Use of immobilized algae for estimating bioavailable phosphorus released by zooplankton

Cells of the green alga Selenastrum capricomuxum were immobilizedin permeable alginate beads to prevent them from being grazedby zooplankton. Algae were able to grow in these beads and wereused as a new technique to estimate bioavailable phosphorus(P) released by zooplankton. P-limited algal cells were encapsulatedin alginate beads and used to measure P-release by Daphnia pulexfeeding on P-saturated and P-limited free algal cells. Daphnidsgrazing on P-saturated cells released 20 times more P availablefor the immobilized algae than animals grazing on P-limitedcells (0.06 versus 0.003 µg P mg^–1 Daphnia-DW h^–1).     

Feeding rate of Diaptomus sicilis and its relation to selectivity and effective food concentration in algal mixtures and in Lake Michigan

The concept of effective food concentration (EFC), a means ofpredicting food consumption from selectivity and food concentrationdata, is explained, tested, and applied to understanding foodconsumption by the freshwater copepod Diaptomus sicilis on mixturesof algae of different sizes and on Lake Michigan seston. Experimentson mixtures of different sized Chlamydomonas spp. showed thatselection (W') was an invariant function of particle size whenthe algae were counted microscopically. When the Coulter counterwas used, a more variable pattern of selectivity — similarto the peak tracking response reported by some investigators- was obtained. This was due to bias of zooplankton-producedparticles. Size-selective selectivity coefficients (W') wereused to weight the food concentration in each size categoryand the weighted values summed to give EFC. Food consumptionin experiments with seston and with cultured algae was betterdescribed by EFC than by total food concentration (TFC), theunweighted sum. Moreover, use of EFC diminished the magnitudeof the apparent threshold concentration required for feedingto commence. Although selectivity in algal mixtures and lakeseston was approximately the same, the food consumption versusEFC curve saturated more quickly for the algal mixtures thanfor the lake seston. Since expression of food concentrationas EFC allowed direct comparison of experiments having differentparticle-size spectra of food, we concluded the difference resultedfrom the lower food quality of lake seston, that is, its lowerdigestibility and sensory quality for zooplankton capture. ¹GLERL Contribution No. 368 ²Also Division of Biological Sciences, University of Michigan,Michigan, USA     

Post-embryonic developmental rates as a function of food type in the cyclopoid copepod, Mesocyclops thermocyclopoides Harada

We studied under laboratory conditions the post-embryonic developmentalrates (from nauplii to adult stage) of Mesocyclops thermocyclopoides,commonly found in the eutrophic and hypertrophic local ponds,in relation to different food types. Nauplii hatched from theeggs collected from laboratory-maintained mass cultures werereared at 25l.5C, using the following test diets: bacteria(Klebsiella aerogenes), cyanobacteria (Microcystis aeruginosa,as single cells and in the form of colonies), green algae (Chlorella,Scenedesmus and Chlorogonium), ciliates (Tetrahymena) and rotifers(Brachionus angularis). The developmental rates were fastest,and the proportion of nauplii reaching the copepodid stage andthat of copepodids reaching the adult stage the highest, withChlorogonium, followed by ciliates and rotifers. Developmentwas incomplete with bacteria and Microcys-tis.Ttie algae Chlorellaand Scenedesmus supported complete post-embryonic developmentonly when cultured in media enriched with beef extract, bactotryptoneand yeast extract. With rotifers as the exclusive food, >70%of the nauplii reached the copepodid stage, and >85% of thecopepodids the adult stage. The ratio (D_c/D_n) of copepodid duration(D_c) and naupliar duration (D_n) was significantly differentfrom 1.0, indicating the absence of isochronal development inM.thermocyclopoides. Our results show that post-embryonic developmentin this species is not possible with certain algal diets, probablybecause of nutritional deficiencies, and that the later stagenauplii are capable of capturing rotifers and utilizing themas food to complete their development to the adult stage.     

