialization by estimating the mutualistic, climatic, and edaphic niche breadths of sister plant species, combining phylogenetic, environmental, and experimental data on Acmispon strigosus and Acmispon wrangelianus genotypes across their overlapping ranges in California. We found that specialization along all three niche axes was asymmetric across species, such that the species with broader climatic and edaphic niches, Acmispon strigosus, was also able to gain benefit from and invest in associating with a broader set of microbial mutualists. Our data are consistent with the oscillation model of specialization, and a parallel narrowing of the edaphic, climatic, and mutualistic dimensions of the host species niche. Our findings provide novel evidence that the evolution of specialization in mutualism is accompanied by specialization in other niche dimensions.

Decreased coevolutionary potential and increased symbiont fecundity during the biological invasion of a legume-rhizobium mutualism.

Although most invasive species engage in mutualism, we know little about how mutualism evolves as partners colonize novel environments. Selection on cooperation and standing genetic variation for mutualism traits may differ between a mutualism's invaded and native ranges, which could alter cooperation and coevolutionary dynamics. To test for such differences, we compare mutualism traits between invaded- and native-range host-symbiont genotype combinations of the weedy legume, Medicago polymorpha, and its nitrogen-fixing rhizobium symbiont, Ensifer medicae, which have coinvaded North America. We find that mutualism benefits for plants are indistinguishable between invaded- and native-range symbioses. However, rhizobia gain greater fitness from invaded-range mutualisms than from native-range mutualisms, and this enhancement of symbiont fecundity could increase the mutualism's spread by increasing symbiont availability during plant colonization. Furthermore, mutualism traits in invaded-range symbioses show lower genetic variance and a simpler partitioning of genetic variance between host and symbiont sources, compared to native-range symbioses. This suggests that biological invasion has reduced mutualists' potential to respond to coevolutionary selection. Additionally, rhizobia bearing a locus (hrrP) that can enhance symbiotic fitness have more exploitative phenotypes in invaded-range than in native-range symbioses. These findings highlight the impacts of biological invasion on the evolution of mutualistic interactions.

Agriculture and the Disruption of Plant-Microbial Symbiosis.

Domestication has transformed hundreds of wild plant species into productive cultivars for human utility. However, cultivation practices and intense artificial selection for yield may entail a hidden cost: the disruption of interactions between plants and beneficial microbiota. Here, we synthesize theory predicting that evolutionary trade-offs, genetic costs, and relaxed selection disrupt plant-microbial symbiosis under domestication, and review the wealth of new data interrogating these predictions in crops. We describe the agronomic practices, ecological scenarios, and genomic attributes that can result in the disruption of symbiosis, and highlight new work probing its molecular basis. To improve agricultural output and sustainability, research should develop breeding methods to optimize symbiotic outcomes in crop species.

A high-throughput method of analyzing multiple plant defensive compounds in minimized sample mass.

PREMISE OF THE STUDY: Current methods for quantifying herbivore-induced alterations in plant biochemistry are often unusable by researchers due to practical constraints. We present a cost-effective, high-throughput protocol to quantify multiple biochemical responses from small plant tissue samples using spectrophotometric techniques. METHODS AND RESULTS: Using Solanum lycopersicum and Medicago polymorpha leaves pre- and post-herbivory, we demonstrate that our protocol quantifies common plant defense responses: peroxidase production, polyphenol oxidase production, reactive oxygen species production, total protein production, and trypsin-like protease inhibition activity. CONCLUSIONS: Current protocols can require 500 mg of tissue, but our assays detect activity in less than 10 mg. Our protocol takes two people 6 h to run any of the assays on 300 samples in triplicate, or all of the assays on 20 samples. Our protocol enables researchers to plan complex experiments that compare local versus systemic plant responses, quantify environmental and genetic variation, and measure population-level variation.

Dynamic genomic architecture of mutualistic cooperation in a wild population of Mesorhizobium.

Research on mutualism seeks to explain how cooperation can be maintained when uncooperative mutants co-occur with cooperative kin. Gains and losses of the gene modules required for cooperation punctuate symbiont phylogenies and drive lifestyle transitions between cooperative symbionts and uncooperative free-living lineages over evolutionary time. Yet whether uncooperative symbionts commonly evolve from within cooperative symbiont populations or from within distantly related lineages with antagonistic or free-living lifestyles (i.e., third-party mutualism exploiters or parasites), remains controversial. We use genomic data to show that genotypes that differ in the presence or absence of large islands of symbiosis genes are common within a single wild recombining population of Mesorhizobium symbionts isolated from host tissues and are an important source of standing heritable variation in cooperation in this population. In a focal population of Mesorhizobium, uncooperative variants that lack a symbiosis island segregate at 16% frequency in nodules, and genome size and symbiosis gene number are positively correlated with cooperation. This finding contrasts with the genomic architecture of variation in cooperation in other symbiont populations isolated from host tissues in which the islands of genes underlying cooperation are ubiquitous and variation in cooperation is primarily driven by allelic substitution and individual gene gain and loss events. Our study demonstrates that uncooperative mutants within mutualist populations can comprise a significant component of genetic variation in nature, providing biological rationale for models and experiments that seek to explain the maintenance of mutualism in the face of non-cooperators.

