Spatial variation of ostracod (Crustacea, Ostracoda) egg banks in temporary lakes of a tropical flood plain

Additional keywords: dormancy, hatching, microcrustaceans, passive community.

Some microcrustaceans produce resting eggs to survive periods of habitat desiccation (Brendonck and De Meester 2003). These eggs accumulate in the sediment, forming egg banks (Brock et al. 2003). Such resting egg banks constitute ecological and evolutionary reservoirs that contribute to the (re)colonisation and resilience of aquatic environments after disturbances (Brendonck and De Meester 2003; Brock et al. 2003).

Differential accumulation of resting eggs in the sediment can lead to differences in the composition and abundance of passive communities in distinct compartments of waterbodies (Brendonck and De Meester 2003; Vandekerkhove et al. 2005; Gerhard et al. 2017). The spatial patterns of the active microfaunal communities are well studied, but those of passive communities generally remain little known (Gerhard et al. 2017; Portinho et al. 2017). Nevertheless, some studies on active communities suggest that spatial variation in the environment is regulated by the production of resting eggs and the dispersal of these propagules (Hairston 1996).

Spatial variation within lakes and pools in the abundance and composition of resting eggs can occur as a result of two processes: (1) resting eggs can become part of the floating debris in drying pools and thus accumulate along the edges (Martens et al. 1992); (2) during the drying of these environments, the eggs accumulate in the deepest regions, mostly the centre of waterbodies, where the water remains the longest before full desiccation (Martens et al. 1992; Bright and Bergey 2015). However, in flood plains, during flood periods with high water levels, most of the environment is inundated and becomes connected to a main river channel. It is then expected that the water flow promotes the dispersal of resting eggs and consequently homogenises the egg banks in the sediments of the lakes (Thomaz et al. 2007; Bozelli et al. 2015), so that composition at the edges and centre of the waterbody would become similar in any season of the year.

Resting eggs that settle at the water–sediment interface of the waterbody may adhere to sediment particles (e.g. organic matter) and may be covered by them (Brendonck and De Meester 2003). Thus, the abiotic variables of the sediment may affect the viability, dispersal and hatchability of resting eggs. For example, the amount of organic matter has an effect on oxygen concentrations in the sediment, affecting the viability and hatchability of resting eggs (Rossi et al. 2004). In addition, the smaller the sediment particles, the greater their capacity for suspension, which increases the probability of dispersal of resting eggs (Constable 1999).

Some groups of freshwater ostracods can produce resting eggs. Most species of the family Cyprididae, which comprises approximately half the total number of extant non-marine ostracod species (Meisch et al. 2019), are known to produce resting eggs (Horne and Martens 1998). Several experiments have investigated the viability and/or hatching phenology of ostracod eggs of Eucypris virens (Jurine, 1820) and Heterocypris incongruens (Ramdohr, 1808) in temperate regions in the Palaearctic (e.g. Rossi et al. 2012; Vandekerkhove et al. 2013). However, no studies have focused specifically on resting eggs of ostracods in the Neotropical region, although some studies on the resting eggs of zooplankton and other invertebrates have occasionally also recorded ostracods hatching from egg banks (Stenert et al. 2010; Cardoso Ávila et al. 2015; Santangelo et al. 2015; Vargas et al. 2019).

Studies on ostracod resting eggs can provide vital information on the biology of the group. Ecological information about the recruitment of organisms from egg banks can contribute to science-based recommendations for the conservation of aquatic environments, such as temporary lakes in flood plains. These ecosystems are threatened by anthropic impacts (e.g. the construction of artificial dams) that affect the hydrological dynamics of the ecosystem (droughts and flood events; Agostinho et al. 2004a), and thus require protective measures. In this study we evaluated the spatial variation of ostracod resting eggs in different regions of five temporary lakes of the Upper Paraná River flood plain. Based on the homogenisation effect of flooding in the flood plain lakes, we hypothesised that the composition and abundance of ostracod eggs in the centre of the temporary lakes would be similar to those in the edge regions.