Feeding of nauplius stages of Eudiaptomus gracilis on mixed plastic beads

Eudiaptomus gracilis makes up 30–40 and 80–90% ofthe zooplankton in Lake Balaton during the summer and the winterrespectively. More than half of the species population consistsof nauplii We studied feeding and size selectivity of naupliiin suspensions which contained polystyrol latex beads in a concentrationdose to the natural seston. Guts of NI nauplii were free ofboth beads and remnants of natural food Of NII–NVI nauplii,67–87% took in beads. Older animals consumed more andlarger particles. The maximum diameter of ingested beads reached29 µm On an average, NII nauplii took in 128 µm³of beads in 10 min, whereas older animals consumed 615–900µm³. The clearance rate remained below 0 01 µl h^–1NII nauplii strongly preferred 1 2 µm particles Oldernauplii did not show any preference or selected only slightlyfor the smallest particles. Nauplii rejected 4–11 µmbeads. In some cases a weak positive selection could be observedtoward 12 µm or larger beads.     

Remote detection of algae by copepods: responses to algal size, odors and motility

Selective feeding on large algae by copepods involves remotedetection of individual particles and subsequent active captureresponses In this study we use radiotracer experiments to quantifythe clearance rates of five coexisting freshwater copepods andto investigate the relative merits of the chemoreception andmechanoreception hypotheses of remote detection Tropocyclopsand three diaptomid copepods exhibited relatively high clearancerates when feeding on low concentrations of large algae, suggestingthat most previous studies with freshwater copepods have underestimatedmaximal clearance rates and the degree of size selectivity.All five species of copepods exihibited strong selection foran intermediate-sized flagellate (25 µm Cartena) or alarge-sized nonmotile alga (80 µ.m Pediastrum) over asmall-sized flagellate (6 µ.m Chlamydomonas). The weight-specificclearance rate for Tropocylops prasmus feeding on motile Cartena(271 ml mg^–1 h^–1) was about twice that of threediaptomid copepods and more than an order-of-magnitude higherthan the estimate for Epischura lacustris feeding on its preferredalga, Pediastrum Assuming that distance chemoreception (‘smell’)is important in remote detection, we predicted that the additionof high concentrations of ‘algal odors’ would obscureany chemical gradients emanating from individual algal cellsand would thereby hinder the remote detection and active captureof large algae Contrary to this hypothesis, the addition ofamino acids, sucrose, and algal extracts had no effect on theclearance rates and selectivity of Diaptomus birgei. These results,together with recent cinematographic studies (Vanderploeg etal.,1990), suggest that mechanoreception is the primary mechanismfor the remote detection of large particles by diaptomid copepods.A raptorial cyclopoid, Tropocyclops prasinus, exhibited strongpreferences for motile algae, whereas a suspension-feeding calanoid,D birgei, did not select between motile and nonmotile cells.Motility appears to be an important factor in algal detectionfor small cyclopoid copepods but not for suspension-feedingdiaptomids     

Acanthamoeba discriminates internally between digestible and indigestible particles

The capacity of Acanthamoeba to distinguish nutritive yeast particles from non-nutritive plastic beads during phagocytosis was investigated. When cells were allowed to phagocytose yeast to capacity, endocytosis stopped and subsequent presentation of particles (either yeast or beads) did not result in further uptake. By contrast, when cells were allowed to phagocytose plastic beads to capacity and a second dose of particles was presented (either yeast or beads), the cells exocytosed the internal particles and took up new ones. Yeast rendered indigestible by extensive chemical cross-linking were taken up at rates similar to those of untreated yeast, but, like beads, they were exocytosed when a second dose of particles was presented. The results show that an internal distinction is made between vacuoles containing yeast and vacuoles containing plastic beads, and they are consistent with the hypothesis that the presence within the vacuoles of material capable of being digested prevents exocytosis.     

Feeding rates and selectivity among nauplii, copepodites and adult females of Calanus finmarchicus and Calanus helgolandicus

