Application of Stable Isotope Tracers to Studies of Zooplankton Feeding,using the Rotifer <Emphasis Type="Italic">Brachionus calyciflorus</Emphasis> as an Example

We present a protocol and calculation methods for the determination of zooplankton ingestion and assimilation rates with stable isotope tracers. These methods have been developed from experiments with the rotifer Brachionus calyciflorus that had been fed ¹³C-labelled Scenedesmus obliquus. Stable isotope tracers offer the same advantages as radioisotopes. These include the possibility for direct and accurate quantification of ingestion and assimilation rates, short sample analysis times and low animal densities requirements. However, the use of stable isotope tracers requires relatively long sample preparation times and specialist equipment and is, thus, relatively costly for most laboratories. The application of stable isotope tracers in zooplankton feeding studies offers several advantages in comparison with radioisotopes. Firstly, they do not emit harmful radiation and can therefore be applied safely both in the laboratory and in the field. Secondly, the samples can be dried for safe storage and easy transportation. Thirdly, no aggressive chemicals are required for sample analysis.     

Feeding responses of Asplanchna brightwelli (rotifera): laboratory and field studies

Laboratory-reared A. brightwelli were offered 16 different prey rotifer species from the genera Anuraeopsis, Brachionus, Epiphanes, Filinia, Hexarthra, Lecane, Lepadella, Pompholyx, Proalides and Trichocerca. Feeding responses (prey encounter, escape, capture, rejection and ingestion) were recorded. In general, the predator's ingestion time was positively correlated with prey length. B. falcatus and H. intermedia were never captured. Once captured, smaller rotifer species were ingested. Gut contents of field-collected A. brightwelli revealed increased occurrence of prey numbers when concentrated plankton samples were preserved after some delay.     

The relationship between food density and short term assimilation rates in Potamopyrgus antipodarum and Deleatidiumsp.

Much of the variation in individual growth rates can be attributed to differences in individual feeding rates. Therefore, in order to build predictive models of individual, or population growth, the factors influencing an individual's feeding rate must be described. An important determinant of the feeding rate is the relationship between the local abundance of food and the individual's ingestion rate – otherwise known as the functional response. We determined functional responses for two species of invertebrate grazers: the snail Potamopyrgus antipodarum and the mayfly Deleatidium sp., by measuring their assimilation rate with increasing densities of radiolabelled periphyton. The assimilation rates were consistent with the Holling Type II or Michelis Menten functional response curve. The parameters of the functional response yielded estimates of the search area and handling time for the stream invertebrates. Our functional response data indicate that the half-saturation food density for P. antipodarum and Deleatidium sp. were 980 mg and 3200 mg AFDM m^–2, respectively, suggesting that Deleatidium growth may be subject to food limitation more often than is P. antipodarum – despite the lower assimilation efficiency of the latter species.     

Experimental study on the impact of dinoflagellate Alexandrium species on populations of the rotifer Brachionus plicatilis

