Diet Composition and Feeding Intensity of Four-Spotted Megrim, Lepidorhombus boscii (Risso, 1810), in the Eastern Adriatic Sea

Abstract

Feeding habits of the four-spotted megrim, Lepidorhombus boscii, from the eastern Adriatic Sea were examined. A total of 1070 individuals collected using a bottom trawl net between July 2020 and June 2021 were analyzed. A high percentage of empty stomachs (63.27%) was recorded. The food composition proved that L. boscii is a carnivorous species. The identified prey of L. boscii was divided into seven groups: Teleostei, Bivalvia, Gastropoda, Cephalopoda, Isopoda, Mysida, and Decapoda. Decapods were the most important prey (%IRI = 58.18), followed by cephalopods (%IRI = 10.93). At the species level, the most important prey were the decapods Parapeneus longirostris (%IRI = 11.48) and Goneplax rhomboides (%IRI = 5.92). Statistically significant seasonal variations in diet were recorded; decapods dominated in spring, summer, and winter, whereas cephalopods prevailed in autumn. There were no statistically significant differences in the food composition between the three size categories. Decapods were the most important prey in all size categories (%IRI > 50). The lowest feeding intensity, as well as the highest percentage of empty stomachs, was recorded in winter, which is also the main spawning season of this species in the Adriatic Sea.

1. Introduction

The four-spotted megrim, Lepidorhombus boscii (Risso, 1810), is a flatfish from the family Scophthalmidae, which inhabits the Mediterranean Sea and the eastern Atlantic from the British Isles to Angola [1]. In the Adriatic, it is distributed in the central and southern parts. It is a benthic species that inhabits soft sandy-muddy bottoms, at depths from 60 to 500 m, mainly between 200 and 300 m [2]. L. boscii has asynchronous ovarian development and a prolonged spawning season in the Adriatic Sea, with the main spawning season between November and March [3]. L. boscii is a commercially important species in the Mediterranean and the Adriatic Sea, and it is almost exclusively exploited with bottom trawl nets. In Croatia, L. boscii is classified under the category of “megrims”, along with other related flatfish species such as Lepidorhombus whiffiagonis, with annual catches amounting to approximately 5 tons [4]. Like most other commercially significant demersal species, L. boscii is primarily threatened by intensive bottom trawling and the degradation and/or loss of habitat caused by fishing activities [4]. Data on the biology and ecology of this species in the Adriatic are very scarce, which is similar to the Mediterranean. Previous studies on this species have primarily focused on age and growth [5,6,7,8,9,10], otolith morphology [5,10,11], biometry [12,13,14,15], reproductive biology [3,9,16,17,18], and diet [17,18,19,20,21].

However, except for the general information that the species feeds on fish, crustaceans, and cephalopods in the Adriatic [2,4], there are no available data or detailed studies on the diet of L. boscii in the Adriatic Sea.

Fish diet composition study is useful for developing trophic models as a tool for understanding the complexity of marine ecosystems, but also for determining the intensity of the interspecific interactions in marine fish communities [22]. This study aims to provide the first detailed information on the diet of L. boscii in the eastern Adriatic Sea, including insights into the qualitative and quantitative composition of food and food variations depending on season and fish size.

2. Materials and Methods

Monthly samples of L. boscii were collected in the eastern part of the Central Adriatic Sea between July 2020 and June 2021 (Figure 1). Fish sampling was performed by commercial bottom trawl nets with a mesh opening of 50 mm at the cod-end, at depths between 90 and 120 m. A total of 1070 specimens were analyzed: 324 sampled in spring, 256 in summer, 230 in autumn, and 260 in winter. The specimens were measured and dissected fresh, immediately after capture. For each sampled specimen, the total length (Lt) to the nearest 0.1 cm and body weight (W) to the nearest 0.01 g were noted. Sex was determined by examining the gonad morphology under a dissecting microscope. Only prey found in the stomach was analyzed, excluding materials from other parts of the gastrointestinal tract. Dissected stomachs were stored in a 4% formaldehyde solution. Prey was identified at the species level whenever possible. The number and wet weight (±0.001 g) of the prey specimens isolated from stomachs were recorded. In most of the stomachs analyzed, only one prey item was present, and mainly in good condition. In the case of multiple damaged prey items, individual fragments were analyzed separately to determine the taxonomic group as accurately as possible. Crustaceans were identified by exoskeleton fragments, appendages, and eyes. Fish were recognized by scales and otoliths, and cephalopods were determined using beaks and statoliths. The size of the analyzed fish ranged from 12.5 to 34.0 cm Lt (Figure 2). The data were first pooled together to provide a comprehensive insight into the dietary composition of the species based on the entire data set. For the purposes of a more detailed analysis, the samples were subsequently sorted by size category of individuals and by season. To evaluate variation in food composition as a function of size, fish specimens were separated into three size categories: 12.5–19 cm (n = 374), 19–24 cm (n = 541), and 24–34 cm (n = 155).