This study focuses on selective feeding by developmental stages of two oceanic copepods, Calanus finmarchicus and Calanus helgolandicus from nauplii to adults. A mixture of four algal species of different biochemical composition, Prorocentrum nanum (dinoflagellate), Thalassiosira minima (diatom), Rhodomonas baltica (cryptophyte) and Dunaliella tertiolecta (chlorophyte), added in an equal biovolume, was used in three different experimental set-ups. In set-up 1 the algal species were present as single cells of similar size (14 μm). In set-up 2 the diatom T. minima was present in chains of two or three cells and was therefore larger than the other algae, while the biovolume of all species remained the same. In set-up 3, the diatom T. minima was excluded from the mixture. Feeding selectivity of the copepods was assessed in relation to the quality of the algal species expressed in terms of carbon and nitrogen content, fatty acid composition, and chain length of the diatom. The results show that younger stages and adult females of C. finmarchicus and C. helgolandicus did not show a preference for an algal species when the algae were of similar size. In the feeding experiments where the diatoms were offered as chains, both copepod species showed a selective behaviour only on the basis of algal size. Individual ingestion rates increased from 0.4 to 0.7 μg C day^–1 for nauplii of both species to 5 μg C day^–1 for adult females of C. helgolandicus to 12 μg C day^–1 for C. finmarchicus. Individual filtration rates ranged from 5 ml day^–1 for C. finmarchicus nauplii to 70–98 ml day^–1 for adult females, and from 3 ml day^–1 for C. helgolandicus nauplii to 35–46 ml day^–1 for adult females. Ingestion and filtration rates per unit body carbon decreased gradually in both copepod species with increasing body carbon. The daily ingested amount of food decreased for C. finmarchicus from 124–134% of the body carbon for nauplii to 19% of the body carbon for adult females, and for C. helgolandicus from 117–137% of the body carbon for nauplii to 13–26% of the body carbon of adult females. Electronic Publication     

The influence of food resources on the development, survival and reproduction of the two cyclopoid copepods: Cyclops vicinus and Mesocyclops leuckarti

Laboratory experiments were conducted to determine whether thetwo cyclopoid copepods. Cyclops vicious and Mesocyclops leuckaru.exploit the same food resources. The food requirements of juvenilesof the two cyclopoid copepods were investigated. Moreover, theimportance of algae for the predaceous adults was studied. Naupliiof both M leuckaru and C.vicinus successfully developed intocopepodites when fed the motile algae Chlamydomonas reinhardtii.Chlamydomonas sphaeroides and Cryptomonas sp. Threshold foodconcentrations for naupliar development varied between offeredalgae and between the two cyclopoid species. The food thresholdfor successful naupliar development, when reared on C.reinhardui,was lower for M.leuckarti (0.3 mg C 1^–1) than for C.vicinus(0.5 mg C l^–1) whereas a similar food threshold was foundusing Cryptornonas sp (0.3 mg C ^–1) and C.sphaeroides(<0.2 mg C 1^–1), Naupliar development time was inverselyrelated to food concentration. Food required for copepoditedevelopment differed for the two cyclopoid species. Cyclopsvicinus was able to develop to the adult stage on a pure dietof any one of the three algal species. whereas M.leuckarti requireda prey supply of the rotifer Brachionus rubens. Food composition.i.e. algal species, algal concentration and rotifer abundance,influenced copepodite survivorship of both cyclopoids and wasalways higher in the presence of B.rubens. Under similar foodconditions, mortality was higher for M.leuckarti than for Cvicinus. Mesocyclops leuckaru females were very dependent onanimal food. The predation rate of M.leuckaru was not lowerin the presence of algae. Egg production of M.leuckarti waslow on a pure algal diet and significantly higher when B rubenswas present. The results were used to discuss the life cyclestrategy and the possibility of exploitative competition ofthe two cyclopoid copepods.     

Influence of two Daphnia species on summer phytoplankton assemblages from eutrophic lakes

We conducted grazing experiments to test whether larger-bodiedDaphnia pulicaria have a different effect from smaller-bodiedDaphnia galeata mendotae on the composition of summer algalassemblages in eutrophic lakes. Three separate cubitainer experimentswere run for 5 days in a replicated factorial design utilizingtwo algal community types and the two Daphnia species. Inorganicphosphorus and nitrogen were added to prevent nutrient limitationof the algae. Both edible and inedible size fractions of chlorophylla increased in cubitainers without Daphnia spp. Grazer additionusually resulted in a reduction in edible chlorophyll; reductionswere greater in D.pulicaria cubitainers. Grazing by Daphniaspp. on presumed inedible chlorophyll was variable. Algal sizewas not always a good predictor of grazeability. The resultsof this study indicate that D.pulicaria, because of its greaterfiltration potential and ability to ingest larger particles,provides a stronger control on inedible-sized algae when comparedto equal numerical densities of D.g.mendotae. However, Aphanizomenonincreased as a response to heavy grazing pressure by D.pulicariaon other algal species. This suggests that biomanipulation effortsthat promote large-bodied Daphnia may not produce desirableresults if nutrient inputs remain high.     

