Vertical eDNA distribution of cold-water fishes in response to environmental variables in stratified lake.

In summer, the survival zones of cold-water species are predicted to narrow by both increasing water temperatures from the surface and by expanding hypoxic zones from the lake bottom. To examine how the abundance of cold-water fishes changes along environmental gradients, we assessed the vertical environmental DNA (eDNA) distributions of three salmonid species which may have different water temperature tolerances during both stratification and turnover periods using quantitative PCR (qPCR). In addition, we examined on the vertical distribution of diverse fish fauna using an eDNA metabarcoding assay. The results suggested that the kokanee salmon (Oncorhynchus nerka) eDNA were abundant in deep, cold waters. On the other hand, rainbow trout (O. mykiss) eDNA were distributed uniformly throughout the water column, suggesting that they may have high water-temperature tolerance compared with kokanee salmon. The eDNA concentrations of masu salmon (O. masou) were below the detection limit (i.e., <10 copies µL-1) at all stations and depths and hence could not be quantified during stratification. Together with the finding that the eDNA distributions of other prey fish species were also constrained vertically in species-specific ways, our results suggest that climate change will result in substantial changes in the vertical distributions of lake fish species and thus affect their populations and interactions.

Evolutionary Dynamics of Host Organs for Microbial Symbiosis in Tortoise Leaf Beetles (Coleoptera: Chrysomelidae: Cassidinae).

Diverse insects host specific microbial symbionts that play important roles for their growth, survival, and reproduction. They often develop specialized symbiotic organs for harboring the microbial partners. While such intimate associations tend to be stably maintained over evolutionary time, the microbial symbionts may have been lost or replaced occasionally. How symbiont acquisitions, replacements, and losses are linked to the development of the host's symbiotic organs is an important but poorly understood aspect of microbial symbioses. Cassidine leaf beetles are associated with a specific gammaproteobacterial lineage, Stammera, whose reduced genome is streamlined for producing pectin-degrading enzymes to assist the host's digestion of food plants. We investigated the symbiotic system of 24 Japanese cassidine species and found that (i) most species harbored Stammera within paired symbiotic organs located at the foregut-midgut junction, (ii) the host phylogeny was largely congruent with the symbiont phylogeny, indicating stable host-symbiont association over evolutionary time, (iii) meanwhile, the symbiont was not detected in three distinct host lineages, uncovering recurrent losses of the ancient microbial mutualist, (iv) the symbiotic organs were vestigial but present in the symbiont-free lineages, indicating evolutionary persistence of the symbiotic organs even in the absence of the symbiont, and (v) the number of the symbiotic organs was polymorphic among the cassidine species, either two or four, unveiling a dynamic evolution of the host organs for symbiosis. These findings are discussed as to what molecular mechanisms and evolutionary trajectories underpin the recurrent symbiont losses and the morphogenesis of the symbiotic organs in the herbivorous insect group. IMPORTANCE Insects represent the biodiversity of the terrestrial ecosystem, and their prosperity is attributable to their association with symbiotic microorganisms. By sequestering microbial functionality into their bodies, organs, tissues, or cells, diverse insects have successfully exploited otherwise inaccessible ecological niches and resources, including herbivory enabled by utilization of indigestible plant cell wall components. In leaf beetles of the subfamily Cassininae, an ancient symbiont lineage, Stammera, whose genome is extremely reduced and specialized for encoding pectin-degrading enzymes, is hosted in gut-associated symbiotic organs and contributes to the host's food plant digestion. Here, we demonstrate that multiple symbiont losses and recurrent structural switching of the symbiotic organs have occurred in the evolutionary course of cassidine leaf beetles, which sheds light on the evolutionary and developmental dynamics of the insect's symbiotic organs and provides a model system to investigate how microbial symbionts affect the host's development and morphogenesis and vice versa.

Bacterial symbionts support larval sap feeding and adult folivory in (semi-)aquatic reed beetles.