The Paraná River is formed by the junction of the Grande and Parnaíba rivers in south-central Brazil, and is the second largest river in South America (4695 km long; Agostinho et al. 2008). The Upper Paraná River has a large catchment (~802 150 km²) in Brazil, which encompasses large parts of the states of Paraná, São Paulo, Mato Grosso do Sul, Minas Gerais and Goiás (Souza-Filho and Stevaux 2004). The upper part of this river comprises a flood plain, which includes a series of small islands and a variety of environments, such as channels, rivers, connected and isolated lakes and temporary lakes. The Upper Paraná River flood plain is located in the Area de Proteção Ambiental das Ilhas de Várzea do Rio Paraná (Environmental Protection Area; Agostinho et al. 2004b). The climate of the region is tropical–subtropical with distinct rainy (November–March) and dry (April–October) seasons (Köppen Cfa; Eletrosul 1986; Agostinho et al. 2004b).

In this study we investigated the passive ostracod communities of five temporary and isolated lakes located on Porto Rico Island (Pontal: 22°45′05.7″S, 53°15′23.6″W; Clara: 22°45′20.7″S, 53°15′27.7″W; Figueira: 22°45′22.7″S, 53°15′34.0″W) and Mutum Island (Pousada: 22°44′43.4″S, 53°14′06.9″W; Osmar: 22°46′28.6″S, 53°19′58.8″W) in the Upper Paraná River flood plain (Fig. 1). These lakes are shallow (not exceeding a depth of 2.2 m) and small (area ≤0.15 ha). The limnological variables (e.g. dissolved oxygen (DO), electrical conductivity (EC), pH) and vegetation cover formed by herbaceous and arboreal species, with a higher presence of emergent macrophytes of the Family Poaceae (Kita and De Souza 2003), are similar among the five lakes. In the dry season of 2017, four lakes were dry (Pontal, Clara, Figueira and Pousada) and one lake (Osmar) had a low water level (<30 cm) during sampling.

Fig. 1.  Location of the temporary lakes of the Upper Paraná River flood plain. Flow direction is from right to left.

Sediments were collected during the dry season (September 2017) at the edge and in the centre of the five temporary lakes. The edge region was defined as the area in direct contact with the adjacent terrestrial ecosystem, mostly with shallower sites, whereas the centre region was defined as the open area of the lake, associated with deeper sites. Sediment was sampled to a depth of 6 cm (~250 g) using a core sampler (volume 194.5 cm³) in each region of the lakes, because resting eggs with higher viability are usually found in the top 3 cm of the sediment (García-Roger et al. 2006).

Twelve samples were collected at the edge of each of the lakes in order to cover the entire edge region of the lake, whereas five samples were collected in the centre. These samples were taken every 3 m (4.250 kg sediment per lake). A larger number of samples was collected at the edge of each lake compared with the central region to better cover the spatial representation of this variable environment. For each lake, samples of each region (edge and centre) were separately pooled to form a composite sample, yielding 10 samples in total (2 regions × 5 lakes). The sediment was stored in plastic bottles and was kept refrigerated for 2 months, following the methods described by Maia-Barbosa et al. (2003). Despite the fact that the lakes were dry (no or very little water), the sediment was mostly still moist.

For the hatching procedure in the laboratory, the composite samples from each region (edge and centre) of the five lakes were separately homogenised and 300 g sediment was individually oven-dried at 50°C (this sediment temperature can be easily reached in the floodplain lakes on hot and dry days) and then placed in individual plastic trays, which acted as artificial microcosms (n = 10 microcosms in total). Each dry sediment sample was hydrated with 500 mL distilled water (see Supplementary Material Fig. S1a, available at the journal’s website) and was maintained in the microcosm at 25°C (Rossi et al. 2004) for 91 days (fig. S1b) in a germination chamber (Model SOLAB, SL.225, Solab, São Paulo, Brazil) under a 12-h light–dark regimen (Rossi et al. 2012).

