Growth and production of heterotrophic nanoflagellates in a meso-eutrophic lake

The growth of heterotrophic nanoflagellates (HNF) in mesotrophicLake Constance was measured in situ during a 13 month period.Experiments were conducted with 10 µm pre-filtered lakewater incubated in diffusion chambers at 3 m water depth atthe sampling location for 24 h. Growth rates were calculatedfrom changes in cell numbers occurring during the period ofincubation. Growth rates of all dominant taxa showed pronouncedseasonal variation (–0.13 to 1.76 day^–1 and weregenerally highest in summer at high water temperatures. In situgrowth rates were well below maximum growth rates known forthe respective and similar species from laboratory experiments.While water temperature was a key parameter positively relatedto the growth of all HNF species, the effect of various potentialfood items was taxon specific and less clear. Bacterial abundancewas equally important as temperature for growth in the smallbactenvorous Spumella sp., but was insignificant for growthrates of the larger omnivorous Kathablepharis sp. In Spuniellasp., 84% of the observed seasonal variation of its growth ratecould be explained by temperature and bacterial food supply.Based on these results, a multiple linear regression equationwith temperature and bacterial concentration as dependent variableswas calculated for the growth rate of Spumella. Taxon-specificproduction rates were derived from growth rates and averagebiomass of these two species, and compared to total HNF productionestimated from previously measured community growth rates andbiomass in Lake Constance. Production peaks of Spumella sp.and Kathablepharis sp. alternated seasonally. Total HINF productionranged from –0.01 to 10 mg C m^–3 day^–1. Theaverage seasonal production varied between 1.4 and 33 mg C m^–3day^–1 over 6 consecutive years. These small protozoa thuscontribute a substantial amount to total zooplankton productionin Lake Constance.     

Changes in cell volume of bacteria and heterotrophic nanoflagellates in a hypereutrophic pond

Changes in cell volume of planktonic bacteria and heterotrophic nanoflagellates (HNF) were examined in a hypereutrophic pond from April to October, 1997. There were marked changes in the abundance of bacteria, HNF and ciliates and in protistan bacterivory during this period. The cell volume of free-living bacteria (0.121 ± 0.031 m³, mean ± SD) was large relative to that reported in the literature. The cell volumes of HNF was 71.1 ± 24.8 m³. Both cell volumes did not follow a seasonal trend. The dominant size class of bacteria was seasonally variable, whereas density of filamentous bacteria was relatively high between August and September. Biomass of filamentous bacteria accounted for up to 33.6% of total bacterial biomass. A correlation analysis for cell volume of bacteria and HNF, density of filamentous bacteria and some microbial variates was performed. The positive correlations detected (p<0.05) were between density of bacteria and cell volume of HNF, and between density of filamentous bacteria and cell volume of HNF.     

Grazing of bacteria and phytoplankton by heterotrophic nanoflagellates in a Baltic Sea sample

Grazing by heterotrophic nanoflagellates on bacteria and phytoplankton was studied in a laboratory experiment, using a natural pelagic community originating from the Tvärminne sea area off the southern coast of Finland. Water was prescreened to remove larger grazers. Four experimental treatments were used: light and dark, with and without added nutrients. The growth of the large heterotrophic flagellates was stimulated by increased production of < 3 m phytoplankton. Clearance rates for heterotrophic nanoflagellates were estimated and were found to be within the range of previously reported values.     

The dynamics of heterotrophic nanoflagellates and bacterioplankton in a large ultra-oligotrophic Antarctic lake

