Journal of Sea Research 72 (2012) 14–21 Contents lists available at SciVerse ScienceDirect Journal of Sea Research journal homepage: www.elsevier.com/locate/seares Food sources of macro-invertebrates in an important mangrove ecosystem of Vietnam determined by dual stable isotope signatures Nguyen Tai Tue a, b,⁎, Hideki Hamaoka b, Atsushi Sogabe c, Tran Dang Quy d, Mai Trong Nhuan d, Koji Omori b a Graduate School of Science and Engineering, Ehime University, 2‐5 Bunkyo-cho, Matsuyama, Japan Center for Marine Environmental Studies, Ehime University, 2‐5 Bunkyo-cho, Matsuyama, Japan Graduate School of Biosphere Science, Hiroshima University, 1-4-4 Kagamiyama Higashi-Hiroshima 739‐8528, Japan d Faculty of Geology, Hanoi University of Science, 334 Nguyen Trai, Thanh Xuan, Hanoi, Viet Nam b c a r t i c l e i n f o Article history: Received 12 February 2012 Received in revised form 24 April 2012 Accepted May 2012 Available online 16 May 2012 Keywords: Stable Isotopes Invertebrate Food Sources Mangrove Ecosystem Vietnam a b s t r a c t Dual stable isotope signatures (δ13C and δ15N) were applied to determine the contribution of mangrove materials and other organic carbon sources to the invertebrate community in an ecologically important mangrove ecosystem of Vietnam We have analyzed 181 specimens of 30 invertebrate species and found δ13C and δ15N ranging from − 14.5 to − 26.8‰ and from 1.3 to 12.1‰, respectively From taxa measured for stable isotopes, polychaete, gastropods, bivalves, and grapsid crabs living in mangrove forest showed relative low δ13C values, while fiddler crabs inhabiting in the land–water ecotone showed the highest δ13C values The δ13C showed that just a few mangrove inhabitants directly relied on the mangrove materials The wide ranges of δ13C and δ15N signatures indicated that the invertebrates utilized heterogeneous diets, comprising benthic microalgae, marine phytoplankton, particulate organic matter, sediment organic matter, mangrove detritus, and meiofauna and rotten animal tissues as the supplemental nutrient food sources Moreover, the significant correlation between δ13C values and body sizes of invertebrates showed that snails Littoraria melanostoma and Terebralia sulcata, bivalve Glauconome virens, and portunid crab Scylla serrata exhibited ontogenetic shifts in diets The present study showed that adjacent habitats such as tidal flat and mangrove creeks seem to contribute an important microalgal food resource for invertebrates and highlighted the need for conservations of mangrove forests and the adjacent habitats © 2012 Elsevier B.V All rights reserved Introduction Mangrove forests have often been regarded as one of the most productive ecosystems in the (sub)tropical coastal waters They are characterized by the high biodiversity of invertebrates and fish (Nagelkerken et al., 2008; Sasekumar et al., 1992) Several hypotheses have been proposed for the explanation of the high biodiversity of the invertebrates and fish in the mangrove ecosystem, including (1) mangroves characterize by the structural complexity of pneumatophores and/or prop roots which provide shelter from predators for invertebrates and fish (Kon et al., 2009), and (2) mangroves produce large amounts of organic matter that form the basis of the estuarine food webs (Hogarth, 2007; Nagelkerken et al., 2008; Odum and Heald, 1972) Therein, the later hypothesis has long been debated between non-stable isotopic (i.e., stomach content analysis, fecal analysis, and direct observation (Michener and Lajtha, 2007)) and isotopic ⁎ Corresponding author at: 790‐8577 Center for Marine Environmental Studies, Ehime University, 2‐5 Bunkyo-cho, Matsuyama, Japan Tel.: + 81 89 927 9643, + 81 902 894 1610 (Cell); fax: + 81 89 927 9643 E-mail address: tuenguyentai@gmail.com (N.T Tue) 1385-1101/$ – see front matter © 2012 Elsevier B.V All rights reserved doi:10.1016/j.seares.2012.05.006 studies Based on stomach content analysis several studies have demonstrated that the mangrove detritus contributes a significant amount of organic carbon fueling detrital-based food webs (Nanjo et al., 2008; Nordhaus et al., 2011; Odum and Heald, 1972) Nevertheless, the isotopic studies have failed to confirm the ingestion of mangrove materials in the estuarine food webs (France, 1998; Rodelli et al., 1984), apparently because the mangrove detritus is too refractory and the simple ingestion of mangrove detritus does not indicate any direct assimilation of that material (Fry and Ewel, 2003) Invertebrates that play important roles in the mangrove structure processes (Cannicci et al., 2008), organic carbon dynamics (Robertson and Daniel, 1989), biogeochemical processes (Kristensen, 