To investigate harmful effects of the dinoflagellate Alexandrium species on microzooplankton, the rotifer Brachionus plicatilis was chosen as an assay species, and tested with 10 strains of Alexandrium including one known non-PSP-producer (Alexandrium tamarense, AT-6). HPLC analysis confirmed the PSP-content of the various strains: Alexandrium lusitanicum, Alexandrium minutum and Alexandrium tamarense (ATHK, AT5-1, AT5-3, ATCI02, ATCI03) used in the experiment were PSP-producers. No PSP toxins were detected in the strains Alexandrium sp1, Alexandrium sp2.Exposing rotifer populations to the densities of 2000 cells ml⁻¹ of each of these 10 Alexandrium strains revealed that the (non-PSP) A. tamarense (AT-6) and two other PSP-producing algae: A. lusitanicum, A. minutum, did not appear to adversely impact rotifer populations. Rotifers exposed to these three strains were able to maintain their population numbers, and in some cases, increase them. Although some increases in rotifer population growth following exposures to these three algal species were noted, the rate was less than for the non-exposed control rotifer groups.In contrast, the remaining seven algal strains (A. tamarense ATHK, AT5-1, AT5-3, ATCI02, ATCI03; also Alexandrium sp1 and Alexandrium sp2) all have adverse effects on the rotifers. Dosing rotifers with respective algal cell densities of 2000 cells ml⁻¹ each, for Alexandrium sp1, Alexandrium sp2, and A. tamarense strains ATHK and ATCI03 showed mean lethal time (LT₅₀) on rotifer populations of 21, 28, 29, and 36h, respectively. The remaining three species (A. tamarense strains AT5-1, AT5-3, ATCI02) caused respective mean rotifer LT₅₀s of 56, 56, and 71 h, compared to 160 h for the unexposed “starved control” rotifers. Experiments to determine ingestion rates for the rotifers, based on changes in their Chlorophyll a content, showed that the rotifers could feed on A. lusitanicum, A. minutum and A. tamarense strain AT-6, but could graze to little or no extent upon algal cells of the other seven strains. The effects on rotifers exposed to different cell densities, fractions, and growth phases of A. tamarense algal culture were respectively compared. It was found that only the whole algal cells had lethal effects, with strongest impact being shown by the early exponential growth phase of A. tamarense. The results indicate that some toxic mechanism(s), other than PSP and present in whole algal cells, might be responsible for the adverse effects on the exposed rotifers.     

Intensive rotifer cultures using chemostats

Continuous production of the rotifer Brachionus plicatilis rotundiformis (S-type) in an intensive chemostat culture system has been investigated. The production dynamics of rotifers in relation to different flow rates and feed regimes show that the growth rate and production depends on the type of algal feed and flow rate utilized in the culture system. It was possible to achieve a mean production of up to 318.84 × 10⁶ rotifers m^–3 d^–1 at a flow rate of 6 1 h^–1 in 100 1 chemostats and up to 261.21 × 10⁶ rotifers m^–3 d^–1 at a flow rate of 40 1 h –1 while using 1 m³ capacity rotifer chemostats as production units. The 3 fatty acid composition of rotifers while using Chlorella and Nannochloropsis in the culture system has been described. The results of this investigation show that the rotifer productivity in the continuous culture system is considerably higher than in any of the conventional culture systems described to date for aquacultural purposes.This research was financed by the Kuwait Foundation for the Advancement of Sciences (KFAS), Kuwait, under a contract research project code 86-04-02.     

Production and nutritional quality of the rotifer Brachionus plicatilis fed marine Chlorella sp. at different cell densities

The effect of different cell densities of marine Chlorella sp. on the growth rate, doubling time and production of the rotifer Brachionus plicatilis was investigated. A significant increase in rotifer production was achieved at a density of 50 × 10⁶ Chlorella cells ml^–1. The nutritional quality of rotifers grown at different concentrations of Chlorella is discussed.     

Host-handling behavior of the parasitoid,Diglyphus minoeus [Hym.: Eulophidae], parasitizing the honeysuckle leaf-miner,Phytomyza lonicerae [Dipt.: Agromyzidae]

The host-handling behavior of individual female waspsDiglyphys minoeus which attack the larvae of the honeysuckle leaf-miner,Phytomyza lonicerae, include 4 types of behavior: probing, ovipositor insertion, host-feeding, and resting. Ovipositor insertion may either be injection of venom, probing of the host by the sensilla on the ovipositor's tip or egg laying. Three types of attacks were distinguished: oviposition attack for unparasitized larvae, host-feeding attack for unparasitized host larvae and host rejection for previously parasitized host larvae. Oviposition attack was characterized by frequent alternation between probing and ovipositor insertion, long duration of ovipositor insertion and resting, and the long duration of host-handling. Resting behavior is thought to protect the progeny against superparasitism, host-feeding by other wasps or hosts' recovery from paralysis. Host-feeding attack was characterized by frequent alternation between host-feeding and ovipositor insertion and long duration of host-feeding. Host rejection was composed mainly of probing and ovipositor insertion and short handling time.      