Endosymbiosis in Hydra and the Evolution of Internal Defense System

Certain species of Chlorella have exploited an intracellularhabitat and occur naturally as cytobionts in Hydra viridissima.The algae evoke phagocytosis by Hydra digestive cells and aresequestered in individual phagosomes that migrate to the baseof the host cells and resist fusion with lysosomes. The abilityto resist digestion is closely correlated with release of extracellularcarbohydrate (maltose) by the algae. The established populationof algae grows at an average rate equal to or greater than thatof the host and a constant population density is maintained.The host regulates algal population density by expelling ordigesting excess algae, or by controlling algal cell division.The control mechanism is unknown but can be breached by additionof inorganic ions to the Hydra culture medium with the resultthat the algae overgrow the Hydra. The Hydra-Chlorella symbiosis is probably mutually beneficial,but conditions such as prolonged darkness (with or without feeding)can reduce the competitive fitness of the host since this conditionresults in heterotrophy by the algae at the expense of selectedhost substrates. The evolution of selective permeability toorganic substrates is a major feature of the successful colonizationof the intracellular habitat by symbiotic Chlorella.     

Particle capture by Favella sp. (Ciliata, Tintinnina)

Particle capture by the tintinnid ciliate, Favella sp., wasinvestigated with high-speed video microscopy. Experiments withmicrospheres and algal cells indicate that within the size range1–19 p.m. behavioral responses are partly responsiblefor the differential capture of large over small particles.For example, 4 µm microspheres or Isochrysis [spheiicaldiameter (ESD) - 4.5 µ are only captured if they are encounteredin the inner zones of the encounter area. Ten micrometer microspheresor Heterocapsa(14–19 – are captured with a greaterefficiency than smaller particles from the inner zones, butcan also be captured from the outer zone of the encounter area.Capture of particles from the outer zone is associated withcilialy reversal. Microsp1ieres and algal cells of similar sizeare captured in a similar manner.     

Laboratory studies on the larval growth and development of Nyctiphanes capensis (Euphausiacea)

Larvae of Nyctiphanes capensis Hansen were reared in the laboratoryunder different trophic conditions (i.e., algal, animal andmixed diets) from the second calyptopis stage to the first juvenilestage. It is demonstrated that diet is important in determininggrowth rates and development of the larvae. The highest growthrates were achieved on diets of Artemia nauplii mixed with eitherthe flagellate Tetraselmis or Pseudodiaptomus nauplii. Poorerdietary conditions such as with the diatom Phaeodactylum yieldedslower growth rates where additional moults were needed to completefurcilia development. Under favourable trophic conditions theintermoult period was 4–6 days and was not age dependent.Three dominant pathways of pleopod development were found inhealthy individuals.     

Effects of concentration on the feeding of a marine copepod in algal monocultures and mixtures

High-speed microcinematography was used to examine the feedingbehavior of the marine copepod Eucalanus elongatus in a rangeof concentrations of algal monocultures and mixtures. Two celltypes were offered, the 13-µm diatom Thalassiosira weissflogii,which is primarily accumulated passively by low amplitude flappingof the second maxillae, and the 20 450-µm diatom Rhizosoleniaalata, which is actively captured by detection of and orientedresponse to individual cells. E. elongatus rapidly switchedback and forth between these two capture modes in mixtures ofboth diatoms, and flapped the second maxillae at low amplitudesregardless of the absolute or relative abundance of small andlarge cells. However, copepods in both monocultures and mixturesaltered the duration and/or rate of flapping of the feedingappendages with changes in algal concentration, with maximumactivity levels occurring at intermediate concentrations. Themarked reduction in feeding motions observed at the lowest algalconcentrations supports results from traditional grazing studiesand optimal foraging models, and may conserve energy duringprolonged perods of low food availability in continental slopewaters.     