2000), are preyed upon foraging fishes during high tides (Kruitwagen et al., 2010), and serve as important links between mangrove detritus and estuarine secondary production (Lee, 2008) The gastropods and brachyuran crabs are the most dominant invertebrate groups in the mangrove ecosystem They play an important role in leaf litter turnover, for example, sesarmid crabs can consume approximately 70% of the total annual litter fall from the forest floor (Robertson and Daniel, 1989), significantly retaining mangrove organic production and reducing direct export Moreover, recent isotopic studies pointed out that several invertebrate species rely on the mangrove materials and such use of mangrove carbon N.T Tue et al / Journal of Sea Research 72 (2012) 14–21 depends on the morphological characteristics of mangrove forests (Bouillon et al., 2004) and/or microhabitats (Kon et al., 2007) Stable isotopes have been used to track organic matter flows and to measure the food web structures, and ontogenetic shifts in diets (Michener and Lajtha, 2007) Stable isotopes of carbon (δ 13C) and nitrogen (δ15N) have frequently applied for tracing the flow of organic matters in lake (Post, 2002), estuarine (Peterson et al., 1985), and marine (Michener and Lajtha, 2007) food webs Therein, δ13C can be used to evaluate the ultimate sources of carbon for an organism when the isotopic signatures of the sources are different (Post, 2002) The δ13C is enriched approximately 0.5–1‰ in the animal relative to its diet The basic assumption for estimation of trophic position is based on a conservative enrichment of 3–4‰ δ15N in a consumer relative to its diet (Michener and Lajtha, 2007) Stable isotope signatures (δ13C and δ15N) are therefore powerful methods for determining the ingested food sources and the food assimilation over long period of a consumer Stable isotope measurements have been applied to trace mangrove detritus in the estuarine environment (Dittmar et al., 2001), to estimate the assimilation of mangrove detritus by a specific species such as snail (Penha-Lopes et al., 2009; Slim et al., 1997) and fiddler crabs (France, 1998) Yet, stable isotope analysis has been rarely applied to identify food sources of a large range of invertebrate taxa in the mangrove ecosystem (i.e., Bouillon et al., 2002a, 2004; Kon et al., 2007; Kruitwagen et al., 2010), particularly in developing countries such as Vietnam In the present study, we have analyzed dual stable isotope signatures (δ 13C and δ 15N) of 181 specimens of 30 invertebrate species for figuring out the question what degree of mangrove materials is assimilated by invertebrates in an ecologically important mangrove ecosystem of Vietnam The objectives of the present study are (1) to determine the utilization of food sources by invertebrates and (2) to examine the question of whether these invertebrates exhibit the ontogenetic shifts in diets Materials and methods 2.1 Study area The present study was conducted in an estuarine mangrove ecosystem of Xuan Thuy National Park (XTP) in northern Vietnam (Fig 1) The detailed description for the XTP has been shown elsewhere (Tue et al., 2012, 2012) Briefly, XTP covers a total wetland area of 12,000 ha, of which about 3000 are covered by mangrove forests The most abundant mangrove species are Sonneratia caseolaris, Kandelia obovata, 15 Aegiceras corniculatum, and Avicennia marina The mangrove ecosystem provides a broad array of ecosystem services such as maintaining high biodiversity, storm protection, erosion mitigation, local climate regulation, fishery production, and the subsistence livelihood The economic values of the mangrove ecosystem have been estimated from 31,565,720 to 34,620,100 VND/year (price in 2002, currency exchange rate, US$1= VND15,300) (Nhuan et al., 2003) Therein, invertebrate production contributes an important economic value and livelihood for local communities, estimating VND30,000 per local person/day by selling crabs, shrimps, and oysters (Nhuan et al., 2009) In the year 1989, XTP is declared as the first wetland of International importance of Southeast Asia (http://www.ramsar.org) In the year 2004, through UNESCO XTP is designated to be the most important sub-zone of the Red River Delta Biosphere Reserve (http://www.unesco.org) The characteristics of seasons, tides, salinity, and other environmental conditions are well described elsewhere (Tue et al., 2012) Briefly, the XTP has a distinct monsoon climate with a rainy season from June to October, and a dry season from November to May The tides are characterized with tidal amplitudes ranging from 1.5 to 1.8 m, and the maximum and minimum tidal levels are 3.6 m and 0.5 m, respectively