Symbiotic microbes can enable their host to access untapped nutritional resources but may also constrain niche space by promoting specialization. Here, we reconstruct functional changes in the evolutionary history of the symbiosis between a group of (semi-)aquatic herbivorous insects and mutualistic bacteria. Sequencing the symbiont genomes across 26 species of reed beetles (Chrysomelidae, Donaciinae) spanning four genera indicates that the genome-eroded mutualists provide life stage-specific benefits to larvae and adults, respectively. In the plant sap-feeding larvae, the symbionts are inferred to synthesize most of the essential amino acids as well as the B vitamin riboflavin. The adult reed beetles' folivory is likely supported by symbiont-encoded pectinases that complement the host-encoded set of cellulases, as revealed by transcriptome sequencing. However, mapping the occurrence of the symbionts' pectinase genes and the hosts' food plant preferences onto the beetles' phylogeny reveals multiple independent losses of pectinase genes in lineages that switched to feeding on pectin-poor plants, presumably constraining their hosts' subsequent adaptive potential.

Symbiont Digestive Range Reflects Host Plant Breadth in Herbivorous Beetles.

Numerous adaptations are gained in light of a symbiotic lifestyle. Here, we investigated the obligate partnership between tortoise leaf beetles (Chrysomelidae: Cassidinae) and their pectinolytic Stammera symbionts to detail how changes to the bacterium's streamlined metabolic range can shape the digestive physiology and ecological opportunity of its herbivorous host. Comparative genomics of 13 Stammera strains revealed high functional conservation, highlighted by the universal presence of polygalacturonase, a primary pectinase targeting nature's most abundant pectic class, homogalacturonan (HG). Despite this conservation, we unexpectedly discovered a disparate distribution for rhamnogalacturonan lyase, a secondary pectinase hydrolyzing the pectic heteropolymer, rhamnogalacturonan I (RG-I). Consistent with the annotation of rhamnogalacturonan lyase in Stammera, cassidines are able to depolymerize RG-I relative to beetles whose symbionts lack the gene. Given the omnipresence of HG and RG-I in foliage, Stammera that encode pectinases targeting both substrates allow their hosts to overcome a greater diversity of plant cell wall polysaccharides and maximize access to the nutritionally rich cytosol. Possibly facilitated by their symbionts' expanded digestive range, cassidines additionally endowed with rhamnogalacturonan lyase appear to utilize a broader diversity of angiosperms than those beetles whose symbionts solely supplement polygalacturonase. Our findings highlight how symbiont metabolic diversity, in concert with host adaptations, may serve as a potential source of evolutionary innovations for herbivorous lineages.

Evolution of a key trait greatly affects underground community assembly process through habitat adaptation in earthworms.

Underground community assemblies have not been studied well compared with aboveground communities, despite their importance for our understanding of whole ecosystems. To investigate underground community assembly over evolutionary timescales, we examined terrestrial earthworm communities (Oligochaeta: Haplotaxida) in conserved mountainous primary forests in Japan as a model system. We collected 553 earthworms mostly from two dominant families, the Megascolecidae and the Lumbricidae, from 12 sites. We constructed a molecular taxonomic unit tree based on the analysis of three genes to examine the effects of a biogeographic factor (dispersal ability) and an evolutionary factor (habitat adaptation) on the earthworm community assembly process. The phylogenetic distance of the earthworm communities among sites was positively correlated with geographic distance when intraspecific variation was included, indicating that the divergence within species was affected by biogeographic factors. The community assembly process in the Megascolecidae has also been affected by environmental conditions in relation to an evolutionary relationship between habitat environment and intestinal cecum type, a trait closely related to habitat depth and diet, whereas that in the Lumbricidae has not been affected as such. Intestinal cecum type showed a pattern of niche conservatism in the Megascolecidae lineage. Our results suggest that investigating the evolution of a key trait related to life history can lead to the clear description of community assembly process over a long timescale and that the community assembly process can differ greatly among related lineages even though they live sympatrically.

Drastic Genome Reduction in an Herbivore's Pectinolytic Symbiont.