During the incubation period the microcosm were monitored weekly. Every 7 days, the water from the microcosm was filtered using a plankton net (68 μm; fig. S1c). Because the life cycle of non-marine ostracods is at least 3 weeks (Meisch 2000), 7 days was not sufficient for the sexual maturation of individuals for reproduction. The water in the microcosm was replaced with fresh distilled water. The filtered material, retained in the net, was sorted under a stereomicroscope. Hatched juveniles were grown separately in glass bottles with distilled water, fed fresh spinach and reared to the adult stage (when the juveniles did not die) in separate chambers for identification and counting. The ostracod species were identified following Higuti et al. (2010, 2013) and using the references in Martens and Behen (1994).

Sediment from each region (edge and centre) was used to determine particle size and organic matter content. Sediment composition was determined according to the method of Suguio (1973), using the Wentworth (1922) scale. The samples were sorted in a nested series of sieves (size ranging between 2 and <0.63 mm) and weighed. The size of sediment particles was classified as gravel (>2 mm), very coarse sand (2–1 mm), coarse sand (1.0–0.5 mm), medium sand (0.50–0.25 mm), fine sand (0.250–0.125 mm), very fine sand (0.125–0.063 mm) and mud (<0.063 mm). Organic matter in the sediment was obtained from 10 g dry sediment by incineration at 560°C for 4 h (Moretto et al. 2013). The difference between the initial and final weight of the sediment indicates the amount of organic matter present in the sediment.

DO (mg L^–1; YSI oximeter 550A, Yellow Springs, Ohio, USA), EC (μS cm^–1; Conductivimeter-Digimed; Digimed, São Paulo, Brazil) and pH (pH meter-Digimed; Digimed) were measured weekly in the microcosms.

A non-parametric Wilcoxon test for paired samples was used to test the significance of differences in ostracod abundance (number of ostracod specimens hatched from resting eggs) between lake regions because the assumptions of normality and homoscedasticity, required for parametric tests, were not met. For these analyses, the total number of ostracods hatched in all weeks for each region and in each lake was used.

The frequency of ostracod species that hatched from resting eggs was calculated using the constancy index (Dajoz 1973) as follows:

where C is constancy, n is the number of samples in which the species was recorded and N is the total number of samples. The following categories were assigned to ostracod species according to Dajoz (1973): constant (C ≥ 50%), accessory (50% > C ≥ 25%) and accidental or rare (C <25%).

A principal coordinate analysis (PCoA) was performed to visualise (dis)similarity of ostracod species composition between the lakes’ regions (edge and centre) using presence and absence data of the ostracods hatched weekly (Legendre and Legendre 1998). Permutational multivariate analysis of variance (PERMANOVA) was used to evaluate differences in ostracod species composition between the edge and centre (Anderson 2005). This test was based on a dissimilarity matrix using the Jaccard distance. A total of 999 permutations was performed to assess significance.

The relationship between ostracod abundance and sediment quality (sediment composition and organic matter) was examined using generalised additive models (GAMs). The models were constructed with negative binomial distribution to avoid overdispersion, using the particle size of the sediment as an explanatory variable and ostracod abundance as a response variable. Before the models were constructed, pairwise correlations among explanatory variables were evaluated using Spearman’s rank correlation coefficients to avoid multicollinearity.

One model was constructed for each explanatory variable because of the low number of samples. The best models identified were based on comparisons of Akaike’s information criterion (AIC), confidence intervals (CIs) and significant values of the variance test (analysis of variance (ANOVA)).

Finally, the Kruskal–Wallis test was used to evaluate possible significant differences in each limnological variable between the edge and centre. For this, DO, pH and EC data measured weekly in each microcosm for the edge and centre samples were used. In addition, possible differences in organic matter content between edge and centre regions were evaluated using paired-sample t-tests; t-tests were used because they are appropriate for the number and dependency of samples in the present study.

ANOVAs, PCoA and GAM analyses were conducted using R 3.4 (R Development Core Team 2013) using the vegan (ver. 2.4-0, see https://CRAN.R-project.org/package=vegan, verified 20 June 2018; Oksanen et al. 2018), permute (ver. 0.9-4, see https://CRAN.r-project.org/package=permute, verified 20 June 2018; Simpson 2018) and mgcv (ver. 1.8-12, see https://CRAN.R.project.org/package=mgcv, verified 20 June 2018; Wood 2018) packages.