To evaluate the rate of feeding activity, the vacuity index %V = (Ne/Ns) × 100 [12] was calculated, where Ne is the number of empty stomachs and Ns is the total number of examined stomachs. A Pearson’s Chi-squared test was applied to test differences in the number of empty stomachs by season and fish size. The contribution of each prey taxon to the fish diet was evaluated by calculating the following: percentage frequency of occurrence (%F), as the percentage of stomachs containing a particular prey taxon; percentage of numerical abundance (%N), as the number of each prey taxon expressed as a percentage of the total number of identified taxa; percentage gravimetric composition (%W), the weight of each kind of prey, expressed as a percentage of the total weight of the prey ingested [23,24,25]. These values were used to calculate the index of relative importance, IRI = (%N + %W) × %F [26]. The index was expressed as a percentage: $\%\mathrm{I}\mathrm{R}\mathrm{I}=\left({\displaystyle \frac{\mathrm{I}\mathrm{R}\mathrm{I}}{\sum \mathrm{I}\mathrm{R}\mathrm{I}}}\right)\times 100$.

Diet composition was analyzed by aggregating stomach content remains into seven taxonomic groups: Cephalopoda, Decapoda, Bivalvia, Teleostei, Mysida, Isopoda, and Gastropoda. Remains that could not be identified at this taxonomic level were removed from the analyses, leaving a final dataset with 318 individuals of fish. The Sørensen–Dice similarity index was calculated between all samples, and data were visualized using non-metric multidimensional scaling (nMDS) in two-dimensional space. ANOSIM was used to perform a robust non-parametric permutation test for significant differences among groups, seasons, and length classes, with 9999 permutations [27,28]. The Benjamini–Hochberg procedure was used to control the false discovery rate at 0.05, after pairwise comparisons [29]. To better observe differences between seasons, group means were visualized using metric MDS, with a measure of uncertainty of their position obtained as 95% bootstrap regions. The averages of repeated bootstrap samples (n = 100) for each group were ordinated into two dimensions to form a region estimate for the means, and then smoothed and marginally bias-corrected [30]. This is implemented in PRIMER7 software as a bootstrap averages routine. Multidimensional scaling and ANOSIM were also performed in PRIMER 7 [31], and the Benjamini–Hochberg correction was performed in R v4.4.1 (https://www.r-project.org/, accessed on 3 February 2025) using the function p.adjust().

3. Results

3.1. Feeding Intensity

Out of 1070 L. boscii stomachs analyzed, 677 were empty (63.27%). This frequency of empty stomachs varied significantly during the year (χ² = 14.19, df = 3, p = 0.002), with a maximum of 73.07% in winter, followed by summer (61.32%), spring (59.87%), and a minimum in autumn (59.13%). Also, statistically significant differences in empty stomachs between different fish sizes were recorded (χ² = 9.74, df = 2, p = 0.007). In smaller fish, 12.5–19 cm, the percentage of empty stomachs was the highest (72.19%); it was lower in the size class 19–24 cm (61.92%), and the lowest was in the largest fishes, i.e., in the size class 24–34 cm (46.45%).

3.2. Diet Composition

The food composition, percentage frequency of occurrence, percentage numerical abundance, percentage gravimetric composition, and %IRI values of prey organisms found in the non-empty stomachs of all analyzed fish are shown in

Table 1. Diet composition from 393 non-empty stomachs of L. boscii (%F—percentage frequency of occurrence, %N—percentage numerical abundance, %W—percentage gravimetric composition, IRI—index of relative importance); “non-identified prey” includes digested remains.