In situ metabolic budget for the calanoid copepod Acartia clausi in a tropical brackish water lagoon (Ebrié Lagoon,Ivory Coast)

Simultaneous measurements of respiration, excretion and production rates were carried out several times over a year period at five representative stations of the Ebrié Lagoon. Assuming a constant assimilation efficiency rate of 69.4%, we derived metabolic budgets for carbon, nitrogen and phosphorus. Daily specific ingestion rates calculated were rather generally high, and ranged between 54 and 159% of body carbon, between 26 and 102% of body nitrogen and between 108 and 307% of body phosphorus. Regional and seasonal variations depended mainly upon variations in trophic conditions. Curvilinear relationships between ingestion production, or net production efficiency K2, and food concentration (as chlorophyll-a + phaeopigments) showed that food could have been a limiting factor. Furthermore, K2 were low when compared with data from the literature (mean of 21% in carbon, 39% in nitrogen and 11% in phosphorus).Complementary laboratory experiments carried out on adults fed with enriched natural particles or algal cultures (Tetraselmis sp. or Dunaliella sp.) showed similar production (egg-production) vs food concentration curvilinear relationships as in the field. However, considerably higher maximal ingestion and production rates were obtained for animals fed algal cultures suggesting that optima for food acquisition and transformation were not reached in field conditions.Consequently, A. clausi, which represents more than 50% of the zooplankton biomass, appears to be rather inefficient in transforming the abundant particulate organic matter produced in the lagoon. This results from its high level of metabolic expenditure through respiration or excretion (about 50% of ingestion in terms of carbon) and from the small size and poor trophic value of food particles (high percentage of detritus).     

Susceptibility of Ephemeral Pool <Emphasis Type="Italic">Hexarthra</Emphasis> to Predation by the Fairy Shrimp <Emphasis Type="Italic">Branchinecta mackini</Emphasis>: can Predation Drive Local Extinction?

Isolated rock pools in the US desert southwest often develop dense populations of a lithophilic species of the rotifer Hexarthra. I hypothesized that rotifers persist in these isolated ponds due to the absence of either competition with or predation by potentially sympatric crustaceans, especially anostracans. I tested the latter idea with laboratory and field experiments, in each case exposing rotifers to adult fairy shrimp (Branchinecta mackini, the dominant anostracan in this region in winter ponds) in 200 ml microcosms. In␣most cases individual female fairy shrimp had distinct negative effects on rotifer suspensions due to direct predatory consumption of the smaller animals. Estimated effective water processing rates ranged from ca. 50 to over 300 ml ind⁻¹ h⁻¹ and rotifer consumption rates by female B. mackini were between 280 and > 600 Hexarthra consumed per individual predator per hour. Male B. mackini never significantly reduced rotifer numbers in either laboratory or field microcosms. The results indicate that, while perhaps not the sole determinant of rotifer distribution in these ephemeral pools, fairy shrimp predation can have a strong negative influence on natural Hexarthra populations.     

Optimal diet theory: behavior of a starved predatory snail

Summary The tenets of optimal foraging theory are used to contrast the behavior of the predatory snail Acantina spirata when feeding on the barnacles Balanus glandula and Chthamalus fissus under conditions of satiation and starvation. As predicted in optimal diet models, A. spirata is less selective (ratio of attack frequency on a prey species to number of individuals available) when the higher ranking prey has low abundance. When given a choice, starved snails attack both barnacle species equally, whereas satiated individuals preferentially attack B. glandula, the more profitable prey (ash-free dry weight of barnacles ingested per unit handling time). Under starvation conditions, equal attack frequency does not result in equal prey species consumption because Acanthina spirata is more successful at attacking C. fissus than B. glandula.The assumption of constant prey encounter rates in optimal diet models is not met when A. spirata goes from a state of satiation to starvation. The encounter rate on B. glandula is lowered due to a decrease in attack success. A loss of feeding skills in starved A. spirata is responsible for the greater difficulty snails have in gaining access through the opercular plates of B. glandula.Behavioral changes in A. spirata as snails pass from satiation to hunger translate into an energetic disadvantage during feeding for hungry snails for two reasons. First, higher prey handling times result in a decreased rate of biomass intake. Second, alteration in the relative attack frequency between barnacle species, combined with a decrease in attack success on the more profitable prey leads to more frequent ingestion of the less profitable prey.     