The abundance of both heterotrophic nanoflagellates (HNAN) andbacterioplankton in a large (9km²) ultraoligotrophic Antarcticlake (Crooked Lake) were investigated from December 1992 untilNovember 1993. HNAN abundance peaked in spring, summer and autumn,falling to lowest numbers during the winter. Numbers rangedbetween 0 and 50.9x10⁴ l^–1. Bacterioplankton abundancewas highest during the late summer and then fell progressivelytowards winter and autumn (range 1.19–4.46x10⁶ l^–1)In contrast to numbers, mean cell volumes (MCV) of the bacteriareached their highest in spring, and consequently highest bacterialbiomass occurred at this time. MCV ranged between 0.052 and0.224µm³. Bacterial production measurements followingthe incorporation of [³H] thymidine into DNA and [¹⁴C] leucineinto protein using a doubling-labelling procedure were undertakenin January, June, August, October and November. Rates variedbetween 2.8 and 52 ng C l¹ h¹. On occasions, a significant differencein production rates based on the uptake of leucine and thymidinewas observed, suggesting unbalanced growth. Highest rates ofproduction coincided with times of high dissolved organic carbonlevels in the water column and lowest production with low levelsof DOC. HNAN grazing rates were measured by following the uptakeof fluorescently labelled bacteria and averaged 4.8 bacterialcells individual¹ day¹ at 2 and 4°C. Specific growth rates(h¹) ranged around 0.00070–0.00077 in both the field andlaboratory, giving doubling times of 37.3 and 41.0 days, respectively.These low rates of grazing and growth indicate that there isno adaptation to low temperatures in these freshwater protists.Based on these data, the gross production efficiency is 24%.HNAN removed between 0.1 and 9.7% of bacterial production perday.     

Trophic relationships between planktonic bacteria,heterotrophic nanoflagellates and viruses in a mesoeutrophic reservoir

The abundance, biomass, and production (Р _В) of bacrerioplankton; the taxonomic composition, abundance, biomass of heterotrophic nanoflagellates (HNF) and the rate of consumption of bacteria by HNFs; and the abundance of virioplankton, frequency of visibly infected bacterial cells, virus-induced mortality of bacterioplankton, and viral production were estimated in the mesoeutrophic Rybinsk Reservoir. The rate of bacterial mortality due to viral lysis (7.8–34.1%, on average 17.2 ± 2.0% of daily Р _В) was lower than the consumption of bacteria by the HNF community (15.4–61.3%, on average 32.0 ± 4.2% of daily Р _В). While consuming bacteria, HNFs simultaneously absorbed a significant number of viruses residing on the surface and inside the bacterial cells.     

Seasonal dynamics of free and attached heterotrophic nanoflagellates in an oligomesotrophic lake

1. The seasonal development of heterotrophic nanoflagellates (HNF), bacteria, rotiferans and crustacean zooplankton was studied in the epilimnion of Lake Pavin, an oligomesotrophic lake in the Massif Central of France.
2. HNF abundance varied from 0.1 to 2.5 × 10³ mL^–1. Free-living HNF reached their highest density in spring when the copepod Acanthodiaptomus denticornis dominated the metazooplankton. They were present in low numbers when rotifers and cladocerans were numerous.
3. Attached HNF, consisting of bicoecids and choanoflagellates, were fixed to large diatoms and to the colonial cyanobacterium Anabaena flos-aquae . The abundance of attached HNF was significantly correlated to bacterial abundance, which fluctuated between 1.1 and 2.7 × 10⁶ mL^–1. Highest abundance of these epiphytic protists was recorded when free-living heterotrophic nanoflagellates declined.
4. The comparison of the dynamics of heterotrophic nanoflagellates, bacteria, and the impact of zooplankton grazing suggested that prey abundance, the presence of suitable attachment sites and limited competition from the free-living forms were the main factors controlling the development of the epiphytic flagellate protists. In contrast, the low abundance of free-living forms during the period of rotiferan and cladoceran development suggests the prevalence of a top-down control by predation of the metazoopankton.     

Seasonal dynamics of crustacean zooplankton, heterotrophic nanoflagellates and bacteria in a shallow, eutrophic lake