Twelve species of ostracods hatched from the egg banks of the five temporary lakes. Cyprididae was the richest and most abundant family, represented by 11 species. The family Candonidae was represented only by the species Physocypria schubarti Farkas, 1958. Cypridopsis vidua (O. F. Müller, 1776), Cypricercus sp. nov. and Bradleytriebella trispinosa (Pinto & Purper, 1965) were only reared from sediment from the centre of the lakes (Table 1).

Table 1.  Constancy index of ostracod resting eggs hatched in the temporary lakes of the Upper Paraná River flood plain
Constant species (▪) were present in more than 50% of samples, accessory species (▪) were present in 25–50% of samples and accidental species (▪) were present in <25% of samples. (□), absent from samples

In all, 553 ostracod specimens hatched from the sediments of the five temporary lakes; 144 and 409 ostracods were recorded from the edges and centres respectively (Fig. 2). There was no significant difference in the number of individuals between the two regions (Wilcoxon test, P = 0.07). Chlamydotheca colombiensis Roessler, 1985 was the most abundant species at the edge and Strandesia mutica (Sars, 1901) was the most abundant in the centre of the lakes. According to the constancy index, S. mutica and C. colombiensis were the most common species in both regions, whereas Strandesia velhoi Higuti & Martens, 2013 was common only in the centre of the lakes (Table 1). The results of PERMANOVA did not show significant differences in the species composition of the egg banks between the centre and edge regions (F = 0.62, P = 0.87; Fig. 3).

Fig. 2.  Mean (±s.e.m.) abundance of ostracods hatched from egg banks at the edge and in the centre of the temporary lakes.

Fig. 3.  Ordination diagram of the principal coordinate analysis (PCoA) of the passive ostracod communities at the edge and in the centre of the five temporary lakes.

Very fine sand was the most dominant type of sediment in the edge regions, whereas mud (clay and silt) was the most dominant type of sediment in the centre regions of the five temporary lakes (Fig. 4). In general, the sediment of the lakes was composed primarily of sediment particles <0.25 mm.

Fig. 4.  Mean (±s.e.m.) sediment composition at the edge and in the centre of the temporary lakes. Mud, silt and clay; VFS, very fine sand; FS, fine sand; MS, medium sand; CS, coarse sand; VCS, very coarse sand; G, gravel.

Coarse particulate organic matter (roots and leaves) was observed in all five temporary lakes. The organic matter content of the sediment was higher at the edge than in the centre (t-test, t = –21.92, P = 0.00; Fig. 5).

Fig. 5.  Mean (±s.e.m.) organic matter content in the sediment at the edge and in the centre of the temporary lakes.

The non-generalised linear model showed positive effects of very fine sand on the number of hatchlings. Conversely, the hatching of eggs was negatively related to the amount of organic matter and sand with a medium grain size (Table 2).

Table 2.  Generalised additive models, model-averaged standardised coefficients, 95% confidence intervals (CIs), Akaike’s information criterion (AIC) and P-values of predictors of hatching of ostracod resting eggs

According to the results of Kruskal–Wallis tests, environmental variables did not vary significantly between the microcosms of the edge and centre regions (DO: H = 3.58, P = 0.058; pH: H = 0.07, P = 0.79; EC: H = 0.01, P = 0.91). Mean DO, pH and EC values in the edge microcosms were 4.95 mg L^–1, 6.74 and 20.77 μS cm^–1 respectively, compared with 4.45 mg L^–1, 6.68 and 18.27 μS cm^–1 respectively in the centre microcosms (Table S1).

Macrophytes germinated from sediments of both regions of the lakes during the incubation period (Fig. S2). Nymphaea amazonum Mart. & Zucc. was the most common macrophyte species recorded in 8 of the 10 microcosms (Table S1).