Selective feeding of Arctodiaptomus salinus (Copepoda, Calanoida) on co-occurring sibling rotifer species

1. Using two‐ and three‐dimensional video recordings, we examined the steps involved in predation that lead to the differential vulnerability of three sympatric rotifer sibling species (Brachionus plicatilis, B. ibericus and B. rotundiformis) to a co‐occurring, predatory, calanoid copepod (Arctodiaptomus salinus). 2. Brachionus rotundiformis, the smallest prey tested, was the most vulnerable with the highest encounter rate, probability of attack, capture and ingestion, and the lowest handling time. 3. Comparison of our results with those of a previous study shows that A. salinus is a more efficient predator than a co‐occurring cyclopoid copepod (Diacyclops bicuspidatus odessanus) feeding on these same rotifer species. However, despite its higher capture rates, A. salinus seems to be less selective than D. b. odessanus based on attack distances and prey handling times. 4. The differential vulnerability to both calanoid and cyclopoid copepod predation can help explain the coexistence and seasonal succession of these co‐occurring rotifer species.     

Leafhoppers on leaves: An analysis of feeding behavior using conditional probabilities

Feeding behavior of four deltocephaline leafhoppers,Graminella nigrifrons, G. oquaka, Amblysellus grex, andDalbulus maidis on maize and johnsongrass was analyzed using an electronic monitoring device. Five distinct waveform patterns were identified: secretion of sheath saliva (salivation), nonvascular probing, nonsieve element ingestion, x-waveform, and phloem ingestion. Waveforms were associated with feeding activities by correlation with light microscopic examination of salivary sheath termination points in leaf tissue and analysis of honeydew excreted by monitored leafhoppers. In previous studies x-waveforms have been reported to occur only when the stylets of homopterans are in contact with the phloem; the function of x-waveforms is poorly understood. There were no differences in time spent salivating or ingesting from nonsieve elements amongG. nigrifrons, G. oquaka andA. grex on either plant.D. maidis differed from other species in phloem probing and feeding behavior; only a small proportion produced x-waveforms, although those that did spent significantly more time in this behavior than other species. Also,D. maidis spent more time than other leafhoppers ingesting from tissues other than sieve elements. Kinetic diagrams of transition probabilities show that probing activities of all species were not random regarding the sequence of behaviors culminating in phloem ingestion. Thirty-five percent ofG. nigrifrons x-waveforms were followed by nonsieve element ingestion. This was consistent with observations showing that salivary sheaths of leafhoppers producing x-waveforms sometimes do not terminate in the phloem, but rather in nearby cells. Phloem ingestion was always preceeded by x-waveforms. The quantitative differences in probing behavior are discussed in relation to ability of these leafhoppers to transmit the phloem-associated maize chlorotic dwarf waikavirus.     

Foraging behavior by <Emphasis Type="Italic">Daphnia</Emphasis> in stoichiometric gradients of food quality

Mismatches in the elemental composition of herbivores and their resources can impact herbivore growth and reproduction. In aquatic systems, the ratio of elements, such as C, P, and N, is used to characterize the food quality of algal prey. For example, large increases in the C:P ratio of edible algae can decrease rates of growth and reproduction in Daphnia. Current theory emphasizes that Daphnia utilize only assimilation and respiration processes to maintain an optimal elemental composition, yet studies of terrestrial herbivores implicate behavioral processes in coping with local variation in food quality. We tested the ability of juvenile and adult Daphnia to locate regions of high-quality food within a spatial gradient of algal prey differing in C:P ratio, while holding food density constant over space. Both juveniles and adults demonstrated similar behavior by quickly locating (i.e., <10 min) the region of high food quality. Foraging paths were centred on regions of high food quality and these differed significantly from paths of individuals exposed to a homogeneous environment of both food density and food quality. Ingestion rate experiments on algal prey of differing stoichiometric ratio show that individuals can adjust their intake rate over fast behavioral time-scales, and we use these data to examine how individuals choose foraging locations when presented with a spatial gradient that trades off food quality and food quantity. Daphnia reared under low food quality conditions chose to forage in regions of high food quality even though they could attain the same C ingestion rate elsewhere along a spatial gradient. We argue that these aspects of foraging behavior by Daphnia have important implications for how these herbivores manage their elemental composition and our understanding of the dynamics of these herbivore–plant systems in lakes and ponds where spatial variation in food quality is present. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Effect of temperature on the filtration rate and percentage of assimilation ofSalpa fusiformis Cuvier (Tunicata: Thaliacea)