1. The seasonal development of crustacean zooplankton, heterotrophic nanoflagellates (HNF) and bacteria was examined in Grosser Binnensee, a shallow, eutrophic lake in northern Germany. The grazing impact of Daphnia on bacteria and nanoflagellates was estimated from field data on population abundances and from clearance rates obtained in laboratory experiments. 2. The seasonal succession of zooplankton showed distinct peaks of Daphnia magna, cyclopopid copepods, Bosmina longirostris and Daphnia galeata and D. hynlina. The population dynamics of Dapfinia had the strongest impact on all sestonic components. Daphnia maxima coincided with clearwater phases, and were negatively correlated with particulate organic carbon (POC), HNF and phytoplankton. Bacterial abundance was only slightly affected although daphnids were at times more important as bacterial consumers than HNF, as estimated from measured bacterial clearance rates. Other crustaceans (copepods, Bosmina) were probably of minor importance as grazers of bacteria and nanoplankton. 3. HNF abundance varied from 550 ml^?1 to more than 30000 ml^?1. HNF appeared to be suppressed by daphnids and reached highest densities when copepods dominated the metazooplankton. The variation in HNF abundance was not reflected in the concentration of heterotrophic bacteria, which fluctuated rather irregularly between 5 and 20 ± 10⁶ ml^?1. Long filamentous bacteria which were probably resistant to protozoan grazing, however, appeared parallel to the development of HNF. These bacterial cells, although small in number, could comprise more than 30% of the total bacterial biomass.     

Grazing by ciliates and heterotrophic nanoflagellates on picocyanobacteria in Lago Maggiore, Italy

During the oligotrophication of Lago Maggiore, picocyanobacteria(Pcy) increased in abundance and production. In their bimodal,seasonal cycle, the spring peak was due almost exclusively tosingle cells of Pcy, whereas in late-summer/autumn the varietyof morpho-types increased and larger Aphanothece-like rods appeared.Rates of Pcy cell removal by heterotrophic nanoflagellates andciliates were measured by using fluorescently labelled Pcy (FLC)in four experiments performed during the Pcy population shiftfrom small cocci to larger rods. The ciliate community appearedto be composed mainly of oligotrichs in the first two experiments,and subsequently of scuticociliates; heterotrophic nanoflagellatesdecreased in number from May to September, and there was a sizeshift which might reflect species composition change. Peritrichsemerged as the most efficient Pcy grazers. For the other ciliategroups we observed higher individual ingestion rates duringthe spring experiments than during those performed in late summer/autumn.The heterotrophic nanoflagellates ingested from 0.5 to3 Pcyh^-1 while ciliates ingested from 18 to 80 Pcy h^-1. The grazingimpact of the heterotrophic nanoflagellate community rangedfrom 1.9 x 10³ to 8 x 10³ Pcy ml^-1 h^-1, whereas the ciliatecommunity ingestion rate was one order of magnitude lower (0.2x 10³–0.4 x 10³ Pcy ml^-1 h^-1). A significant inverse correlationbetween Pcy size and the clearance rate of heterotrophic nanoflagellatesand ciliates was found. Our results indicate that protozoa areless efficient in cell uptake when the Pcy are composed of largercells. In Lago Maggiore, the carbon flux from Pcy to protozoadecreased from 29.8 to 10.2 µg C l^-1 day^-1 (May and Septemberrespectively). A tentative balance on an annual basis suggeststhat around 80% of the carbon produced by Pcy is taken up byprotozoa and channelled to metazooplankton.     

Pollen and resource limitation in a gynodioecious species

Differences between plant sex morphs in pollen or resource availability may affect their relative fitness and thereby the sex ratio of dimorphic species. In gynodioecious species, in which hermaphroditic and female plants coexist, a variety of factors (e.g., hermaphrodite self-fertility or rarity or pollinator discrimination against females) might be expected to lead to stronger pollen limitation in females than in hermaphrodites. On the other hand, females have been found to be superior compared to hermaphrodites in low-nutrient conditions. The effects of supplemental hand-pollination and resource addition on the reproductive output of the self-fertile gynodioecious perennial Geranium sylvaticum (Geraniaceae) were tested for several populations that differ in their female frequency (4.4-23.0%). Both pollen and resource availability limited fruit set and the number of seeds produced per plant; however, seed set (i.e., the number of seeds produced per fruit) was limited only by resources. Because pollen limitation in females did not correlate with female frequency, our results suggest that pollen limitation in females does not depend on the frequency of the pollen-producing hermaphrodites. Furthermore, because pollen and resource availability limited reproductive output of both sex morphs, these factors may not contribute significantly to maintenance and evolution of gynodioecy in G. sylvaticum.     