Pectin, an integral component of the plant cell wall, is a recalcitrant substrate against enzymatic challenges by most animals. In characterizing the source of a leaf beetle's (Cassida rubiginosa) pectin-degrading phenotype, we demonstrate its dependency on an extracellular bacterium housed in specialized organs connected to the foregut. Despite possessing the smallest genome (0.27 Mb) of any organism not subsisting within a host cell, the symbiont nonetheless retained a functional pectinolytic metabolism targeting the polysaccharide's two most abundant classes: homogalacturonan and rhamnogalacturonan I. Comparative transcriptomics revealed pectinase expression to be enriched in the symbiotic organs, consistent with enzymatic buildup in these structures following immunostaining with pectinase-targeting antibodies. Symbiont elimination results in a drastically reduced host survivorship and a diminished capacity to degrade pectin. Collectively, our findings highlight symbiosis as a strategy for an herbivore to metabolize one of nature's most complex polysaccharides and a universal component of plant tissues.

Predators regulate prey species sorting and spatial distribution in microbial landscapes.

The role of predation in determining the metacommunity assembly model of prey communities is understudied relative to that of interspecific competition among prey. Previous work on metacommunity dynamics of competing species has shown that sorting by habitat patch type and spatial patterning can be affected by disturbances. Microcosms offer a useful model system to test the effect of multi-trophic interactions and disturbance on metacommunity dynamics. Here, we investigated the potential role of predators in enhancing or disrupting sorting and spatial pattern among prey in experimental landscapes. We exposed multi-trophic protist microcosm landscapes with one predator, two competing prey, two patch resource types, and localized dispersal to three disturbance regimes (none, low, and high). Then, we used variation partitioning and spatial clustering analysis to analyse the results. In contrast with previous experiments that did not manipulate predators, we found that patch type did not structure prey communities very well. Instead, we found that it was the distribution of the predator that most strongly predicted the composition of the prey community. The predator impacted species sorting by (1) preferentially consuming one prey, thereby acting as a strong local environmental driver, and by (2) indirectly magnifying the impact of patch food resources on the less preferred prey. The predator also enhanced spatial signal in the prey community because of its limited dispersal. Our results indicate that predators can strongly influence prey species sorting and spatial patterning in metacommunities in ways that would otherwise be attributed to stochastic effects, such as dispersal limitation or demographic drift. Therefore, whenever possible, predators should be explicitly included as separate explanatory factors in variation partitioning analyses.

Disturbance-mediated colonization-extinction dynamics in experimental protist metacommunities.

Colonization-extinction dynamics and species sorting among habitats deter- mine the distribution of species within metacommunities. Theory suggests that disturbances reduce the importance of species sorting and enhance spatial patterning and stochastic effects, however this has not yet been experimentally shown. We examined how extinctions in a heterogeneous landscape of patches affects the influence of environmental, spatial, and stochastic factors on community composition in a simple two-species, two-habitat, protist metacommunity where each species dominates in a different habitat type. We imposed four different levels of random extinctions on local patches and monitored changes in the metacommunity through time. We found that near-steady state patterns of community variability developed relatively rapidly (within nine colonization-extinction cycles) and that increased extinction rate produced altered patterns of community regulation by reducing environmental control and increasing spatial and stochastic effects. Our results indicate a possible explanation for the combination of environmental, spatial and stochastic effects observed in natural metacommunities.

Remarkable spatial memory in a migratory cardinalfish.

The ability to orient and navigate within a certain environment is essential for all animals, and spatial memory enables animals to remember the locations of such markers as predators, home, and food. Here we report that the migratory marine cardinalfish Apogon notatus has the potential to retain long-term spatial memory comparable to that of other animals. Female A. notatus establish a small territory on a shallow boulder bottom to pair and spawn with males. We carried out field research in two consecutive breeding seasons on territory settlement by individually marked females. Females maintained a territory at the same site throughout one breeding season. After overwintering in deep water, many of them (82.1%) returned to their breeding ground next spring and most occupied the same site as in the previous season, with only a 0.56 m shift on average. Our results suggest that female A. notatus have long-distance homing ability to pinpoint the exact location of their previous territory, and retain spatial memory for as long as 6 months.