Rapid establishment of a flowering cline in Medicago polymorpha after invasion of North America.

To establish and spread in a new location, an invasive species must be able to carry out its life cycle in novel environmental conditions. A key trait underlying fitness is the shift from vegetative to reproductive growth through floral development. In this study, we used a common garden experiment and genotyping-by-sequencing to test whether the latitudinal flowering cline of the North American invasive plant Medicago polymorpha was translocated from its European native range through multiple introductions, or whether the cline rapidly established due to evolution following a genetic bottleneck. Analysis of flowering time in 736 common garden plants showed a latitudinal flowering time cline in both the native and invaded ranges where genotypes from lower latitudes flowered earlier. Genotyping-by-sequencing of 9,658 SNPs in 446 individuals revealed two major subpopulations of M. polymorpha in the native range, only one of which is present in the invaded range. Additionally, native range populations have higher genetic diversity than invaded range populations, suggesting that a genetic bottleneck occurred during invasion. All invaded range individuals are closely related to plants collected from native range populations in Portugal and southern Spain, and population assignment tests assigned invaded range individuals to this same narrow source region. Taken together, our results suggest that latitudinal clinal variation in flowering time has rapidly evolved across the invaded range despite a genetic bottleneck following introduction.

Co-invading symbiotic mutualists of Medicago polymorpha retain high ancestral diversity and contain diverse accessory genomes.

Exotic, invasive plants and animals can wreak havoc on ecosystems by displacing natives and altering environmental conditions. However, much less is known about the identities or evolutionary dynamics of the symbiotic microbes that accompany invasive species. Most leguminous plants rely upon symbiotic rhizobium bacteria to fix nitrogen and are incapable of colonizing areas devoid of compatible rhizobia. We compare the genomes of symbiotic rhizobia in a portion of the legume's invaded range with those of the rhizobium symbionts from across the legume's native range. We show that in an area of California the legume Medicago polymorpha has invaded, its Ensifer medicae symbionts: (i) exhibit genome-wide patterns of relatedness that together with historical evidence support host-symbiont co-invasion from Europe into California, (ii) exhibit population genomic patterns consistent with the introduction of the majority of deep diversity from the native range, rather than a genetic bottleneck during colonization of California and (iii) harbor a large set of accessory genes uniquely enriched in binding functions, which could play a role in habitat invasion. Examining microbial symbiont genome dynamics during biological invasions is critical for assessing host-symbiont co-invasions whereby microbial symbiont range expansion underlies plant and animal invasions.

Invasive legumes can associate with many mutualists of native legumes, but usually do not.

Mutualistic interactions can strongly influence species invasions, as the inability to form successful mutualisms in an exotic range could hamper a host's invasion success. This barrier to invasion may be overcome if an invader either forms novel mutualistic associations or finds and associates with familiar mutualists in the exotic range. Here, we ask (1) does the community of rhizobial mutualists associated with invasive legumes in their exotic range overlap with that of local native legumes and (2) can any differences be explained by fundamental incompatibilities with particular rhizobial genotypes? To address these questions, we first characterized the rhizobial communities naturally associating with three invasive and six native legumes growing in the San Francisco Bay Area. We then conducted a greenhouse experiment to test whether the invasive legume could nodulate with any of a broad array of rhizobia found in their exotic range. There was little overlap between the Bradyrhizobium communities associated with wild-grown invasive and native legumes, yet the invasive legumes could nodulate with a broad range of rhizobial strains under greenhouse conditions. These observations suggest that under field conditions in their exotic range, these invasive legumes are not currently associating with the mutualists of local native legumes, despite their potential to form such associations. However, the promiscuity with which these invading legumes can form mutualistic associations could be an important factor early in the invasion process if mutualist scarcity limits range expansion. Overall, the observation that invasive legumes have a community of rhizobia distinct from that of native legumes, despite their ability to associate with many rhizobial strains, challenges existing assumptions about how invading species obtain their mutualists. These results can therefore inform current and future efforts to prevent and remove invasive species.

