DNA metabarcoding reveals diet diversity and niche partitioning by two sympatric herbivores in summer

Abstract

Background: Food provides essential nutrients and energy necessary for animals to sustain life activities. Accordingly, dietary niche analysis facilitates the exploration of foraging strategies and interspecific relationships among wildlife. The vegetation succession has reduced understory forage resources (i.e., shrubs and herbs) available to sika deer (Cervus nippon kopschi). Little is known about the summer foraging strategies or the interspecific relationship between sika deer and Reeves' muntjac (Muntiacus reevesi).

Methods: The present study used high-throughput sequencing and DNA metabarcoding techniques to investigate the feeding habits and interspecific relationships between sika deer and Reeves' muntjac in our study.

Results: A total of 458 amplicon sequence variants (ASVs) were identified from fecal samples, with 88 ASVs (~19.21%) unique to sika deer and 52 ASVs (~11.35%) unique to Reeves' muntjac, suggesting the consumption and utilization of specific food items for the two species. The family Rosaceae was the most abundant for both species, especially Rubus spp. and Smilax china. Alpha diversity (local species richness) indicated that the dietary species richness of sika deer was higher than that of Reeves' muntjac, but the difference was not statistically significant. Sika deer also exhibited a higher evenness index (J' = 0.514) than Reeves' muntjac (J' = 0.442). Linear discriminant effect size analysis revealed significant differences in forage plants between the two herbivores. The niche breadths of sika deer and Reeves' muntjac were 11.36 and 14.06, respectively, and the dietary niche overlap index was 0.44. Our findings indicate the diet partitioning primarily manifested in the differentiation of food items and the proportion, which ultimately reduces the overlap of nutritional niches and helps avoid conflicts resulting from resource utilization. This study provides a deeper insight into the diversity of foraging strategies and the interspecific relationship of herbivores from the food dimension.

Keywords: DNA metabarcoding; Dietary partitioning; Herbivores; Niche overlap; Summer.

Figure 1. Ecological photographs from the reserve.

(A) Suitable scrub-grassland habitat of sika deer; (B) reduced understory food resources available to sika deer. (C) Vegetation dwarfing experimental plot; (D) comparison between the dwarfed experimental plots and the non-dwarfed areas.

Figure 2. Sampling sites at the Taohongling Sika Deer National Nature Reserve.

(MP: Nursery bases; XLA: XianLingAn; SS: Fir forests; WGS: WuGuiShi; NJS: NieJiashan; ZY: Bamboo garden). Built map with ArcGIS Pro V3.0.0: https://www.esri.com/zh-cn/arcgis/products/arcgis-pro/overview.

Figure 3. (A) Box-plot of the alpha diversity index using Kruskal-Wallis and Wilcox tests. In each panel, the abscissa is the group, and the ordinate is the value of the corresponding alpha diversity index. (B) Alpha rarefaction curves: Observed species index and Shannon index.

*Significant different between groups (p < 0.05).

Figure 4. The bar chart distribution of the dominant forage plant in sika deer and Reeves’ muntjac groups at the genus level.

The x-axis stands for individual samples.

Figure 5. Top 20 forage plants with the highest proportions in sika deer and Reeves’ muntjac groups at the species level.

The x-axis stands for individual samples.

Figure 6. (A) Cladogram based on LEfSe analysis, showing ASVs with the significance of two herbivores (green: sika deer; dark green: Reeves’ muntjac). (B) Log10-transformed LDA scores for ASVs, i.e., with a threshold value >4.0.

Figure 7. (A) NMDS analysis of sika deer and Reeves’ muntjac with 95% confidence ellipse. (B) Network analysis of forage plants.

The size of the circles represents relative abundance, the lines indicate a significant correlation between two species (p < 0.05). Red lines mean positive correlations and blue means negative correlations.