Patterns in the top-down versus bottom-up regulation of heterotrophic nanoflagellates in temperate lakes

The importance of resources versus that of predators in determiningthe abundance of organisms is modulated by several factors whichvary over time and space. Here, we evaluate the abundance ofheterotrophic nanoflagellates (HNF), the main predators of planktonicbacteria, in the plankton of 16 Quebec lakes as a function ofthe abundance of resources and zooplankton (predators). We analyzethe data at two different scales: among lakes varying in resourcesupply and within lakes. Resource factors (total phosphorus,chlorophyll, bacterial concentration) best explained the abundancesof HNF among lakes differing in resource availability. Whenthe effect of resource availability was removed, the biomassof cladocerans became an important determinant of HNF abundance,with abundance declining and the ratio of bacteria per flagellateincreasing, as the cladocerans increased. Within lakes, HNFabundance was not a direct function of resource concentration.Multiple regression models based on data for many lakes showedthat both bottom-up (food) and top-down (predation) variablespredicted HNF abundance quite well. An examination of the seasonalpattern among individual lakes showed each group of variablesdiffering in importance: bottom-up variables were more importantin spring and top-down variables in midsummer. An examinationof the effects of different assemblages of zooplankton on HNFshowed cladocerans to exert the highest negative impact on HNFand to be responsible for keeping low HNF numbers during mostof the summer with that probably affecting the pathways of energyflux in those lakes dominated by cladocerans versus those dominatedby other zooplankton. The absolute, as well as the relative,importance of the different biotic and abiotic factors examinedin predicting the abundance of HNF is a function of the scaleof the investigation. Among lakes varying greatly in resourcesupply, resources are found to better correlate with HNF abundance.When the analysis is performed at another scale, within lakes,the relative importance of resources and predation in determiningHNF abundances changes depending on the variability superimposedby other factors (seasonality and lake food web structure).     

Top–down control of benthic heterotrophic nanoflagellates by oligochaetes and microcrustaceans in a littoral freshwater habitat

1. We investigated trophic interactions between benthic heterotrophic nanoflagellates (HNF) and oligochaetes and microcrustaceans (cladocerans and copepods) transferred from a silty, littoral freshwater habitat to laboratory microcosms. With a newly adapted experimental design we were able to compute (i) predation rates on benthic HNF by the tested metazoan organisms and (ii) growth rates of the natural benthic HNF population when losses because of this predation were excluded. 2. The experiments covered a temperature range of 4–27 °C and a fivefold variation of predator densities (September 2000–February 2002). For 60% of these experiments, significant predation of oligochaetes and microcrustaceans on benthic HNF was revealed. Predation rates on HNF ranged from 0 to 0.256 day^?1. Growth rates of the benthic HNF assemblage varied from ?0.098 to 0.353 day^?1; they were used to estimate the significance of the measured losses in comparison with possible other loss factors. 3. The data suggested that during the major part of the year a high percentage of the HNF production was consumed in the surficial sediment of the investigated system, resulting in a relatively constant and low HNF standing stock and an uncoupling of benthic bacteria and their protistan grazers. Top–down control by microcrustaceans and oligochaetes was identified as one significant, frequently prevailing regulatory factor, while other parameters responsible for the control of benthic HNF densities remain to be examined.     

Seasonal and vertical distribution of planktonic bacteria and heterotrophic nanoflagellates in the middle Adriatic Sea

Temporal and spatial patterns of bacteria and heterotrophic nanoflagellates (HNF) were studied monthly from January 1997 to December 1998 in the middle Adriatic Sea. Bacterial and HNF relationships with phytoplankton biomass and temperature were analyzed to examine how the relative importance of bottom-up and top-down factors may shift over seasons and locations. For the coastal area, an inconsistent relationship between bacterial abundance and chlorophyll a and a stronger relationship between bacterial abundance and bacterial production suggest that other substrates than those of phytoplankton origin are important for bacteria. The analysis of simultaneous effects of temperature and bacterial production on bacterial abundance showed that the effect of temperature obscured the effects of bacterial production, suggesting that bacterial growth itself is highly temperature-dependent. The relationship between HNF abundance and bacterial abundance was slightly improved by the inclusion of in situ temperature, bacterial production or both parameters, as additional independent variables. About 60% of the variability in HNF abundance can be explained by bacterial abundance, bacterial production and temperature. In the open sea, tight coupling of bacterial abundance with chlorophyll a concentrations implied that phytoplankton-derived substrates have a dominant role in controlling bacterial abundance. During the colder months, bacterial abundance was high enough to support higher HNF abundance than observed, suggesting that predation exerted a minor depressing influence on bacterial abundance during that period. During the spring-summer period, HNF controlled bacterial standing stocks by direct cropping of bacterial production.Communicated by: H.-D. Franke     

