Trapping times and body size comparisons
Trapping times ranged from 2.5–13.5 h with a mean of 5.7 h; processing times ranged from 0.5–5.5 h with a mean of 1.9 h. Total trapping plus processing times (time from trap placed in water to specimen placed into formalin) ranged from 3.5–14.5 h with a mean of 7.6 h. Trapping times in excess of 10 h occurred overnight during summer in the lake and stream. Overall prey consumption and trapping hours were not significantly related (F (9, 244) = 1.779, p = 0.073; Table 1), indicating a representative sampling.
Table 1 Multivariate effects on diet of blackfish. The MANCOVA is based on three factors (waterbody, season, and sex), two covariates (SL, trapping hours), and nine response variables (nine prey categories whose % IRI ≥ 1). SL = standard length, ns = not significant
From a total of 470 blackfish collected and fixed in formalin, 302 fish were dissected for stomach content analysis: 84 from the lake, 104 from the stream, and 114 from the wetland (Suppl. Table 1). In spring, only wetland fish were captured and analyzed due to unsafe ice conditions at the other sites. Seventy-eight percent of dissected blackfish were trapped during daytime compared to 22 % trapped overnight, in summer, from the lake and stream. Among specimens examined, males (SL mean = 98.90 mm, SD = 15.51, n = 145) were larger (t (297) = 7.23, p < 0.001) than females (SL mean = 91.67 mm, SD = 15.44, n = 154), though sizes broadly overlapped (Suppl. Fig. 1). Seventy-five percent of all fish were in 75–115 mm SL size range. The three sites varied significantly (ANOVA, F = 7.14, p < 0.001) in SL (lake mean SL = 90.96 mm, stream mean SL = 97.61 mm, wetland mean SL = 94.61 mm). Blackfish did not vary significantly in SL by season.
Prey categories and dietary diversity
Blackfish gut contents from the three study sites contained prey from 20 taxonomic groups distributed among five animal phyla (mollusks, annelids, arthropods, bryozoans, and chordates) and one plant division (Suppl. Table 2). In the stomach content analysis, insects were classified as Insecta adults (Order unknown), Diptera larvae, Trichoptera larvae, Odonata larvae, and Ephemeroptera larvae. Order Coleoptera included both larvae and adults. Bryozoa/Plumatellida from DeLong Lake consisted of over-wintering cysts (statoblasts). Angiosperms in blackfish gut contents were represented by seeds of unknown plants. Occasional plant stem and leaf tissue found in the guts of blackfish were excluded from analysis because of their rarity.
Consistent with the prediction of high diet diversity, blackfish in these populations typically had many prey types in their guts. The number of different prey types, based on 20 broad prey categories (Suppl. Table 2) found in each nonempty gut (esophagus + stomach + intestines) ranged from 1 to 10 (Fig. 3). The mode for number of prey categories per examined gut was three in the lake and stream samples, and four in the wetland samples. Waterbody and season were significant predictors of the nine response variables (nine prey categories whose % IRI ≥ 1), as was the interaction between them (Table 1). Blackfish size and sex were not significant predictors of prey types across all nine prey categories, although the frequency of fish in diet was related to blackfish size and sex (Chi-square = 9.487, df = 2, p = 0.009).
Stomach contents by combined sites and seasons
Figure 4 provides a graphic representation of the IRI from all samples combined for prey categories with ≥3 % IRI. For combined waterbodies and seasons, gastropods were by far the dominant prey group (51 % IRI). Ostracods contributed 28 % of the percent IRI, while all other prey taxa recorded 7 % IRI or less. Twelve other categories made minor contributions to blackfish diets at these sites (%IRI < 0.5 %; Table 2). Ostracods were the primary prey in terms of total number, while gastropods and teleosts were the two most important prey in terms of biomass. Fish as prey ranked fourth in importance by percent IRI, while frequency of fish consumption was less than 10 %. Dipterans were found most frequently in guts (62 % frequency), followed by gastropods (46 % frequency). Overall, six prey types were consumed by at least 25 % of all blackfish: dipteran larvae, gastropods, copepods, trichopteran larvae, ostracods, and bivalves. Plant seeds were found in 14 % of all guts.
