Noise pollution alters the diet composition of invertebrate consumers both in and beyond a noise-exposed grassland ecosystem.

Anthropogenic noise is ubiquitous globally. However, we know little about how the impacts of noise alter fundamental ecosystem properties, such as resource consumption by invertebrate consumers. Using experimental noise manipulation and faecal DNA metabarcoding, we assessed how the direct and cross-trophic indirect effects of noise altered the dietary richness and specializations of omnivorous grasshoppers in a grassland ecosystem. We found that the experimental noise treatment expanded grasshoppers' dietary richness and resulted in dietary generalizations in both noise-exposed and adjacent relatively quieter areas. Unexpectedly, however, these dietary changes were primarily explained by the direct effect of noise not only in the noise-exposed areas but also in the adjacent quieter areas and were relaxed by indirect effects of noise such as reduced birds and predation risk and increased grasshoppers. Our work suggests that noise pollution can be key in explaining the variation of invertebrate consumers' diets across a gradient of noise-exposed environments.

Temporal and interspecific dietary variation in wintering ducks in agricultural landscapes.

Farmlands are becoming more important as waterfowl foraging habitats, while natural wetlands are being lost globally. However, it is unclear how waterfowl coexist in agricultural landscapes by resource partitioning. We evaluated the diets of seven sympatric dabbling ducks foraging in rice paddy and lotus fields around Lake Kasumigaura, the second largest lake in Japan, during two wintering seasons (from November to February) by faecal DNA metabarcoding using chloroplast trnL and mitochondrial CO1 region sequences. We examined 420 faecal samples and found different patterns of dietary diversity and composition among the duck species. The pattern also differed between plant and invertebrate food. Dietary niche partitioning was clear in plant food. Large-bodied ducks intensively use crop plants, and other ducks might mediate competition by using terrestrial and aquatic plants that are suitable for their foraging behaviours or microhabitats. Dietary segregation among species was the most apparent in February, when the abundance of foraging ducks was the largest. This study illustrated the complex pattern of dietary niche partitioning of dabbling ducks in agricultural landscapes, which might be difficult to evaluate by conventional approaches. The availability of crop plants, as well as other plant food resources in flooted areas and farmland dikes, may enable ducks to coexist by spatial or behavioural resource partitioning.

Evaluating the existence and benefit of major histocompatibility complex-based mate choice in an isolated owl population.

How mate preferences evolve in the first place has been a major conundrum for sexual selection. Some hypotheses explaining this assume fitness benefit derived from subsequent generations. Major histocompatibility complex (MHC)-based mate choice is a representative example of the mate choice that is associated with such trans-generational mechanisms. To provide evidences for fitness benefit of MHC-based mate choice, previous studies assessed the association between own MHC genotype and own fitness components. However, the association between MHC-based mate choice in the parental generation and fitness components in the resultant offspring generation has only rarely been measured in wild populations. Focusing on the isolated population of the monogamous Ryukyu Scops Owl (Otus elegans interpositus) on Minami-daito Island, Japan, we found evidence of MHC-based mate choice. However, we found no evidence of MHC-based mate choice increasing own reproductive success or offspring survival. This is a rare case study that directly examines the existence of the trans-generational indirect benefit of MHC-based mate choice for genetic compatibility from trans-generational data in a wild bird population. By investigating the fitness benefits of mate choice, this study serves to facilitate our understanding of the evolution of MHC-based mate choice.

Evaluation of plant contamination in metabarcoding diet analysis of a herbivore.

Fecal DNA metabarcoding is currently used in various fields of ecology to determine animal diets. Contamination of non-food DNA from complex field environments is a considerable challenge to the reliability of this method but has rarely been quantified. We evaluated plant DNA contamination by sequencing the chloroplast trnL P6 loop region from food-controlled geese feces. The average percentage of contaminant sequences per sample was 1.86%. According to the results of generalized linear models, the probability of contamination was highest in samples placed in wet soil. The proportion of contaminant sequences was lowest at the earliest sampling point and was slightly higher in samples placed in open conditions. Exclusion of rare OTUs (operational taxonomic units) was effective for obtaining reliable dietary data from the obtained sequences, and a 1% cutoff reduced the percentage of contaminated samples to less than 30%. However, appropriate interpretation of the barcoding results considering inevitable contamination is an important issue to address. We suggest the following procedures for fecal sampling and sequence data treatment to increase the reliability of DNA metabarcoding diet analyses: (i) Collect samples as soon as possible after deposition, (ii) avoid samples from deposits on wet soil, and (iii) exclude rare OTUs from diet composition estimations.

DNA barcoding reveals seasonal shifts in diet and consumption of deep-sea fishes in wedge-tailed shearwaters.