The salinity ranges from 12.5 to 25.6‰ for the rainy season and from 21.1 to 26.5‰ for the dry season 2.2 Field sampling Field work was conducted from 28 January to 10 February 2008 Invertebrates were collected at two sites in the XTP (Fig 1a) The sampling transect is started at the tidal flat and creek bank toward the dense mangroves at sites (Fig 1b) and (Fig 1c), respectively The benthic invertebrate specimens were manually collected by hand during low tides Total invertebrate specimens were 181, belonging to 30 species (Table 2) The species of polychaete, bivalves (Geloina coaxans and Laternula truncata), gastropods, and grapsid crabs were collected within mangrove forests The crabs of Leucosiidae, Ocypodidae families, and portunid crab (Scylla serrata) were collected from fringe mangrove forests, tidal flat, and creek banks Prawns and portunid crab (Charybdis helleri) were collected from Tra Creek by the gill nets during spring and ebb tides The number of invertebrate samples is shown in Table These invertebrates were selected in the present study because they are predominant and high economic values in the XTP (Cuong and Khoa, 2004) In general, the benthic microalgae (BMA) production is low in the mangrove forests due to the high tannin concentration and low light Fig Sampling sites within the mangrove ecosystem of Xuan Thuy National Park, Vietnam (a), the sampling transect at site (b) and at site (c) In each sampling transect the habitats and invertebrate samples are shown (see the sampling methods for details) Refer to Table for acronyms of macro-invertebrate samples 16 N.T Tue et al / Journal of Sea Research 72 (2012) 14–21 penetration (Alongi, 1994) However, the BMA production has been considered as an important food source for invertebrates in the mangrove ecosystem (Bouillon et al., 2002a; France, 1998) In present study, the BMA samples were collected at five sites in the tidal flat and creek bank to examine the contribution of this organic matter source to the invertebrates The BMA samples were extracted from conspicuous layers on surface sediments according to the procedure described by Bouillon et al (2002a) The invertebrate and BMA samples were packed in labeled polyethylene bags and immediately stored in iceboxes, transported to the laboratory for further processing and were frozen at −20 °C until analysis 2.3 Sample preparation and analysis In the laboratory, the polychaete species were kept alive for 24 h in filtered seawater to allow them to excrete their gut contents The invertebrate specimens were washed by distilled water and wiped with paper And then, the carapace width of crabs and prawns and shell size of mollusks were measured The invertebrates were dissected and only their muscle tissues were taken for stable isotope analysis The muscle tissues were dried at 60 °C for 24 h, and then ground to a fine powder by an agate mortar and pestle Post et al (2007) showed that δ 13C can be altered by changes in lipid contents of invertebrates In the present study, the lipid content is extracted from the invertebrate tissues prior to stable isotope analysis The pulverized muscle tissue was then placed in an Eppendorf tube and immersed in a 2:1 chloroform:methanol solution for 24 h to extract lipids After lipid extraction, the muscle tissue was dried in an electric oven at 60 °C for 24 h BMA was treated with N HCl for δ 13C analysis according to the procedure for sediments described by Tue et al (2012), and sub-sample of BMA for δ 15N analysis was not treated with acid The muscle tissues and BMA samples were packed in tin capsules and analyzed for stable isotope signatures (δ 13C and δ 15N) by using a stable isotope mass spectrometer (ANCA-GSL; Sercon Inc., UK) Stable isotopic compositions are expressed by δ values, which are measured as a ratio of the heavy to light isotopes in a sample relative to a standard À À by an equation: X ị ẳ Rsample =Rstandard 1ị 1;000, where X is the isotope value in permil (‰) and R is the ratio of the heavy to the light isotope of the sample (Rsample) to a standard (Rstandard) For stable carbon and nitrogen isotope ratios, R is 13C/12C and 15N/14N, respectively The standards were Pee Dee Belemnite (PDB) limestone carbonate for δ13C and atmospheric nitrogen for δ15N During analysis processes, L-histidine was used as certified reference material Analytical errors were 0.1‰ for δ 13C and 0.2‰ for δ 15N, respectively Linear regression analysis was used to examine the size-specific shifts in δ 13C and δ 15N within each invertebrate species The significance level of linear regression was 0.05 (p b 0.05) for each statistical procedure Results and discussion 3.1 Background data