The effect of temperature on filtration and ingestion rates ofSalpa fusiformis Cuvier was determined over the temperature range 13–22°C at a constant algal concentration. Filtration and ingestion rates increased with temperature up to an optimum (19.7 to 20.2°C) beyond which they decreased. Food assimilation for this salp species was determined while fed onPhaeodactylum tricornutum Bohlin orHymenomonas elongata Stein at different algal concentrations. The percent assimilation varied between 28 and 39% (mean: 32%) with the diatomP. tricornutum as food, and between 39 to 81% (mean: 64%) with the flagellateH. elongata. Assimilation did not seem to depend on the algal concentration but it was greater with the flagellate than with the diatom, the latter having a higher ash content.     

A laboratory study of phosphorus and nitrogen excretion of Euchlanis dilatata lucksiana

Phosphorus (PO₄-P) and nitrogen (NH₄-N) excretion rates of Euchlanis dilatata lucksiana, a rotifer, isolated from Lake Loosdrecht (The Netherlands) and cultured in the lake water at 18–19 °C, were measured in the laboratory.In a series of experiments, the effects of experiment duration on the P and N excretion rates were examined. The rates measured in the first half-hour were about 2 times higher for P and 2–4 times for N than the rates in the subsequent three successive hours which were quite comparable.Eight experiments were carried out in triplicate, 4 each for P and N excretion measurements, using animals of two size ranges: 60–125 µm and > 125 µm. The specific excretion rates varied from 0.06 to 0.18 µg P.mg^–1 DW.h^–1 and 0.21 to 0.76 µg N.mg^–1 DW.h^–1. Generally an inverse relationship was observed between the specific excretion rates and the mean individual weight. The excretion rates of Euchlanis measured by us are lower than those reported for several other rotifer species, most of which are much smaller than Euchlanis.Extrapolating the excretion rates of Euchlanis to the other rotifer species in Lake Loosdrecht, and accounting for their density, size and temperature, rotifer excretion appears to be a significant, potential nutrient (N,P) source for phytoplankton growth in the lake. The excretion rates for the rotifers appear to be about two thirds of the total zooplankton excretion, even though the computed rotifer mean biomass is about one-third of the total zooplankton biomass.     

Interactions between red tide microalgae and herbivorous zooplankton: effects of two bloom-forming species on the rotifer <Emphasis Type="Italic">Brachionus plicatilis</Emphasis> (O.F. Muller)

Experiments were carried out to investigate interspecific interactions between the rotifer Brachionus plicatilis and two harmful algal bloom (HAB) species using single and mixed culture methods. B. plicatilis populations and the growth of two algae were compared at different algal cell densities. The results demonstrate that B. plicatilis obtained sufficient nutrition from Alexandrium tamarense to support net population increase. When exposed to a density of 8 × 10⁴ cells ml⁻¹ A. tamarense, the number of B. plicatilis increased faster than it did when exposed to other four algal densities (16 × 10⁴, 24 × 10⁴, 32 × 10⁴, and 40 × 10⁴ cells ml⁻¹). Cell densities of A. tamarense decreased due to the grazing of B. plicatilis. In contrast, Heterosigma akashiwo had an adverse effect on the B. plicatilis population and its growth was largely unaffected by rotifer grazing. In this case, the B. plicatilis population decreased and H. akashiwo grew at a rate similar to that of a control without addition of rotifers. Mixed culture experiments showed that A. tamarense could partly counteract the effect of H. akashiwo in limiting the rate of population increase of rotifer. In addition, the effect of different initial cell densities on interspecific competition between A. tamarense and H. akashiwo in mixed culture(s) was also investigated. The results show that A. tamarense competed very successfully when the inoculation proportions of A. tamarense and H. akashiwo were 40:5 and 40:30. Handling editor: D. Hamilton     