Linking herbivore-induced defences to population dynamics

1. Theoretical studies have shown that inducible defences have the potential to affect population stability and persistence in bi‐ and tritrophic food chains. Experimental studies on such effects of prey defence strategies on the dynamics of predator–prey systems are still rare. We performed replicated population dynamics experiments using the herbivorous rotifer Brachionus calyciflorus and four strains of closely related algae that show different defence responses to this herbivore. 2. We observed herbivore populations to fluctuate at a higher frequency when feeding on small undefended algae. During these fluctuations minimum rotifer densities remained sufficiently high to ensure population persistence in all the replicates. The initial growth of rotifer populations in this treatment coincided with a sharp drop in algal density. Such a suppression of algae by herbivores was not observed in the other treatments, where algae were larger due to induced or permanent defences. In these treatments we observed rotifer population densities to first rise and then decline. The herbivore went extinct in all replicates with large permanently defended algae. The frequency of herbivore extinctions was intermediate when algae had inducible defences. 3. A variety of alternative mechanisms could explain differential herbivore persistence in the different defence treatments. Our analysis showed the density and fraction of highly edible algal particles to better explain herbivore persistence and extinctions than total algal density, the fraction of highly inedible food particles or the accumulation of herbivore waste products or autotoxins. 4. We argue that the rotifers require a minimum fraction and density of edible food particles for maintenance and reproduction. We conjecture that induced defences in algae may thus favour larger zooplankton species such as Daphnia spp. that are less sensitive to shifts in their food size spectrum, relative to smaller zooplankton species, such as rotifers and in this way contributes to the structuring of planktonic communities.     

Resistance to blue-green algal toxins by Bosmina longirostris

Bosmina longirostris was resistant to strains of Microcysrtsaeruginosa and Anabaena flosaquae previously reported to havelethal toxic effects on cladocerans. These blue-green algaewere of poor nutritional value to B.longirostris; survivorshipwas increased when fed these algae but reproduction was negligible.Results of feeding selectivity experiments showed that B.longirostrisdid not avoid consuming these blue-green algae, indicating thatthe mechanism for resistance must be post-ingestion. These resultssuggest that, unlike the other cladocerans tested, B.longirostriscould potentially coexist with toxic blue-green algal blooms.     

Experimental identification and in silico prediction of bacterivory in green algae

While algal phago-mixotrophs play a major role in aquatic microbial food webs, their diversity remains poorly understood. Recent studies have indicated several species of prasinophytes, early diverging green algae, to be able to consume bacteria for nutrition. To further explore the occurrence of phago-mixotrophy in green algae, we conducted feeding experiments with live fluorescently labeled bacteria stained with CellTracker Green CMFDA, heat-killed bacteria stained with 5-(4,6-dichlorotriazin-2-yl) aminofluorescein (DTAF), and magnetic beads. Feeding was detected via microscopy and/or flow cytometry in five strains of prasinophytes when provided with live bacteria: Pterosperma cristatum NIES626, Pyramimonas parkeae CCMP726, Pyramimonas parkeae NIES254, Nephroselmis pyriformis RCC618, and Dolichomastix tenuilepis CCMP3274. No feeding was detected when heat-killed bacteria or magnetic beads were provided, suggesting a strong preference for live prey in the strains tested. In parallel to experimental assays, green algal bacterivory was investigated using a gene-based prediction model. The predictions agreed with the experimental results and suggested bacterivory potential in additional green algae. Our observations underline the likelihood of widespread occurrence of phago-mixotrophy among green algae, while additionally highlighting potential biases introduced when using prey proxy to evaluate bacterial ingestion by algal cells.Subject terms: Microbial ecology, Cellular microbiology     

In situ egg production and hatching success of the marine copepod Pseudocalanus newmani in Funka Bay and adjacent waters off southwestern Hokkaido, Japan: associated to diatom bloom