for potential organic carbon food sources of invertebrates The characteristics of major primary producers and other potential organic carbon food sources of invertebrates in the XTP have been reported elsewhere (Tue et al., 2012, 2012) The stable isotope compositions of the potential food sources are sumarized in Table The δ13C of the potential food sources ranged from −29.9± 0.5 to −20.2± 0.6‰ The δ 13C values were increased as follows: mangrove leaves, mangrove sediments, tidal flats, creek bank and bottom sediments, particulate organic matter (POM), marine phytoplankton, and BMA The δ15N values of the potential food sources ranged from 0.7 ± 0.6 to 3.9 ± 0.9‰ The δ 15N values showed an increasing trend from Table Stable carbon and nitrogen ratios of the potential organic carbon food sources in the mangrove ecosystem of Xuan Thuy National Park, Vietnam Organic matter sources ACR δ13C δ15N n References 0.9 0.6 0.2 0.3 0.9 0.6 0.4 9 24 12 Tue et al (2012) 3.9 3.3 0.9 1.1 5 3.4 1.2 13 Mean SD Mean SD − 27.2 − 27.3 − 29.9 − 21.2 − 23.9 − 20.2 − 25.9 0.9 0.6 0.5 0.5 0.8 0.6 1.4 1.6 0.7 2.4 3.6 2.5 2.3 4.3 Cbs Bcs − 24.1 − 24.0 1.2 0.9 Tfs − 24.2 1.0 Mangrove leaves Kandelia obovata Aegiceras corniculatum Sonneratia caseolaris Marine phytoplankton Creek POM Benthic microalgae Mangrove sediments Mang Kao Aec Soc Phyto POM BMA Msed Adjacent habitat sediments Creek bank sediments Bottom creek sediments Tidal flat sediments Ased This study Tue et al (2012) Tue et al (2012) ACR: acronym; n: number of samples; SD: standard deviation mangrove leaves, though to BMA, POM, phytoplankton, and to creek bank and mangrove sediments The stable isotope compositions of BMA from the present study were consistent with previous reports in mangrove ecosystems from Coringa Sanctuary, India (Bouillon et al., 2002a, 2004) and Sikao Creek, Thailand (Kon et al., 2007) 3.2 Food sources of invertebrates in the mangrove ecosystem 3.2.1 Stable isotope compositions of invertebrates From the taxa measured for stable isotopes, the δ 13C values of invertebrates ranged from −14.5 to −26.8‰ (Table 2, Figs and 3) The lowest δ 13C values were expressed in two polychaete species, followed by bivalves L truncata and G coaxans, grapsid crabs Episesarma versicolor and Metopograpsus messor, snails Cassidula aurisfelis, Cerithidea ornata, and Littoraria melanostoma, and grapsid crab Sesarma dehaani Highest δ 13C values were expressed in ocypodid crabs Uca acuta, Uca borealis, Uca flammula, grapsid crab Metaplax elegans, and followed by ocypodid crab Uca urvillei, slug Onchidiidae spp., crab of family Leucosiidae, ocypodid crab Uca arcuata, the portunid crab C helleri, the grapsid crab Metaplax longipes, prawns, and the portunid crab S serrata (Table 2, Fig 2) The δ15N values ranged from 1.3 to 12.1‰, the grapsid crab M elegans expressed the lowest average δ15N value, followed by the snails L melanostoma, C ornata, C aurisfelis, the grapsid crabs M messor and S dehaani, slug Onchidiidae spp., and bivalves L truncata and G coaxans Highest δ15N values were expressed in prawns of Palaemonidae and Penaeidae families, and polychaete (Diopatra neapolitana), followed by the portunid crabs C helleri and S serrata, polychaete (Nephthys polybranchia), fiddler crabs, and tidal flat bivalves Glauconome virens and Ensis magnus (Fig 2, Table 2) 3.2.2 Food sources of invertebrates in mangrove ecosystem 3.2.2.1 Polychaete The polychaete N polybranchia had the δ 13C value in the range of mangrove leaves (Fig 2) The δ 13C value of polychaete D neapolitana was similar to that of mangrove sediments or more enriched 1–2‰ than that of mangrove leaves These results indicated that two polychaete species may obtain the carbon food sources from mangrove materials such as decomposed leaves on the forest floor and/ or scavenge mangrove detritus in the mangrove sediments δ13C signatures of polychaete confirmed that the feeding guild of D neapolitana is herbivore and/or scavenger (Fauchald and Jumars, 1979), and suggested the feeding guilds of N polybranchia may be a motile subsurface depositfeeder More depleted in 13C of polychaete (i.e., N polybranchia) compare N.T Tue et al / Journal of Sea Research 72 (2012) 14–21 Table Stable isotope signatures (δ13C and δ15N) of invertebrates in Xuan Thuy National Park, Vietnam Species Polychaete Diopatra neapolitana Nephthys polybranchia Gastropods Onchidiidae sp Cassidula aurisfelis Dostia violecea Littoraria melanostoma Terebralia sulcata Nassarius olivaceus Cerithidea ornata Bivalves Ensis magnus Glauconome virens Laternula truncata Geloina coaxans Crabs Grapsidae Metopograpsus messor Sesarma dehaani Episesarma versicolor Metaplax elegans Metaplax