Association mapping reveals novel serpentine adaptation gene clusters in a population of symbiotic Mesorhizobium.

The genetic variants that underlie microbial environmental adaptation are key components of models of microbial diversification. Characterizing adaptive variants and the pangenomic context in which they evolve remains a frontier in understanding how microbial diversity is generated. The genomics of rhizobium adaptation to contrasting soil environments is ecologically and agriculturally important because these bacteria are responsible for half of all current biologically fixed nitrogen, yet they live the majority of their lives in soil. Our study uses whole-genome sequencing to describe the pan-genome of a focal clade of wild mesorhizobia that show contrasting levels of nickel adaptation despite high relatedness (99.8% identity at 16S). We observe ecotypic specialization within an otherwise genomically cohesive population, rather than finding distinct specialized bacterial lineages in contrasting soil types. This finding supports recent reports that heterogeneous environments impose selection that maintains differentiation only at a small fraction of the genome. Our work further uses a genome-wide association study to propose candidate genes for nickel adaptation. Several candidates show homology to genetic systems involved in nickel tolerance and one cluster of candidates correlates perfectly with soil origin, which validates our approach of ascribing genomic variation to adaptive divergence.

Cheaters must prosper: reconciling theoretical and empirical perspectives on cheating in mutualism.

Cheating is a focal concept in the study of mutualism, with the majority of researchers considering cheating to be both prevalent and highly damaging. However, current definitions of cheating do not reliably capture the evolutionary threat that has been a central motivation for the study of cheating. We describe the development of the cheating concept and distill a relative-fitness-based definition of cheating that encapsulates the evolutionary threat posed by cheating, i.e. that cheaters will spread and erode the benefits of mutualism. We then describe experiments required to conclude that cheating is occurring and to quantify fitness conflict more generally. Next, we discuss how our definition and methods can generate comparability and integration of theory and experiments, which are currently divided by their respective prioritisations of fitness consequences and traits. To evaluate the current empirical evidence for cheating, we review the literature on several of the best-studied mutualisms. We find that although there are numerous observations of low-quality partners, there is currently very little support from fitness data that any of these meet our criteria to be considered cheaters. Finally, we highlight future directions for research on conflict in mutualisms, including novel research avenues opened by a relative-fitness-based definition of cheating.

The ecological genomic basis of salinity adaptation in Tunisian Medicago truncatula.

BACKGROUND: As our world becomes warmer, agriculture is increasingly impacted by rising soil salinity and understanding plant adaptation to salt stress can help enable effective crop breeding. Salt tolerance is a complex plant phenotype and we know little about the pathways utilized by naturally tolerant plants. Legumes are important species in agricultural and natural ecosystems, since they engage in symbiotic nitrogen-fixation, but are especially vulnerable to salinity stress. RESULTS: Our studies of the model legume Medicago truncatula in field and greenhouse settings demonstrate that Tunisian populations are locally adapted to saline soils at the metapopulation level and that saline origin genotypes are less impacted by salt than non-saline origin genotypes; these populations thus likely contain adaptively diverged alleles. Whole genome resequencing of 39 wild accessions reveals ongoing migration and candidate genomic regions that assort non-randomly with soil salinity. Consistent with natural selection acting at these sites, saline alleles are typically rare in the range-wide species' gene pool and are also typically derived relative to the sister species M. littoralis. Candidate regions for adaptation contain genes that regulate physiological acclimation to salt stress, such as abscisic acid and jasmonic acid signaling, including a novel salt-tolerance candidate orthologous to the uncharacterized gene AtCIPK21. Unexpectedly, these regions also contain biotic stress genes and flowering time pathway genes. We show that flowering time is differentiated between saline and non-saline populations and may allow salt stress escape. CONCLUSIONS: This work nominates multiple potential pathways of adaptation to naturally stressful environments in a model legume. These candidates point to the importance of both tolerance and avoidance in natural legume populations. We have uncovered several promising targets that could be used to breed for enhanced salt tolerance in crop legumes to enhance food security in an era of increasing soil salinization.

Selection for cheating across disparate environments in the legume-rhizobium mutualism.

The primary dilemma in evolutionarily stable mutualisms is that natural selection for cheating could overwhelm selection for cooperation. Cheating need not entail parasitism; selection favours cheating as a quantitative trait whenever less-cooperative partners are more fit than more-cooperative partners. Mutualisms might be stabilised by mechanisms that direct benefits to more-cooperative individuals, which counter selection for cheating; however, empirical evidence that natural selection favours cheating in mutualisms is sparse. We measured selection on cheating in single-partner pairings of wild legume and rhizobium lineages, which prevented legume choice. Across contrasting environments, selection consistently favoured cheating by rhizobia, but did not favour legumes that provided less benefit to rhizobium partners. This is the first simultaneous measurement of selection on cheating across both host and symbiont lineages from a natural population. We empirically confirm selection for cheating as a source of antagonistic coevolutionary pressure in mutualism and a biological dilemma for models of cooperation.