Utilization of cyanobacteria by Brachionus calyciflorus: Anabaena flos-aquae (NRC-44-1) as a sole or complementary food source

The rotifer Brachionus calyciflorus can utilize the cyanobacterium Anabaena flos-aquae as either a sole or supplementary food source in laboratory culture. Positive population growth rates accompany food densities of 10 or 100 µg dry weight ml^–1, but slightly negative rates are found at a lower density (1.0 µg ml^–1). These results are consistent for rotifers feeding on two strains of A. flos-aquae, UTEX-1444 and NRC-44-1, with slightly enhanced survivorship and reproduction with the latter food. A 1:1 mixture (by dry weight) of Euglena gracilis and A. flos-aquae (NRC-44-1) produces survivorship comparable to that of control rotifer cohorts fed E. gracilis alone, but elicits significantly greater fecundity and population growth rates than found with the control food suspension at the same biomass density.     

Geocoris punctipes as a predator of Bemisia tabaci: a laboratory evaluation

Geocoris punctipes (Say), a predaceous lygaeid not previously docmented as a whitefly predator, was tested in the laboratory as a natural enemy of the sweet potato whitefly, Bemisia tabaci (Genn.). Its stalking behavior with whiteflies as prey was similar to that observed with aphids. A previously unobserved behavior was noted that involves the predators using salivary secretions to fasten the wings of prey to various surfaces, allowing labial probing and feeding. Prey consumption as a function of prey number appeared to follow the pattern of the Holling type II functional response. Handling time per prey item ranged from about 180 to 240 seconds. No changes were observed in handling time devoted to earlier versus later catches. Nutritional quality of whiteflies was measured using crude protein, lipids and carbohydrates as criteria. Performance, in terms of predator behavior, total daily handling time, functional response, energy budget and nutritional quality all support the hypothesis that G. punctipes is a promising candidate for biological control of sweet potato whiteflies.     

Microbial decomposition of urea in the Menai Straits

The assimilation of urea by the microbial population of the Menai Strait (Anglesey, U.K.) was studied over a seasonal cycle. Samples incubated in the light produced higher assimilation rates than samples incubated in darkness. Higher ratios assimilation of urea: chlorophyll a during a time of a probable nitrogen limiting condition may be indicative of the importance of urea for phytoplankton nutrition. Assimilation rates of urea were higher during the diatom and Phaeocystis bloom than in times of low standing crop.Phytoplankton seem to be the major utilizers of urea in the Menai Strait waters. Bacteria may also be important in the decomposition of urea. However, with the methods employed it was not possible to estimate the extent of their participation in the urea decomposition.     

Review of rotifers and crustaceans in highly acidic environments of pH values ≤3

A review of the literature on rotifers and crustacean zooplankton in highly acidic environments revealed that data from eleven aquatic environments on three continents (America, Europe, Japan) with a pH 3 are available. Seven sites are influenced by volcanism or weathering processes in the catchment area, four others originated from human mining activities. Species richness was generally low. Only 16 species are found and 1–11 species are reported for each area. These studies clearly show that small littoral or benthic rotifers predominate over crustaceans under highly acidic conditions. In the Lusatian mining area (Germany), all lakes are colonized by zooplankton, even the most acidic one with a pH of 2.3. The core community consists of the rotifers Cephalodella hoodi, C. gibba, Elosa worallii and Rotaria rotatoria, with C. hoodi and E. worallii the most abundant. Larger species, such as the rotifer Brachionus sericus or the cladoceran Chydorus sphaericus, occur at a pH close to 3. A similar pattern is reported from acidic mining lakes in Illinois, U.S.A. Many of these species can also be found in less acidic softwater or even alkaline environments due to the tolerance of a broad range of pH values. Elosa worallii and Brachionus sericus are probably the most acidophilic rotifer species, though at least the latter can also grow at neutral pH in the laboratory. Clear understanding of the pH limits of B. sericus in nature may also have been complicated by the fact that it has probably in the past been wrongly named as B. urceolaris (phenotype `sericus'). The typical B. urceolaris cannot tolerate extremely low pH. Overall, generalist species with a worldwide distribution seem to play the major role in the colonization of anthropogenic highly acidic lakes.