longipes Leucosiidae Ebalia malefactrix Ocypodidae Uca urvillei Uca flammula Uca borealis Uca acuta Uca arcuata Portunidae Scylla serrata Charybdis helleri Prawns Palaemonidae Macrobrachium rosenbergii Penaeidae Metapenaeus joyneri Penaeus merguensis Penaeus monodon ACR L (mm) δ15N δ13C Mean SD Mean SD Mean n SD 0.8 − 25.5 0.1 − 26.8 Dn Np 52.7 42.0 2.1 11.3 1.0 8.7 On Ca Dv Lm Ts No Co 40.3 25.2 16.6 20.6 26.1 16.0 23.0 2.0 2.2 1.9 3.6 4.9 1.0 1.4 6.2 5.6 7.3 4.1 9.3 8.3 6.8 Em Gv Lt Gc 52.1 30.3 49.7 28.3 6.5 4.2 1.2 1.9 7.5 9.2 5.7 7.1 0.8 0.5 0.8 0.3 Mm Sd Ev Me Ml 11.8 17.7 18.0 7.9 16.0 0.7 1.2 0.5 3.2 5.0 7.8 8.7 3.3 9.9 1.0 − 22.7 0.9 − 22.6 − 23.8 0.7 − 16.7 0.3 − 18.5 Ebm 18.2 0.7 9.3 0.6 − 17.6 1.1 Uu Uf Ub Ua Uar 13.7 25 25.5 21.5 25.7 2.1 8.0 9.0 8.9 9.0 8.8 0.6 − 17.1 0.5 − 16.3 − 15.7 − 14.5 0.5 − 19.0 0.9 2 Ss Ch 49.5 51.8 18.0 10.0 14.4 10.6 Mr 74.9 9.3 10.9 Mj 81.6 Pm 62.6 Pmo 57.1 6.5 11.0 14.1 11.1 4.3 10.4 0.5 0.5 2.7 − 17.7 − 22.8 − 19.2 − 22.1 − 19.6 − 20.6 1.3 − 22.4 1.1 1.5 11 0.6 10 2.5 18 0.7 − 21.8 − 21.3 − 24.6 − 24.3 0.2 0.5 11 0.7 0.3 0.7 3.5 2.8 0.9 0.9 0.8 1.3 1.1 11 0.5 − 19.4 1.8 17 0.6 − 17.6 1.0 0.3 − 19.5 1.2 0.3 − 19.8 0.3 0.6 − 19.0 1.2 11 0.3 − 17.9 0.6 ACR: acronym; n is number of samples; mean, and mean and SD values are given where n = 2, and n ≥ 3, respectively; L: body length, for crab is carapace length, and other species is total body length 17 to that of other benthic fauna was also reported in Bouillon et al (2002b), apparently a preferential food source of polychaete living in dense mangrove forests was the mangrove detritus The polychaete δ15N values were high (Table 2, Fig 2), indicating that they occupied higher trophic position in the benthic food web The trophic position suggested these polychaete species were also nourished on the enriched 15 N food sources (i.e., animal tissues) The Nephtyids can feed on small invertebrates, including mollusks, crustaceans, and other polychaete The Onuphids may nourish by the rotten animal tissues as the supplemental nutrient food sources (Fauchald and Jumars, 1979) Nevertheless, this explanation needs to be confirmed by an experimental determination of the variation of isotopic compositions of polychaete 3.2.2.2 Gastropods Two individuals of slug Onchidium spp had an average δ13C value (−17.7‰) close to that of the BMA, and δ 15N values were higher from to 4‰ than that of BMA (Fig 2) The isotopic compositions obviously indicated that Onchidium spp grazed on the conspicuous layer of BMA on surface sediments The mangrove snails showed a wide range of δ 13C, from −23.7 to −16.4‰ (Fig 3a) The wide range of δ 13C signatures indicated the heterogeneous diets of snails (Penha-Lopes et al., 2009; Rodelli et al., 1984) that can be explained by several factors, such as (1) feeding guilds (Guest et al., 2004), (2) ontogeny (Fratini et al., 2004; Slim et al., 1997), and (3) microhabitats (Fratini et al., 2004; Penha-Lopes et al., 2009) As shown in Fig 3a, the δ13C values of the snails were within the ranges of the BMA, marine phytoplankton, POM, and sediment organic matter of adjacent habitats or slightly higher than the mangrove sediments The δ13C values from the present study indicated the preferential BMA, marine phytoplankton, and POM over the mangrove leaves of the mangrove snails These findings are consistent with the results of diets of Terebralia sulcata and L melanostoma from mangrove ecosystems of Okinawa, Japan (Meziane and Tsuchiya, 2000) and Coringa Wildlife Sanctuary, India (Bouillon et al., 2002a), respectively The food sources of snail T sulcata from the present study were contradictory with the stomach content analysis that reported for mangrove snails of genus Terebralia such as T palustris (Fratini et al., 2004; Penha-Lopes et al., 2009; Slim et al., 1997) Slim et al (1997) showed that the stomach of adult and juvenile snail T palustris contained up to 62.5% and 19% of mangrove materials, respectively The contradictory results between the two methods can be explained by the snails that ingested the mangrove litters but not always assimilated (Fry and Ewel, 2003; Lee et al., 2001) Hence, the mangrove materials probably did not form an important contribution to energy intake by these mangrove snails Fig Dual isotope plot of the δ15N and δ13C (mean ± SD, ‰) values of invertebrates from the mangrove ecosystem of Xuan Thuy National Park, Vietnam The point denotes the mean value and error bar denotes SD Boxes indicate the range of stable isotope compositions of the potential organic carbon food sources of invertebrates Refer to Tables and for acronyms of the potential organic carbon food sources and invertebrates, respectively 18 