Table 2 Prey values for combined sites and seasons (n = 302). Major and minor prey categories are given in descending order by percent IRI. Grey-colored rows are prey categories shown in IRI diagram (Fig. 4). Diptera, Odonata, Trichoptera, and Ephemeroptera represent larvae
Stomach contents by site
Prey consumption differed by waterbody for seasons pooled (F (18, 490) = 4.56, p < 0.001; Table 1). A different prey taxon dominated gut contents for each site: dipterans (66 % IRI) in lake fish, gastropods (78 % IRI) in stream fish, and ostracods (59 % IRI) in wetland fish (Suppl. Fig. 2, Suppl. Tables 3, 4, 5). All other prey values at each site were less than 20 % IRI. Teleosts contributed at least 60 % of the percent biomass in gut contents from lake fish, while gastropods contributed 70 % of overall biomass in gut contents for stream fish. No single taxon dominated by percent biomass for wetland fish. Dipteran larvae were the most frequently consumed prey of lake fish, compared to gastropods and dipteran larvae for stream fish. Wetland blackfish had the greatest variety of prey in their guts, consuming three prey categories at 50 % or greater frequency and three additional taxa at frequencies of 24 %–35 %. Fish ranked third in prey importance for both lake and stream blackfish but were of minor importance for wetland blackfish.
Stomach contents by season
Prey consumption differed by season (spring excluded) for waterbodies pooled (F (18, 490) = 3.56, p < 0.001; Table 1). During summer, gastropods were the single major prey (69 % IRI), while all other groups contributed less than 10 % IRI (Suppl. Tables 6). Gastropods also dominated in summer by biomass (79 % mass), followed by teleosts (10 % mass). More than half of all guts in summer contained gastropods and dipterans. Small ostracods and cladocerans outnumbered larger prey.
Dominant prey groups in autumn consisted of gastropods and ostracods, which contributed 48 % and 24 % of the percent IRI values, respectively (Suppl. Table 7). Gastropods were the most important prey in terms of biomass (61 % mass). At least 45 % of all guts in autumn contained dipterans, gastropods, trichopterans, and ostracods.
Prey in winter switched to Ostracoda (39 % IRI) followed by Diptera (24 % IRI) and Teleostei (19 % IRI; Suppl. Table 8). Fish were the most dominant winter prey by biomass (72 % mass), although their total count (13) was small compared to ostracods (4452). The most frequently consumed winter prey were dipteran larvae and copepods.
We chose to interpret a significant (randomization test p = 0.004) two-dimensional NMS solution with mean stress of 12 after verifying consistency of interpretation among the three NMS solutions. Blackfish with a greater mass of gastropods in their gut are associated with greater positive Axis 1 values; blackfish with a greater mass of teleosts in their gut are associated with greater positive Axis 2 values; blackfish with a greater mass of dipterans in their gut are associated with lower Axis 2 values (Fig. 5). Blackfish with more mixed diets and low mass of the six arthropod categories (including ostracods, copepods, dipterans, and trichopterans) are located in the center of the biplots, while blackfish with an empty gut are represented by the lowest Axis 1 value.
The biplots of waterbodies across the three seasons (summer, autumn, winter) show overlap in diet for many blackfish (Fig. 5). However, lake blackfish are concentrated on the upper-left side (greater mass of teleosts, and mixed arthropods); stream blackfish extend to the right side (greater mass of gastropods and teleosts, lower mixed arthropods), and wetland blackfish occupy the center portion of the biplots (greater mass of mixed arthropods, low mass of gastropods and teleosts). In general, diets varied considerably within each waterbody, though wetland blackfish had a less variable diet, especially in summer. Little difference was evident among the summer and fall biplots. In winter, however, diets for the stream and wetland blackfish constricted along Axis 1 and expanded along Axis 2. This is associated with a reduction in gastropods and increase in teleosts and copepods in their diets. Diets among fish in the three waterbodies converged more in winter. Lake blackfish showed little change in diet composition across seasons in the ordination.
Fish in diet
Gut contents from pooled waterbodies and seasons showed that 9 % of all blackfish had recently consumed a total of 35 fish (Table 2, Suppl. Table 9). Seven percent of the lake blackfish had fish in their guts—four threespine stickleback, one juvenile blackfish, and two unidentifiable fish (Suppl. Table 3). Stream blackfish were the most piscivorous (17.3 % frequency; Suppl. Table 4); fish prey included 10 threespine stickleback, four ninespine stickleback, one coho, and three unidentified fish. Piscivory among blackfish in the wetland was lowest at 3.5 % frequency across the year (2.2 % when spring samples are excluded; Suppl. Table 5), with threespine stickleback, juvenile blackfish, and one unidentifiable fish. Overall, the frequency of cannibalism for all waterbodies and seasons was 1.3 %, excluding unidentifiable prey fish. Less than 1 % of blackfish had salmonids in their guts.
The likelihood of fish in blackfish diet was related to both sex and blackfish size (Chi-square = 9.487, df = 2, p = 0.009). The Wald statistic showed that both sex (p = 0.043) and size (p = 0.045) made significant contributions to the model. Fish consumption first appeared in blackfish with a SL of at least 78 mm, although few blackfish this small had fish in their guts (Suppl. Table 9). Fish were more important in the diets of blackfish greater than 105 mm SL. Males tended to consume more fish than did females. Approximately 13 % of males consumed fish, while 6 % of females consumed fish.