Resource limitation,predation risk and compensatory growth in a damselfly

Periods of poor nutrition during early development may have negative fitness consequences in subsequent periods of ontogeny. In insects, suppression of growth and developmental rate during the larval stage are likely to affect size and timing of maturity, which in turn may lead to reduced reproductive success or survivorship. In light of these costs, individuals may achieve compensatory growth via behavioural or physiological mechanisms following food limitation. In this study, we examined the effects of a temporary period of food restriction on subsequent growth and age and size at maturity in the larval damselfly Ischnura verticalis (Odonata: Coenagrionidae). We also asked whether this temporary period of reduced nutrition affected subsequent foraging behaviour under predation risk. I. verticalis larvae exposed to a temporary food shortage suffered from a reduced growth rate during this period relative to a control group that was fed ad libitum. However, increased growth rates later in development ensured that adult body size measurements (head and pronotum widths) did not differ between the treatments upon emergence. In contrast, adult dry mass did not catch up to that of the controls, indicating that the increased growth rates for size dimensions occur at the cost of similar gains in mass. Predators reduced foraging effort of larvae, but this reduction did not differ between control larvae and those previously exposed to poor nutrition.     

Effects of hydrophobic and electrostatic cell surface properties of bacteria on feeding rates of heterotrophic nanoflagellates

The influence of cell surface hydrophobicity and electrostatic charge of bacteria on grazing rates of three common species of interception-feeding nanoflagellates was examined. The hydrophobicity of bacteria isolated from freshwater plankton was assessed by using two different methods (bacterial adhesion to hydrocarbon and hydrophobic interaction chromatography). The electrostatic charge of the cell surface (measured as zeta potential) was analyzed by microelectrophoresis. Bacterial ingestion rates were determined by enumerating bacteria in food vacuoles by immunofluorescence labelling via strain-specific antibodies. Feeding rates varied about twofold for each flagellate species but showed no significant dependence on prey hydrophobicity or surface charge. Further evidence was provided by an experiment involving flagellate grazing on complex bacterial communities in a two-stage continuous culture system. The hydrophobicity values of bacteria that survived protozoan grazing were variable, but the bacteria did not tend to become more hydrophilic. We concluded that variability in bacterial cell hydrophobicity and variability in surface charge do not severely affect uptake rates of suspended bacteria or food selection by interception-feeding flagellates.     

8 Grazing by heterotrophic nanoflagellates on virus- and bacteria-sized particles in two lakes of different trophy

Although viruses are now widely recognized to infect aquatic bacteria as well as prokaryotic and eukaryotic phytoplankton, our understanding of how they fit into aquatic foodwebs remains still deficient. The present study examined the potential nutritional food source that viral particles represent for natural assemblages of heterotrophic nanoflagellates (HNF). We determined the seasonal and depth-related variability in the grazing activity of HNF on virus- and bacteria-sized particles in the oligomesotrophic Lake Pavin and the eutrophic Lake Aydat. Ingestion rates were determined using 50 and 500 nm diameter fluorescent microspheres. Estimated ingestion rates ranged from 0.01 to 1.7 viruses cell⁻¹·h⁻¹ (Aydat) and from 0.01 to 2 viruses cell⁻¹·h⁻¹ (Pavin). Derived clearance rates for viruses represented 3–5% (Aydat) and 10–12% (Pavin) of those for bacteria. In general, HNF grazing on both viruses and bacteria showed a similar pattern characterized by an apparent decrease in summer followed by autumnal peaks, with lowest values always being recorded in the hypolimnion. Viral production, estimated from the frequency of visibly infected cells (using transmission electronic microscopy), was consumed by HNF at rates averaging 2% (Aydat) and 10% (Pavin). This study suggests that, viruses, which are assumed to correspond to 1×10⁻⁷ ng C/particle, are likely to be of low nutritive value for HNF in both lakes. However, these results imply that HNF grazing may contribute significantly to the removal of viruses in freshwaters, especially in low productive systems.     