Food Items	%F	%N	%W	IRI	%IRI
PISCES TELEOSTEI
Argentina sphyraena	0.25	0.24	0.11	0.08	<0.1
Callionymus fasciatus	0.25	0.24	0.09	0.08	<0.1
Cepola rubescens	0.50	0.48	1.06	0.77	<0.1
Lesueurigobius friessi	0.25	0.24	0.13	0.09	<0.1
Merluccius merluccius	9.92	9.48	10.08	194.03	2.16
Non-identified Teleostei	3.30	3.16	2.18	17.62	0.19
Total Teleostei	14.47	13.84	13.65	397.78	4.44
CRUSTACEA DECAPODA
Alpheus glaber	0.25	0.24	0.27	0.12	<0.1
Eriphia spinifrons	0.25	0.24	0.03	0.06	<0.1
Goneplax rhomboides	18.06	18.00	11.38	530.60	5.92
Munida intermedia	5.34	5.10	2.78	42.07	0.46
Munida nexa	1.52	1.49	0.64	3.23	<0.1
Iridonida speciosa	2.03	1.94	1.35	6.67	<0.1
Munida spp.	1.01	1.21	0.55	1.77	<0.1
Parapenaeus longirostris	23.40	22.62	21.34	1028.66	11.48
Processa canaliculata	0.50	0.48	0.33	1.02	<0.1
Solenocera membranacea	1.27	1.21	2.19	4.31	<0.1
Upogebia pulsilla	0.25	0.24	0.20	0.11	<0.1
Non-identified Crustacea	0.76	0.72	0.79	1.14	<0.1
Total Decapoda	54.64	53.49	41.85	5209.37	58.18
MYSIDA
Lophogaster typicus	2.29	2.43	1.88	9.86	0.11
Total Mysida	2.29	2.43	1.88	9.86	0.11
ISOPODA
Anuropodione amphiandra	7.12	7.05	9.77	119.75	1.33
Total Isopoda	7.12	7.05	9.77	119.75	1.33
MOLLUSCA CEPHALOPODA
Alloteuthis media	11.45	10.94	18.18	333.42	3.72
Illex coindetii	1.78	1.70	2.78	7.87	<0.1
Loligo vulgaris	1.01	0.97	2.78	3.78	<0.1
Sepia officinalis	4.58	4.37	2.86	33.11	0.36
Sepiola rondeletii	0.25	0.24	0.21	0.11	<0.1
Non-identified Cephalopoda	1.78	1.70	0.76	4.37	<0.1
Total Cephalopoda	20.85	19.98	27.57	978.57	10.93
GASTROPODA
Non-identified Gastropoda	0.76	0.72	0.36	0.82	<0.1
Total Gastropoda	0.76	0.72	0.36	0.82	<0.1
BIVALVIA
Nucula spp.	0.25	0.24	0.12	0.09	<0.1
Nuculana pella	0.25	0.24	0.06	0.07	<0.1
Non-identified Bivalvia	0.50	0.48	0.18	0.33	<0.1
Total Bivalvia	3.81	3.64	1.95	21.29	0.23

. Food items identified in fish stomachs belonged to seven different prey groups: Teleostei, Bivalvia, Gastropoda, Cephalopoda, Isopoda, Mysida, and Decapoda (Table 1). Decapod crustaceans occurred in 54.64% of stomachs that contained food and represented 53.49% of the total prey number, and 41.85% of the total weight. Cephalopods occurred in 20.85% of stomachs examined and represented 19.98% by number and 27.57% by weight of the total prey. Decapod crustaceans were the most important prey group, constituting 58.18% of the total IRI, and were followed by cephalopods (%IRI = 10.93). Other prey groups had much lower %IRI values and were thus of less importance. Due to the advanced stage of digestion, species-level identification was often not possible. The most common identifiable prey items were decapods Parapeneus longirostris (%IRI = 11.48) and Goneplax rhomboides (%IRI = 5.92) (Table 1). The prey found in fish stomachs indicates a relatively diverse diet of L. boscii in the eastern Adriatic.

3.3. Seasonal Variation in the Diet Composition

The food composition of L. boscii varied between seasons. Decapods were the dominant group in spring (%IRI = 91.69), summer (%IRI = 89.98), and winter (%IRI = 53.32). Other identified crustaceans, i.e., isopods and mysids, were found in most of the seasons; however, except for isopods in spring (%IRI = 3.46), these groups were much less represented (%IRI < 1). Cephalopods were the most important prey during autumn (%IRI = 72.70), and they were also important in the diet of L. boscii during winter (%IRI = 22.23%). Teleosts were present in the diet in all seasons, with the highest value during winter (%IRI = 23.70). Other prey taxa had very low %IRI values and were found only in one season each: bivalves in summer, and gastropods in spring (

Table 2. Index of relative importance IRI (%IRI) of L. boscii prey by season.