The foraging ecology of pelagic seabirds is difficult to characterize because of their large foraging areas. In the face of this difficulty, DNA metabarcoding may be a useful approach to analyze diet compositions and foraging behaviors. Using this approach, we investigated the diet composition and its seasonal variation of a common seabird species on the Ogasawara Islands, Japan: the wedge-tailed shearwater Ardenna pacifica. We collected fecal samples during the prebreeding (N = 73) and rearing (N = 96) periods. The diet composition of wedge-tailed shearwater was analyzed by Ion Torrent sequencing using two universal polymerase chain reaction primers for the 12S and 16S mitochondrial DNA regions that targeted vertebrates and mollusks, respectively. The results of a BLAST search of obtained sequences detected 31 and 1 vertebrate and mollusk taxa, respectively. The results of the diet composition analysis showed that wedge-tailed shearwaters frequently consumed deep-sea fishes throughout the sampling season, indicating the importance of these fishes as a stable food resource. However, there was a marked seasonal shift in diet, which may reflect seasonal changes in food resource availability and wedge-tailed shearwater foraging behavior. The collected data regarding the shearwater diet may be useful for in situ conservation efforts. Future research that combines DNA metabarcoding with other tools, such as data logging, may provide further insight into the foraging ecology of pelagic seabirds.

Development of microsatellite markers for the coastal shrub Scaevola taccada (Goodeniaceae).

PREMISE OF THE STUDY: Microsatellite markers were developed for the coastal shrub species Scaevola taccada to estimate the population genetic structure, which may reflect different seed dispersal patterns. • METHODS AND RESULTS: Thirteen microsatellite primer sets were developed for S. taccada using 454 pyrosequencing. The primer sets were tested on 64 individuals sampled from two populations in Japan. Fragments were amplified using the primers, with one to 10 alleles per locus, and the expected heterozygosity ranged from 0.00 to 0.85. • CONCLUSIONS: These results indicate the utility of markers in S. taccada for broad estimations of the population genetic structure of this species.

Diet analysis by next-generation sequencing indicates the frequent consumption of introduced plants by the critically endangered red-headed wood pigeon (Columba janthina nitens) in oceanic island habitats.

Oceanic island ecosystems are vulnerable to the introduction of alien species, and they provide a habitat for many endangered species. Knowing the diet of an endangered animal is important for appropriate nature restoration efforts on oceanic islands because introduced species may be a major component of the diets of some endangered species. DNA barcoding techniques together with next-generation sequencing may provide more detailed information on animal diets than other traditional methods. We performed a diet analysis using 48 fecal samples from the critically endangered red-headed wood pigeon that is endemic to the Ogasawara Islands based on chloroplast trnL P6 loop sequences. The frequency of each detected plant taxa was compared with a microhistological analysis of the same sample set. The DNA barcoding approach detected a much larger number of plants than the microhistological analysis. Plants that were difficult to identify by microhistological analysis after being digested in the pigeon stomachs were frequently identified only by DNA barcoding. The results of the barcoding analysis indicated the frequent consumption of introduced species, in addition to several native species, by the red-headed wood pigeon. The rapid eradication of specific introduced species may reduce the food resources available to this endangered bird; thus, balancing eradication efforts with the restoration of native food plants should be considered. Although some technical problems still exist, the trnL approach to next-generation sequencing may contribute to a better understanding of oceanic island ecosystems and their conservation.

Genetic diversity of the Japanese wood pigeon, Columba janthina, endemic to islands of East Asia, estimated by newly developed microsatellite markers.

The Japanese wood pigeon Columba janthina is endemic to islands of East Asia and is listed as Near Threatened by the International Union for Conservation of Nature (IUCN). One subspecies, C. janthina nitens, in particular, is at the greatest risk of extinction due to its small population size. To reduce the extinction risk of C. janthina, it is important to understand the species' present genetic status and to develop an appropriate conservation plan based on genetic data. We developed seven new microsatellite markers for two subspecies of C. janthina: C. janthina janthina and C. janthina nitens. We also confirmed the cross-use of one microsatellite marker developed for Columba livia var. domestica. Seven loci were polymorphic in C. janthina janthina, while two loci were polymorphic in C. janthina nitens. Using the markers, we performed preliminary analysis of genetic diversity and genetic structure within each subspecies. The expected heterozygosity ranged from 0.00 to 0.64 in C. janthina janthina and from 0.00 to 0.08 in C. janthina nitens. Each subspecies and each population within C. janthina janthina had different allele frequencies. C. janthina nitens exhibited far lower genetic diversity than C. janthina janthina. Furthermore, C. janthina nitens appears to have experienced strong genetic drift from a common ancestral population, inferred by STRUCTURE analysis. The markers described here may be useful for investigating genetic diversity and genetic structure of C. janthina populations, and could be used to estimate appropriate evolutionary significant unit and to guide development of a captive breeding program based on genetic information.