Relationships between heterotrophic nanoflagellates and the demographic response of Daphnia longispina in a eutrophic lake with poor food quality conditions

1. Based on both field data and laboratory studies, the summer population of Daphnia longispina living in a stratified eutrophic lake was examined in relation to the abundance of algae, nanoflagellates and picocyanobacteria.
2. In early July, the Daphnia population replaced Bosmina and remained the dominant zooplankter during summer 1994. Its development in July was concomitant with an increase of edible algae but, despite the apparent abundance in available food, the Daphnia population decreased throughout August suggesting poor food conditions.
3. From mid-August to the beginning of September, the biomass of inedible phytoplankton was greater than that of the smaller, edible fraction. Consequently the average rate of increase, birth rates and fecundity of Daphnia remained low. Although the biomass of heterotrophic nanoflagellates was consistently low, the demographic parameters of Daphnia were correlated throughout this period with these protozoans.
4. Life table experiments run in the laboratory showed that epilimnetic food supported both the growth in length of individual Daphnia and an increase in fecundity, but metalimnetic food supported only individual growth. D. longispina probably failed to reproduce because of the abundance of detritus mixed with the heterotrophic nanoflagellates in the metalimnetic water at that period of the year. The vertical migration of Daphnia into these deeper layers could be caused by a predator avoidance mechanism.     

Juvenile survival of a planktonic insect: effects of food limitation and predation

• 1 An experiment was conducted to investigate potential impacts of food limitation and copepod predation on juvenile survival of Chaoborus purtctipennis. We tested the hypotheses that: (i) juvenile survival of Chaoborus is influenced more by copepod predation than by starvation in a productive environment, and (ii) food limitation and predation interact to affect survival.
• 2 Effects of food concentration (approximately 800, 1400 and 2300 microzooplankton 1-^?1) and predator density (0, 1 and 2 Mesocydops edax 1-^?1) on Chaoborus development and survival were evaluated using a 3 × 3 factorial design. Jars containing lake water, the appropriate food and predator treatments, and two Chaoborus (<12h old) were rotated on a plankton wheel at 25°C. Survival and developmental stage were monitored daily until all individuals had either died or moulted to instar II.
• 3 Predation by Mesocydops was the major source of mortality, causing 87.5% of Chaoborus deaths over all treatments. Chaoborus mortality was significantly higher in treatments with Mesocydops (67–100%) than in predator-free treatment (0–13%).
• 4 Development time was significantly longer in the low-density food treatment than in the highest food treatment.
• 5 No significant interaction between food limitation and predation was detected.
• 6 These results suggest that predation by copepods may limit recruitment of juvenile Chaoborus in productive lakes.

     

Local and regional processes in low-productive mountain plant communities: the roles of seed and microsite limitation in relation to grazing

It is becoming widely accepted that plant community structure is determined not only by local scale factors, but that regional factors may play considerable role. The research studying the associated processes in different environments with different species assemblages is still limited. We conducted a two-year seed sowing experiment to test whether a plant community in a low-productive mountain snowbed is limited by seed or microsite availability and how these variables depend on natural grazing. In a factorial design, half of the plots received a mixture of seeds of fourteen species naturally occurring at the study site and above ground biomass was removed from half of the plots. These treatments were applied to plots with long term grazer exclosures and to plots accessible to grazers. Both sowing and biomass removal increased the number of seedlings, the species richness of seedlings and total species richness. The number of seedlings was higher in open plots than in exclosures in the second year. Both seedling richness and total species richness were higher in open plots. Seedling recruitment was negatively related to the amount of above ground biomass and positively to the initial species richness. These results suggest that even fairly low-productive environments can be both seed and microsite limited and that these depend on grazing pressure. Natural grazing by mammal herbivores (e.g. lemmings and reindeer) favours species colonization and seedling emergence. Low-productive mountain snowbeds are prone to colonization from the local species pool and even high species richness may not constrain ingression of new species.