N.T Tue et al / Journal of Sea Research 72 (2012) 14–21 Fig Frequency distribution of δ13C values of invertebrate groups from the mangrove ecosystem of Xuan Thuy National Park, Vietnam The gray bar indicates the ranges of the potential organic carbon food sources as referred in Table Subsequently, fecal materials of snails may contain large undigested mangrove materials (Lee et al., 2001) In the ecological aspect, the fecal materials can be quickly increased in bacteria biomass and nutrient contents that can be easily assimilated by other deposit-feeders As a result, snails play as an important linkage between mangrove primary producer and other invertebrates (Lee, 2008) Mangrove snails change their microhabitats with the life stages (Fratini et al., 2004; Penha-Lopes et al., 2009) Juveniles prefer the adjacent habitats such as creek bank and tidal flat and fringe mangrove forest They will migrate into the landward zone after reaching to adult stages (Fratini et al., 2004; Penha-Lopes et al., 2009) In addition, many deposit feeders shift their diets during the development to meet their nutrient demands (Hentschel, 1998) The ontogenetic shift in diets was reported for the snail of genus Terebralia (i.e., T palustris) from mangrove forests of Gazi Bay, Kenya (Slim et al., 1997), Inhaca Island, Mozambique (Penha-Lopes et al., 2009) and Malindi, Kenya (Fratini et al., 2004) These authors reported that the juvenile snails are detritivorous and the adults are mainly leaf-litter consumers Our results showed that δ13C was significantly negatively correlated with the shell size of L melanostoma (regression line: δ13C = −0.1 × shell_size − 20.1; R²= 0.39, p b 0.05, Fig 4a) and of T sulcata (regression line: δ13C = −0.44 × shell_size− 8.1; R² = 0.73, p b 0.05, Fig 4b) These results demonstrated that the ontogenetic shift in diets may also occur with these snail species The small snails may graze on enriched in 13C food sources such as the BMA, marine phytoplankton, and POM They will shift to depleted 13C food sources such as sediment organic matter or mangrove materials when they reach to adult stages For the L melanostoma species, they are generalist grazers, grazing on surfaces of the substrata non-selectively (Lee et al., 2001) In the present study, the snails L melanostoma were found abundant on mangrove roots and stems where they ingested the plant tissues and filamentous algae on the plant surface Unfortunately, we could not measure the stable isotope compositions of the filamentous algae on mangrove roots and stems However, the stable isotopic values of the filamentous algae from Bouillon et al (2002a) suggested that the snail L melanostoma fed on mixed diet that is probably composed of filamentous algae and mangrove materials, with the latter sources increasing as with the size of L melanostoma The trophic dimorphism between juveniles and adults has been explained by a difference in radula morphology of gastropods (Slim et al., 1997) 3.2.2.3 Bivalves Four filter feeding bivalve species can be separated by the δ13C signatures (Table 2, Figs and 3b) The δ13C values of two bivalves G coaxans and L truncate living in the dense mangrove forests were in the range of POM and mangrove sediments (Table 2; Fig 2) The mangrove detritus proportion in POM was >50% during the flood tide in the small creeks of this mangrove ecosystem (Tue et al., 2012) As a result, the mangrove detritus may contribute a significant proportion to diets of these mangrove bivalves This finding is consistent with the report on the diet of bivalve G coaxans in the Okinawa mangrove ecosystem (Bachok et al., 2003) Fig Correlation between shell size and δ13C (pb 0.05) of gastropod and bivalve species L melanostoma (a), T sulcata (b), and G virens (c) N.T Tue et al / Journal of Sea Research 72 (2012) 14–21 The δ 13C signatures of two bivalves G virens and E magnus living in the tidal flat were in the ranges of marine phytoplankton and BMA, and slightly enriched than that of POM Thus, they probably had major diets from the POM, and the microalgae, consisting of marine phytoplankton (Rodelli et al., 1984) and BMA resuspension from surface sediments by hydrodynamics (Cognie et al., 2001; Kang et al., 1999) Interestingly, the measurements of δ 13C values and shell sizes were significantly negatively correlated for individuals of G virens species (regression line: δ 13C = − 0.1 × shell_size − 18.6; R² = 0.57, p b 0.05, Fig 4c), implying the ontogenetic shift in diets of the G virens from the microalgal diets to POM containing high proportion of mangrove detritus The difference in food resource utilization of juvenile and adult G virens may relate to the mechanisms