The hypothesis that the composition and abundance of ostracod resting eggs are similar between the edge and centre regions of temporary lakes in the Upper Paraná River flood plain was supported by the present study. A practical implication of the results of this study is that further investigations into egg banks can be performed using sediment sampled from any region (centre or edge) of temporary lakes in flood plains because of the similar spatial distribution of resting eggs. Theoretically, flood pulses promote the homogenisation of active communities (Thomaz et al. 2007) and may have also contributed to the homogenisation of the passive community (the egg bank). The similar spatial distribution of the egg bank between the edge and centre regions of the lakes may increase the probability of dispersal of ostracod resting eggs by biotic vectors compared with the dispersal of an egg bank that only accumulates eggs in a specific region of the lake. Because several animals (e.g. birds) visit this type of environment, they can promote the dispersal of these structures (Morais Junior et al. 2019).

Spatial variation in the occurrence of resting eggs in waterbodies has been shown for both horizontal (between the edge and centre) and vertical distribution, such as when the sediment floats on the surface of the water after rains, as observed by Martens et al. (1992) in a temporary pool in Israel. Martens et al. (1992) also showed that the floating sediment contained more eggs than the submerged sediments, although the composition did not differ between the regions. In the present study, only samples of submerged sediment (dry or wet) were sampled because the lakes were dry or had very low water levels during the sampling period.

The presence of resting eggs in the centre and at the edge of temporary waterbodies may depend on several factors. For example, drought-resistant eggs can accumulate on floating debris along the edges (Martens et al. 1992), but drying pools will also concentrate fauna at their deepest (mostly central) point towards the end of the hydrological cycle, with resting eggs then produced at a higher rate in these remaining pools. Thus, ostracods may produce a greater number of eggs in the deepest part (centre) of ponds and lakes (Bright and Bergey 2015).

However, both these processes mostly relate to rain-filled, isolated, temporary waterbodies. In the temporary lakes of the Upper Paraná River flood plain, the effect of flood pulses may nullify such processes and may lead to homogenisation (Thomaz et al. 2007), also with regard to the composition and abundance of ostracods between the edge and centre of these lakes. This is because during high-water periods water from the main river invades the lakes, homogenising and dispersing the propagule bank of the dormant communities (Gurnell et al. 2008). The floodwaters will mix the sediments of these temporary lakes and distribute the ostracod resting eggs over the entire lake, thus resulting in a similar composition of the passive ostracod community in central and peripheral areas of the waterbody. Another factor that may contribute to the similarity in species composition is the morphology of the temporary floodplain lakes. These lakes are invariably elongated and narrow, and this possibly facilitates homogenisation between regions in these temporary lakes, because the edge and centre are only a few metres apart.

Nevertheless, differences in the composition and abundance of species of cladocerans in egg banks have been observed in littoral and pelagic zones of shallow waterbodies (Vandekerkhove et al. 2005; Gerhard et al. 2017). Other studies found no differences in the composition of egg banks of invertebrates, including ostracods, among upland, edge and centre regions of playa wetlands (Bright and Bergey 2015). In addition, Bright and Bergey (2015) also reported that the abundance of invertebrate eggs was similar in the edge and centre regions because of environmental factors and passive dispersal by wind and inundation.

All 12 ostracod species hatching from the resting eggs in the present study have been recorded previously from other lotic and lentic environments (e.g. rivers, channels, connected, isolated and temporary lakes) of the river–floodplain system of the Upper Paraná River (Higuti et al. 2010, 2017). Of these 12 species, C. colombiensis, S. mutica, Strandesia variegata (Sars, 1901) and Strandesia bicuspis (Claus, 1892) were originally described from specimens that had been hatched from dried sediment (Sars 1901; Roessler 1985). Interestingly, thus far C. colombiensis was only found in temporary lakes of the Upper Paraná River (Higuti et al. 2010). In the present study, the species hatched equally successfully from sediments from the edge and centre regions of the temporary lakes, indicating that C. colombiensis may be adapted to temporary environments. As expected, most ostracod species that hatched from resting eggs in the present study belonged to the Family Cyprididae, with one species belonging to the Family Candonidae (P. schubarti). A previous study on the diversity of crustacean zooplankton in North America also recorded a species of Physocypria hatching from sediment egg banks (Havel et al. 2000).