Specialization-generalization trade-off in a Bradyrhizobium symbiosis with wild legume hosts.

BACKGROUND: Specialized interactions help structure communities, but persistence of specialized organisms is puzzling because a generalist can occupy more environments and partake in more beneficial interactions. The "Jack-of-all-trades is a master of none" hypothesis asserts that specialists persist because the fitness of a generalist utilizing a particular habitat is lower than that of a specialist adapted to that habitat. Yet, there are many reasons to expect that mutualists will generalize on partners.Plant-soil feedbacks help to structure plant and microbial communities, but how frequently are soil-based symbiotic mutualistic interactions sufficiently specialized to influence species distributions and community composition? To address this question, we quantified realized partner richness and phylogenetic breadth of four wild-grown native legumes (Lupinus bicolor, L. arboreus, Acmispon strigosus and A. heermannii) and performed inoculation trials to test the ability of two hosts (L. bicolor and A. strigosus) to nodulate (fundamental partner richness), benefit from (response specificity), and provide benefit to (effect specificity) 31 Bradyrhizobium genotypes. RESULTS: In the wild, each Lupinus species hosted a broader genetic range of Bradyrhizobium than did either Acmispon species, suggesting that Acmispon species are more specialized. In the greenhouse, however, L. bicolor and A. strigosus did not differ in fundamental association specificity: all inoculated genotypes nodulated both hosts. Nevertheless, A. strigosus exhibited more specificity, i.e., greater variation in its response to, and effect on, Bradyrhizobium genotypes. Lupinus bicolor benefited from a broader range of genotypes but averaged less benefit from each. Both hosts obtained more fitness benefit from symbionts isolated from conspecific hosts; those symbionts in turn gained greater fitness benefit from hosts of the same species from which they were isolated. CONCLUSIONS: This study affirmed two important tenets of evolutionary theory. First, as predicted by the Jack-of-all-trades is a master of none hypothesis, specialist A. strigosus obtained greater benefit from its beneficial symbionts than did generalist L. bicolor. Second, as predicted by coevolutionary theory, each test species performed better with partner genotypes isolated from conspecifics. Finally, positive fitness feedback between the tested hosts and symbionts suggests that positive plant-soil feedback could contribute to their patchy distributions in this system.

Trade-offs, spatial heterogeneity, and the maintenance of microbial diversity.

Specialization and concomitant trade-offs are assumed to underlie the non-neutral coexistence of lineages. Trade-offs across heterogeneous environments can promote diversity by preventing competitive exclusion. However, the importance of trade-offs in maintaining diversity in natural microbial assemblages is unclear, as trade-offs are frequently not detected in artificial evolution experiments. Stressful conditions associated with patches of heavy-metal enriched serpentine soils provide excellent opportunities for examining how heterogeneity may foster genetic diversity. Using a spatially replicated design, we demonstrate that rhizobium bacteria symbiotic with legumes inhabiting contrasting serpentine and nonserpentine soils exhibit a trade-off between a genotype's nickel tolerance and its ability to replicate rapidly. Furthermore, we detected adaptive divergence in rhizobial assemblages across soil type heterogeneity at multiple sites, suggesting that this trade-off may promote the coexistence of phenotypically distinct bacterial lineages. Trade-offs and adaptive divergence may be important factors maintaining the tremendous diversity within natural assemblages of bacteria.

Adaptive divergence in seed color camouflage in contrasting soil environments.

Although adaptive plant population divergence across contrasting soil conditions is often driven by abiotic soil factors, natural enemies may also contribute. Cryptic matching to the native soil color is a form of defensive camouflage that seeds can use to avoid detection by seed predators. The legume Acmispon wrangelianus occurs across a variety of gray-green serpentine soils and brown nonserpentine soils. Quantitative digital image analysis of seed and soil colors was used to test whether genetically based seed color is a closer match to the color of the native soil than to the color of other nearby soils. Lineages bear seeds that more closely match the color of their native serpentine or nonserpentine soil type than the opposing soil type. Further, even within a soil type, lineages bear seeds with a closer color match to the soil at their native site than to other sites. The striking concordance between seed and native soil color suggests that natural selection for locally camouflaged seed color morphs, probably driven by seed predators, may maintain adaptive divergence in pigmentation, despite the opportunity for migration between soil environments.