of selective feeding, strength of inhalant flows, siphon sizes, and lipid contents of filterfeeding bivalves (Kang et al., 1999) 3.2.2.4 Crabs The stable isotope compositions of these brachyurans varied with the habitats, feeding behavior, and taxonomy (Table 2; Fig 2) In general, the crabs fed on variety of food resources, consisting of BMA, marine phytoplankton, POM, and sediment organic matters from adjacent habitats and mangrove forests (Table 1; Figs and 3c) For grapsid crabs, the average δ13C value of M messor was −22.7 ± 0.9‰ (n= 6), indicated that the grapsid crab M messor did not directly rely on the mangrove leaves The Metopograpsus spp has been reported to be an omnivorous that nourished animals, plant materials, and inorganic sediments (Poon et al., 2010) However, our results may not support the omnivorous feeding guild of the M messor in the XTP due to the low δ15N values in the muscle tissues (Table 2) In addition, δ13C was slightly higher than sediment organic matters of mangrove and adjacent habitat sediments, and POM (Fig 2) The stable isotope compositions thus suggested that the grapsid crab M messor fed on a bulk mixture of sediment organic matters with high proportion of mangrove detritus The δ 13C of sesarmid crab S dehaani was slightly enriched relative to that of the mangrove sediment organic matters (Table 2, Fig 2), and the δ 15N was 3.5‰ higher relative to these substrates The isotopic compositions demonstrated that the sesarmid crab S dehaani fed on the organic matters in the mangrove sediments, which composed of the marine phytoplankton and mangrove detritus, with the later sources being predominant (Tue et al., 2012) Only one specimen of sesarmid E versicolor was collected in the present study, thus it is not obvious to indicate the food sources of this species However, the stable isotope compositions from the present study are similar to those reported for the E versicolor from Indian mangrove forest (Bouillon et al., 2002a) and Indonesian mangrove forest (Nordhaus et al., 2011) These patterns suggested that the sesarmid E versicolor may have alike diets in different mangrove forests The diets of E versicolor included sediment organic matter, other invertebrates, carrion, and mangrove litter thereof (Nordhaus et al., 2011) The δ 13C values of two Metaplax species and the pebble crab Ebalia malefactrix were higher than those of other sesarmid and grapsid crabs (Table 2, Fig 2) The high δ 13C values indicated that mangrove organic carbon was not significantly contributed to diets of these crabs The δ13C signatures of two Metaplax species showed that they nourished the BMA, e.g., benthic diatoms and cyanobacteria (Bouillon et al., 2002a) Moreover, the species M longipes had significantly higher δ15N than the species M elegans (pb 0.05) The δ15N values demonstrated that the species M longipes fed on a higher trophic position than the species M elegans The stable isotope compositions showed that the species M longipes ingested the BMA, animal carrions, and juveniles of gastropods and bivalves For fiddler crabs (Uca spp.), δ13C values varied from −20.6 to −14.3‰, with the most enriched in 13C for U acuta species (δ 13C values of two individuals were −14.3‰ and −14.6‰) However, δ 15N of the Uca spp species had a small range, varying from 7.3 to 9.5‰ The small range of δ15N indicated that these fiddler crabs fed on the same 19 trophic level The fiddler crabs are conspicuous residents of the land– water ecotone in the mangrove ecosystem (France, 1998) They are deposit feeders, ingesting organic matter from the exposed mud at low tide (Hogarth, 2007) Therefore, the δ 13C values suggested that they nourished nutrients from surface sediments with a major food source from the BMA (Fig 2) Two species U arcuata and U urvillei collected from the tidal flat at site (Fig 1b) were enriched in 13C compared to the BMA, referring to the preferential diets from the BMA The species U borealis was collected from tidal flat and two species U flammula and U acuta were collected from the creek bank (Fig 1c), had δ13C values higher from 2.6 to 4.6‰ than that of the BMA, suggesting that they may not rely only on the BMA food source Moreover, the δ15N values of these fiddler crabs were 5.0 to 7.2‰ greater than that of the BMA The 15N enrichment has probably ruled out the substantial dietary contribution from the BMA, given the expected fractionation 3–4% enrichment in δ 15N per trophic level (Post, 2002) Obviously, these fiddler crabs nourished other food sources with/without BMA such as bacteria (France, 1998), and ciliate protozoa and nematodes (Hogarth, 2007) In agreement with our study, the fiddler crabs in the