. Clutch size and egg viability were examined for wild femalesof Pseudocalanus newmani collected from four different stationsinside and outside Funka Bay, southwestern Hokkaido, Japan,during pre-phytoplankton-bloom conditions in 1997, and froma single station outside the bay during the period from pre-to post-blooming seasons in 1998, to examine spatio-temporalvariability of the reproductive parameters and to detect deleteriouseffects of diatom blooms on in situ copepod reproduction. Clutchsize and hatching success in P.newmani were relatively similarat the four stations in 1997. The eggs almost always hatchedcompletely, but deformed nauplii occurred in 20–40% ofthe cases. The proportion of deformed nauplii gradually decreasedwhen the females were fed on non-diatom Pavlova sp. in the laboratory,suggesting that the occurrence of deformed nauplii was relatedto the quality of in situ food particles, including diatoms.Clutch size varied from 12 to 30 eggs female^–1 duringthe study period in 1998; more than 90% of the variation couldbe explained by the body size of the females which dependedon ambient water temperature. Hatching success also varied,from 15 to 80%, but was not related to either clutch size, femalebody size, water temperature or diatom biomass. These resultssuggest that not only egg production but also hatchability shouldbe measured routinely when estimating recruitment of the copepodinto the planktonic population, and show that neither clutchsize nor egg viability are directly affected by diatom biomass.Although we could not obtain clear evidence of in situ deleteriouseffects of diatoms on abnormal embryos and nauplii, this mightbe related to methodological problems and properties or characteristicsof prey–predator interactions. Also, deformities of copepodnauplii have not yet been observed for reasons other than feedingon diatoms.     

On the Occurrence of Intracellular Blue-green Algae in Cortical Cells of the Apogeotropic Roots of Macrozamia communis L. Johnson

The occurrence of species of the cyanophytes Nostoc and Anabaenain the cortex near the algal zone is reported for apogeotropicroots of Macrozamia communis L. Johnson. Algae were found tooccur both intercellularly and intracellularly in cells of theinner and outer cortex. This is the first record of intracellularalgae in the cycads and only the second report of this phenomenonin vascular plants. By examination of cells at various stagesof invasion by algae, it is interpreted that algal invasionof cortical cells and intercellular spaces is preceded by mucusapparently secreted by algal zone cells of the host, and depositedin intercellular spaces of cortical parenchyma cells nearby.Also algal penetration of cortical cells is preceded by an algalinvasion front of finely granular mucal material which bypassesmucus already deposited in intercellular spaces and may eitherlyse part of the host cell wall or enter through the plasmo-desmata,filling much of the cell cavity. Subsequently, large numbersof the algal symbionts enter the cell and may be enclosed withinhost wall material. Electron microscopic techniques are nowbeing employed to further clarify these invasion processes.     

Life history consequences for Daphnia pulex feeding on nutrient-limited phytoplankton

1. The growth and feeding of Daphnia pulex De Geer on different algal species was examined. The green algae Chlamydomonas reinhardtii Dangeard and Scenedesmus acutus Meyen, the diatom Synedra tenuissima Kützing, the cryptophyte Cryptomonas pyrenoidifera Geitler and the cyanobacterium Microcystis aeruginosa Kützing were cultured in non-limiting and in N- or P-limiting medium and used as food for D. pulex.
2. Growth limitations were reflected in the elemental and biochemical composition and the morphological characteristics of the algal resources.
3. The clearance rates of D. pulex feeding on nutrient-limited algae were reduced. This was not observed when nutrient-limited mutant Chlamydomonas cells without cell walls were used as food, indicating that the cell wall may play an important part.
4. In comparison with animals grown on nutrient-sufficient cells, nutrient-limited algae resulted in smaller body length, reduced brood sizes, reduced size at maturity, increased age at first reproduction and, consequently, in reduced Daphnia population growth rates.
5. Daphnia population growth rates ( r ) were negatively correlated with the C : P ratio and the carbohydrate content of the food. Moreover, significant correlations between r and clearance rates were found.
6. The observed differences in the grazing and the life history parameters of Daphnia feeding on non-limited and nutrient-limited algae may be the result of both reduced nutritional value and reduced digestibility of nutrient-limited algae.     

In situ measurement of algal growth potential in aquatic ecosystems by immobilized algae

Cells of the green algaSelenastrum capricornutum were immobilized in alginate beads. The alga was able to grow inside these beads without being grazed by zooplankton. For P-limited immobilized cells, however, a lower µ _m and initial slope of the Monod growth curve µ _m /K _s were found than for free cells.To study the feasibility of immobilized algae to estimate algal growth potentialin situ in aquatic ecosystems, a series of experiments were conducted in indoor model ecosystems (microcosms) and in a small stream. The use of immobilized algae allowed a continuous registration of algal growth potential integrated over periods with natural fluctuations in the environment. The method of encapsulation of the algae can, however, still be improved. The alginate matrix is exposed to marked degradation by microorganisms when incubated in polluted streams for a period longer than two weeks. The applicability of other types of matrices should be tested.Author for correspondence