Primitive agriculture in a social amoeba.

Agriculture has been a large part of the ecological success of humans. A handful of animals, notably the fungus-growing ants, termites and ambrosia beetles, have advanced agriculture that involves dispersal and seeding of food propagules, cultivation of the crop and sustainable harvesting. More primitive examples, which could be called husbandry because they involve fewer adaptations, include marine snails farming intertidal fungi and damselfish farming algae. Recent work has shown that microorganisms are surprisingly like animals in having sophisticated behaviours such as cooperation, communication and recognition, as well as many kinds of symbiosis. Here we show that the social amoeba Dictyostelium discoideum has a primitive farming symbiosis that includes dispersal and prudent harvesting of the crop. About one-third of wild-collected clones engage in husbandry of bacteria. Instead of consuming all bacteria in their patch, they stop feeding early and incorporate bacteria into their fruiting bodies. They then carry bacteria during spore dispersal and can seed a new food crop, which is a major advantage if edible bacteria are lacking at the new site. However, if they arrive at sites already containing appropriate bacteria, the costs of early feeding cessation are not compensated for, which may account for the dichotomous nature of this farming symbiosis. The striking convergent evolution between bacterial husbandry in social amoebas and fungus farming in social insects makes sense because multigenerational benefits of farming go to already established kin groups.

Genetic diversity in the social amoeba Dictyostelium discoideum: population differentiation and cryptic species.

The social amoeba Dictyostelium discoideum is a commonly used model organism for the study of social evolution, multicellularity, and cell biology. But the boundaries and structure of the species have not been explored. The lack of morphological traits to distinguish D. discoideum makes even knowing whether a given clone is D. discoideum a challenge. We address this with a phylogeny of a widespread collection of clones from a range of locations and including clones identified previously as potential cryptic species. We sequenced portions of nuclear ribosomal DNA and mitochondrial DNA, analyzing approximately 5500 and 2500 base pairs from the two regions respectively. We compared these sequences to known reference sequences for both D. discoideum and other closely related Dictyostelium species to create Bayesian and neighbor-joining phylogenetic trees representing the evolutionary relationships among the clones. We identified 51 unique D. discoideum concatenated sequences based on the combined mitochondrial and ribosomal sequence data. We also identified four unique D. citrinum concatenated sequences, three of which were previously classified as D. discoideum clones. Our analysis of the data revealed that all D. discoideum clones form a monophyletic group, but there are several well-supported subclades and pronounced genetic differentiation among locations (F(ST)=0.242, P=0.011), suggesting the presence of geographic or other barriers between populations. Our results reveal the need for further investigation into potential tropical cryptic species.

Diversity of Free-Living Environmental Bacteria and Their Interactions With a Bactivorous Amoeba.

A small subset of bacteria in soil interact directly with eukaryotes. Which ones do so can reveal what is important to a eukaryote and how eukaryote defenses might be breached. Soil amoebae are simple eukaryotic organisms and as such could be particularly good for understanding how eukaryote microbiomes originate and are maintained. One such amoeba, Dictyostelium discoideum, has both permanent and temporary associations with bacteria. Here we focus on culturable bacterial associates in order to interrogate their relationship with D. discoideum. To do this, we isolated over 250 D. discoideum fruiting body samples from soil and deer feces at Mountain Lake Biological Station. In one-third of the wild D. discoideum we tested, one to six bacterial species were found per fruiting body sorus (spore mass) for a total of 174 bacterial isolates. The remaining two-thirds of D. discoideum fruiting body samples did not contain culturable bacteria, as is thought to be the norm. A majority (71.4%) of the unique bacterial haplotypes are in Proteobacteria. The rest are in either Actinobacteria, Bacteriodetes, or Firmicutes. The highest bacterial diversity was found in D. discoideum fruiting bodies originating from deer feces (27 OTUs), greater than either of those originating in shallow (11 OTUs) or in deep soil (4 OTUs). Rarefaction curves and the Chao1 estimator for species richness indicated the diversity in any substrate was not fully sampled, but for soil it came close. A majority of the D. discoideum-associated bacteria were edible by D. discoideum and supported its growth (75.2% for feces and 81.8% for soil habitats). However, we found several bacteria genera were able to evade phagocytosis and persist in D. discoideum cells through one or more social cycles. This study focuses not on the entire D. discoideum microbiome, but on the culturable subset of bacteria that have important eukaryote interactions as prey, symbionts, or pathogens. These eukaryote and bacteria interactions may provide fertile ground for investigations of bacteria using amoebas to gain an initial foothold in eukaryotes and of the origins of symbiosis and simple microbiomes.

Aristaless Controls Butterfly Wing Color Variation Used in Mimicry and Mate Choice.

Neotropical Heliconius butterflies display a diversity of warningly colored wing patterns, which serve roles in both Müllerian mimicry and mate choice behavior. Wing pattern diversity in Heliconius is controlled by a small number of unlinked, Mendelian "switch" loci [1]. One of these, termed the K locus, switches between yellow and white color patterns, important mimicry signals as well as mating cues [2-4]. Furthermore, mate preference behavior is tightly linked to this locus [4]. K controls the distribution of white versus yellow scales on the wing, with a dominant white allele and a recessive yellow allele. Here, we combine fine-scale genetic mapping, genome-wide association studies, gene expression analyses, population and comparative genomics, and genome editing with CRISPR/Cas9 to characterize the molecular basis of the K locus in Heliconius and to infer its evolutionary history. We show that white versus yellow color variation in Heliconius cydno is due to alternate haplotypes at a putative cis-regulatory element (CRE) downstream of a tandem duplication of the homeodomain transcription factor aristaless. Aristaless1 (al1) and aristaless2 (al2) are differentially regulated between white and yellow wings throughout development with elevated expression of al1 in developing white wings, suggesting a role in repressing pigmentation. Consistent with this, knockout of al1 causes white wings to become yellow. The evolution of wing color in this group has been marked by retention of the ancestral yellow color in many lineages, a single origin of white coloration in H. cydno, and subsequent introgression of white color from H. cydno into H. melpomene.

Collection and cultivation of dictyostelids from the wild.

Dictyostelium discoideum is a commonly used model organism for the study of biological processes such as chemotaxis, cell communication, and development. While these studies primarily focus on a single clone, recent work has revealed a host of questions that can only be answered from studies of multiple genetically distinct clones. Understanding intraspecific clone conflict, kin recognition, differential adhesion, and other kinds of interactions likely to occur in the natural soil habitat can only come from studies of multiple clones. Studies of populations of wild isolates are also important for understanding the factors contributing to associations such as species co-occurrences and to observed inter- and intraspecific interactions such as those found between bacteria and D. discoideum. Natural isolates of Dictyostelium are easily found in soil and leaf litter in nearly all habitats. Here we describe a simple and successful method for isolating new wild clones from soil, then isolating single clonal strains, and storing them for future use.