mangrove ecosystems of Malaysia (Rodelli et al., 1984), Puerto Rico (France, 1998), and Coringa Wildlife Sanctuary, India (Bouillon et al., 2002a) were highly selective for the BMA and other food sources (i.e., bacteria, ciliate protozoa, and nematodes) than the mangrove detritus The stable isotope compositions of two portunid crabs showed that they are among the top predators in the mangrove ecosystem (Fig 2) In Thailand mangrove forests, the portunid crabs feed on the slow moving benthic animals, including bivalves, snails, other crabs, and polychaete (Thimdee et al., 2004) Our results suggested that the portunid crabs probably relied on the various types of invertebrates (i.e., M longipes, T sulcata, Nassarius olivaceus, Dostia violecea, and G virens) Moreover, our findings showed that δ13C exhibited a significant negative correlation with carapace width >30 mm of portunid crab S serrata (regression line: δ13C = −0.1 × carapace_width − 14.4; R² = 0.6, p b 0.05, Fig 5) This pattern suggested that the larger individuals S serrata extensively fed on other invertebrates in the lower trophic levels (i.e., C ornata, S dehaani, and M messor) and microphytobenthos for enough nutrient demand 3.2.2.5 Prawns Stable isotope composition for three penaeid shrimps and Macrobrachium rosenbergii species were tightly clustered (Fig 2), which indicated of an assessment to the similar food resources As shown in Fig 3d, the major food sources of these penaeid shrimps were BMA, marine phytoplankton, POM, and sediment organic matters In addition, the δ15N signatures of the penaeid shrimps were highest that are indicative of the top predators among the invertebrates in the mangrove ecosystem of XTP The high δ15N signatures were indicative of a feeding on other small preys such as juveniles of crabs, gastropods, and bivalves In agreement with our study, Chong and Sasekumar (1981) showed that penaeid shrimps are opportunistic omnivorous and are known to feed on a variety of food — depending on the locality and availability of food items Fig Correlation between carapace width and δ13C (pb 0.05) of portunid crab S serrata The square symbols are the values of individuals with carapace width b 30 mm, and these individuals were not used to calculate the regression between carapace width and δ13C 20 N.T Tue et al / Journal of Sea Research 72 (2012) 14–21 Conclusions The dual stable isotope signatures (δ 13C and δ 15N) were applied to identify the utilization of food sources by invertebrates in an ecologically important mangrove ecosystem of Vietnam The results showed that the invertebrates had heterogeneous diets and just a few mangrove inhabitants such as polychaete, gastropods, bivalves, and grapsid crabs directly relied on the mangrove detritus In addition, fiddler crabs living in the land-water ecotone were highly selective for the BMA and other food sources (i.e., bacteria, ciliate protozoa, and nematodes) than the mangrove detritus The present study supports other isotopic studies that invertebrates assimilate a greater production of high nutrient food sources such as BMA, marine phytoplankton, meiofauna, and animal carrions in the mangrove ecosystem The ontogenetic shifts in diets were identified for several invertebrate species, consisting of snails L melanostoma and T sulcata, bivalve G virens, and portunid crab S serrata On the basis of the δ13C and δ15N signatures, feeding guilds of invertebrates could be divided into low trophic position, consisting of snails, bivalves, grapsid crabs (exclude: M longipes) and high trophic position (polychaete, grapsid crab M longipes, portunid crabs, and prawns) This study revealed that the snails may play as a linkage between mangroves and other invertebrates in the high trophic positions, and the estuarine food webs (Proffitt and Devlin, 2005) In addition, the land–water ecotone such as tidal flats and creek banks seem to contribute an important microalgal food resource for 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mangrove ecosystem of Xuan Thuy National Park, Vietnam Estuaries and Coasts 35, 1060–1068 ... Conclusions The dual stable isotope signatures (δ 13C and δ 15N) were applied to identify the utilization of food sources by invertebrates in an ecologically important mangrove ecosystem of Vietnam The... materials is assimilated by invertebrates in an ecologically important mangrove ecosystem of Vietnam The objectives of the present study are (1) to determine the utilization of food sources by. .. intake by these mangrove snails Fig Dual isotope plot of the δ15N and δ13C (mean ± SD, ‰) values of invertebrates from the mangrove ecosystem of Xuan Thuy National Park, Vietnam The point denotes