	IRI (%IRI)
Group of Prey	Summer	Autumn	Winter	Spring
Teleostei	156.25 (3.34)	277.39 (4.70)	642.72 (23.70)	201.82 (3.27)
Bivalvia	3.69 (0.07)	-	-	-
Gastropoda	-	-	-	0.56 (0.009)
Cephalopoda	133.13 (2.85)	4283.76 (72.70)	603.05 (22.23)	95.53 (1.55)
Isopoda	33.28 (0.71)	-	17.85 (0.65)	213.69 (3.46)
Mysida	8.32 (0.17)	13.71 (0.23)	1.98 (0.07)	-
Decapoda	4201.18 (89.98)	1316.87 (22.35)	1446.14 (53.32)	5650.97 (91.69)

).

There was a statistically significant difference between diets in different seasons (ANOSIM R = 0.08, p = 0.0001). Diet in warm seasons differed from cold ones (

Table 3. Results of the ANOSIM test comparing the diet of Lepidorhombus boscii by season.

Pairwise Tests
Season	R	p
Spring–Summer	0.004	0.231
Spring–Winter	0.122	0.001
Spring–Autumn	0.149	0.0003
Summer–Winter	0.089	0.0015
Summer–Autumn	0.11	0.0003
Winter–Autumn	0.019	0.0852

, Figure 3 and Figure 4).

3.4. Food in Relation to Fish Size

Decapods were the most dominant prey in all size categories (12.5–19 cm, 19–24 cm, and 24–34 cm), and %IRI increased with the size of the fish. On the other hand, cephalopods and teleosts were the second and third most important prey groups in the 19 to 24 cm size category, but their %IRI decreased in the largest size class. Isopods were often found in the 24–34 cm size category (%IRI = 9.73). Other prey taxa (Mysida, Bivalvia, Gastropoda) were either absent or present but with very low indices of relative importance (%IRI < 0.5) in all size groups (

Table 4. Composition of L. boscii diet among fish size, based on %IRI values of major prey groups.

IRI (%IRI)
Group of Prey	12.5–19 cm	19–24 cm	24–34 cm
Teleostei	439.67 (14.87)	948.34 (5.09)	316.75 (3.31)
Bivalvia	1.97 (0.06)	-	1.99 (0.02)
Gastropoda	-	-	2.22 (0.02)
Cephalopoda	984.8 (33.31)	2524.82 (13.55)	63.05 (0.65)
Isopoda	-	224.51 (1.20)	930.03 (9.73)
Mysida	14.57 (0.49)	2.73 (0.01)	1.99 (0.20)
Decapoda	1515.16 (51.25)	14,920.41 (80.12)	8241.83 (86.23)

). There was no statistically significant difference between diets of different size classes (ANOSIM R = 0.002, p = 0.42).

4. Discussion

The composition of food proved that the four-spotted megrim inhabiting the eastern part of the central Adriatic Sea is a carnivorous species and its diet is based on bottom-living crustaceans, molluscs, and teleosts. Decapod crustaceans were the most abundant prey, constituting 58.18% of the total %IRI. The prey group accounting for more than 50% of the total %IRI can be considered as the main food source [25]. Cephalopods were the second most important prey (%IRI = 10.93), while other prey groups were of less importance. Generally, the prey identified in stomachs indicates that the four-spotted megrim is an opportunistic fish species that feeds on diverse types of available benthic organisms. The stomach contents of the four-spotted megrim reflected the distribution and abundance of several benthic species that inhabit soft bottom sediments, such as decapods P. longirostris and G. rhombides, cephalopods A. media and S. officinalis, teleosts M. merluccius, and L. friessi, which are all common on sandy-muddy bottoms in the Adriatic Sea [4].

Most of the species found in the stomach contents live on the bottom surface or buried in the substrate, so active capturing of prey includes both hunting on the seabed and digging in the bottom in search of food [20]. The four-spotted megrim, like other scopthalamids, has a large mouth, gill rakers, and a stomach with a simple intestinal loop [21], and is well adapted to feed on larger, quick-moving prey [32]. As in the present study, decapods were the most important food of the four-spotted megrim sampled in Portuguese waters [17]. Also, decapods followed by teleosts were the dominant prey of these species in the Aegean Sea [21], Spanish waters [20], and Tyrrhenian Sea [19]. The results of these studies confirm the importance of decapods in the four-spotted megrim diet, and observed variations in prey composition are likely linked to regional differences in the presence and availability of various benthic assemblages [33].