Most studies on the production and hatching of resting eggs of ostracods are from temperate regions of the Palaearctic (Martens et al. 1992; Horne and Martens 1998; Valls et al. 2016) and few studies have focused on (sub)tropical regions. Ostracods are known to lay mixed batches of subitaneous and resting eggs, and it is possible that the ratio of these two types of eggs can be affected by environmental factors (Dumont et al. 2002; Schön et al. 2012). This is unlike, for example, Cladocera, where resting eggs (ephippia) are only produced by the final sexual population at the end of the reproductive period (mostly summer).

Abiotic characteristics of the sediment are important for the active community of benthic invertebrates, mainly by providing habitats and substrate for organisms (Hauer et al. 2018), and thus they may also have an effect on the dormant egg banks. In the present study, the positive relationship between the number of the hatched ostracod resting eggs and particle size (very fine sand) may be related to the fact that this type of sediment has a greater capacity for suspension (Constable 1999). This is due to the movement of water or bioturbation activities, which oxygenate the substrate and result in a higher concentration of water in the sediment (Constable 1999), and thus provide better conditions for the hatching and dispersal of resting eggs. These results agree with those of Masero and Villate (2004), who found a positive correlation between the density of calanoid eggs and smaller sediment particles, thus showing that sediment characteristics can affect egg banks. In addition, Tilbert et al. (2019) found a positive association between active ostracods and fine and very fine sand in a small tropical estuary in Brazil.

The negative effect of organic matter content on the hatching of ostracod resting eggs may be linked to increased decomposition and hypoxia in the sediment and water column, which can negatively affect hatching (Rossi et al. 2004; Watkins et al. 2011). In addition, the organic matter in the sediment of temporary lakes was primarily composed of allochthonous (non-aquatic) material, provided by riparian vegetation, mostly leaves of trees, because these temporary lakes have dense vegetation cover (Kita and De Souza 2003). The layers of leaves accumulated in the sediment can also bury and smother the egg banks, reducing the hatchability and viability of resting eggs (Gleason et al. 2003).

The germination of macrophytes in all microcosms may also contribute to ecological succession, because these aquatic plants provide substrate and food for ostracods (juveniles) after the hatching of resting eggs. Several studies have shown the important effect of macrophytes on the structure of active ostracod communities (Higuti et al. 2010; Matsuda et al. 2015).

A possible limitation of the present study is that the experimental condition imposed for artificial incubation may not have provided the required environmental cues for the hatching of all ostracod species present in the egg banks. Although some abiotic variables were controlled in the laboratory, this still does not exactly reflect the characteristics of the natural environment. However, artificial incubations have been used as an effective method to study the egg banks of different communities, such as rotifers (Fernandes et al. 2012), cladocerans (Stenert et al. 2017) and branchiopods (Pinceel et al. 2019).

In conclusion, the composition and abundance of ostracod resting eggs are similar between the edge and central regions of temporary lakes of riverine flood plains, most likely because flood pulses can lead to homogenisation of the ostracod egg banks. However, natural floods are becoming less frequent in this region because of the effect of a cascade of reservoirs upstream of the flood plain. In addition, because of longer periods of drought, reservoirs will retain the water for the production of energy for longer periods of time. Thus, we can infer that a reduction of floods, caused by both natural and anthropogenic effects, would affect the structure and spatial variation of ostracod egg banks in the future.

For now, however, the homogenised distribution of ostracod resting eggs between lake regions may increase the dispersal of these structures by biotic vectors (e.g. birds) owing to the larger distribution area in the environment. In addition, the results of this study have practical implications for the sampling of ostracod egg banks in floodplain lakes, suggesting that sediment samples can be collected from any region of such lakes (edge or centre).

The authors declare that they have no conflicts of interest.

Jonathan Rosa thanks the National Council for Scientific and Technological development (CNPq) and Ramiro de Campos thanks Coordination of Improvement of Higher Education Personnel (CAPES) for granting them scholarships. The authors thank the National Council for Scientific and Technological development (CNPq) Long-term Ecological Research (Programa Ecológico de Longa Duração (PELD)) for funding the collections.