In our study, stomach content analysis did not reveal distinct differences in the prey selection based on fish size, as decapods dominated the diet across all size categories (%IRI > 50%). Decapods, cephalopods, teleosts, and a very small percentage of bivalves and mysids are represented in the stomachs of smaller individuals. With the increase in size of the fish, there is an increase in the representation of decapods and cephalopods, as well as the appearance of parasitic isopods. The parasitic isopod Anuropodione amphiandra was found in the stomachs of L. boscii together with decapod crustaceans from the family Munididae and the species Parapeneus longirostris. This parasitic species has been found with high incidence in the host I. speciosa in the Adriatic Sea [34] and also in some other species of the family Munididae, such as Munida rutllanti in the eastern Mediterranean [35]. Based on that, we can conclude that isopods, i.e., specimens of A. amphiandra, found in the stomachs of the four-spotted megrim were endoparasites of their decapod prey. The presence of the host-parasite complex in the stomach of L. boscii indicates the transmission of parasites through trophic pathways and increases the understanding of food web dynamics and parasitism in benthic communities [36]. The representation of teleosts in the stomachs decreased with an increase in the total body length of L. boscii. Greater differences in feeding between fish size classes of L. boscii were recorded from the Tyrrhenian Sea [19], Aegean Sea [21], Portuguese [17], and Spanish waters [20]. In these studies, decapods were the most abundant prey in the stomachs of small individuals, and teleosts were represented exclusively in larger individuals (>18 cm) [17,21]. Mysids dominated in smaller size classes, while decapods dominated medium and large size classes [19,20]. Numerous authors conclude that an increase in fish size is associated with an increase in the consumption of larger prey, which is in line with the theory of optimal foraging [37], according to which larger predators consume larger prey to maximize energy gain relative to the effort of catching prey [37,38]. Based on our results, we can conclude that decapods in the Adriatic are the most important source of food for L. boscii, regardless of the size of the individual. However, in our study, teleosts were found only in smaller individuals, which is different from other studies and may indicate specific feeding habits of this species in the Adriatic, but considering that L. boscii is an opportunistic predator, this may also indicate differences in the food availability in different geographical areas [39]. Similar conclusions were drawn by Bișinicu et al. [40], who emphasized that spatial variability in prey resources plays a key role in shaping fish feeding strategies and distribution patterns in marine environments.

Statistical analyses have shown that the composition of the diet of L. boscii from the Adriatic Sea depends on the seasons, i.e., that there are differences between the warmer and colder parts of the year. Decapoda were most abundant in the stomachs in spring, summer, and winter, while cephalopods were more abundant during autumn. Seasonal dietary variation likely reflects prey availability, with prey distribution and abundance being influenced by the dynamics of water masses [41]. Increased consumption of decapods during the summer coincides with the period of new recruits of many decapod species that can be present in high densities [42]. These findings are consistent with broader evidence showing that trophic interactions and food web structures in marine ecosystems are closely linked to temporal and spatial variations in lower trophic levels [43]. Results for Portuguese waters and the Aegean Sea indicate that decapods are the main prey during all seasons [17,21], which suggests the importance of decapods in the seasonal diet of this species. On the other hand, statistically significant differences were not recorded in feeding specimens of different length categories. This suggests that specimens of all sizes have similar food preferences and feed on the same resources in the habitat.

In this study, a very high percentage of empty stomachs was recorded (63.27%), and the percentage of empty stomachs is negatively related to the intensity of feeding [44]. The share of empty stomachs was especially high during winter. Low feeding intensity during winter is partly associated with lower sea temperatures [45], with many demersal fishes showing a decrease in their feeding rates in the cold season [46]. It can also be related to the seasonality of prey abundance and species composition [47]. Moreover, the spawning of the four-spotted megrim is the most intensive in autumn and winter [3], so the low feeding intensity may be a result of the pre-spawning and spawning conditions when fish usually decrease food intake or even cease feeding [48]. The feeding of most fish species oscillates considerably during the year as a consequence of physiological changes during reproduction [49].

5. Conclusions

L. boscii from the eastern Adriatic Sea is a carnivorous fish that feeds on bottom-living crustaceans, molluscs, and teleosts. Decapods were the most important prey in all size categories (%IRI > 50%). Significant seasonal variations in diet were recorded, with significant differences in food composition between warm (spring–summer) and cold (autumn–winter) seasons. Decapods dominated in spring, summer, and winter, whereas cephalopods were the most important prey in autumn. Low feeding intensity during the winter could be associated with the peak of spawning and lower sea temperatures in the season. The presented results improve the scarce knowledge on the feeding ecology of this species in the Adriatic Sea and provide basic information for the development of a multi-species assessment model for this area.