Blepharisma Americanum Research Articles

Colpidium spp. engulfed by a Blepharisma americanum

<i>Colpidium</i> spp. engulfed by a <i>Blepharisma americanum</i>

Predator-prey interactions between the ciliate Blepharisma americanum and toxic (Microcystis spp.) and non-toxic (Chlorella vulgaris, Microcystis sp.) photosynthetic microbes

AME Aquatic Microbial Ecology Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsSpecials AME 83:211-224 (2019) - DOI: https://doi.org/10.3354/ame01913 Predator-prey interactions between the ciliate Blepharisma americanum and toxic (Microcystis spp.) and non-toxic (Chlorella vulgaris, Microcystis sp.) photosynthetic microbes Ian J. Chapman1,2, Daniel J. Franklin1, Andrew D. Turner3, Eddie J. A. McCarthy1, Genoveva F. Esteban1,* 1Bournemouth University, Department of Life and Environmental Sciences, Faculty of Science and Technology, Dorset, BH12 5BB, UK 2NSW Shellfish Program, NSW Food Authority, Taree, NSW 2430, Australia 3Centre for Environment, Fisheries and Aquaculture Science (CEFAS), Weymouth, Dorset, DT4 8UB, UK *Corresponding author: gesteban@bournemouth.ac.uk ABSTRACT: Despite free-living protozoa being a major factor in modifying aquatic autotrophic biomass, ciliate-cyanobacteria interactions and their functional ecological roles have been poorly described, especially with toxic cyanobacteria. Trophic relationships have been neglected and grazing experiments give contradictory evidence when toxic taxa such as Microcystis are involved. Here, 2 toxic Microcystis strains (containing microcystins), 1 non-toxic Microcystis strain and a non-toxic green alga, Chlorella vulgaris, were used to investigate predator-prey interactions with a phagotrophic ciliate, Blepharisma americanum. Flow cytometric analysis for microalgal measurements and a rapid ultra high performance liquid chromatography-tandem mass spectrometry protocol to quantify microcystins showed that non-toxic photosynthetic microbes were significantly grazed by B. americanum, which sustained ciliate populations. In contrast, despite constant ingestion of toxic Microcystis, rapid egestion of cells occurred. The lack of digestion resulted in no significant control of toxic cyanobacteria densities, a complete reduction in ciliate numbers, and no observable encystment or cannibalistic behaviour (gigantism). Individual B. americanum morphological responses (biovolume and cell width) showed a significant decrease over time when sustained on non-toxic Microcystis compared to grazed C. vulgaris populations, supporting previous studies that cyanobacteria may be a relatively poor source of nutrition. Results here provide insight into the ecological interactions of ciliates and cyanobacteria, and for the first time B. americanum is shown to have the capacity to suppress potentially bloom-forming cyanobacteria. However, grazing can be significantly altered by the presence of microcystins, which could have an impact on bloom dynamics and overall community structure. KEY WORDS: Ciliate-cyanobacteria interactions · Toxic cyanobacteria · Ciliate · Grazing · Microcystis · Microcystins Full text in pdf format PreviousNextCite this article as: Chapman IJ, Franklin DJ, Turner AD, McCarthy EJA, Esteban GF (2019) Predator-prey interactions between the ciliate Blepharisma americanum and toxic (Microcystis spp.) and non-toxic (Chlorella vulgaris, Microcystis sp.) photosynthetic microbes. Aquat Microb Ecol 83:211-224. https://doi.org/10.3354/ame01913 Export citation RSS - Facebook - Tweet - linkedIn Cited by Published in AME Vol. 83, No. 3. Online publication date: September 19, 2019 Print ISSN: 0948-3055; Online ISSN: 1616-1564 Copyright © 2019 Inter-Research.

Nuclear Features of the Heterotrich Ciliate Blepharisma americanum: Genomic Amplification, Life Cycle, and Nuclear Inclusion.

Blepharisma americanum, a member of the understudied ciliate class Heterotrichea, has a moniliform somatic macronucleus that resembles beads on a string. Blepharisma americanum is distinguishable by its pink coloration derived from the autofluorescent pigment blepharismin and tends to have a single somatic macronucleus with 3-6 nodes and multiple germline micronuclei. We used fluorescence confocal microscopy to explore the DNA content and amplification between the somatic and germline nuclei of B.americanum through its life cycle. We estimate that the DNA content of the macronucleus and micronucleus are 43±8 Gbp and 83±16 Mbp respectively. This correlates with an approximate DNA content difference of 500-fold from micronucleus to macronucleus and a macronuclear ploidy of ~1,100N as compared to the presumably diploid micronucleus. We also investigate a previously reported macronuclear inclusion, which is present sporadically across all life cycle stages; this inclusion looks as if it contains blepharismin based on its fluorescent properties, but its function remains unknown. We also provide additional detail to our understanding of life cycles changes in B.americanum by analyses of fluorescent images. Overall, the data analyzed here contribute to our understanding of the diversity of nuclear architecture in ciliates by providing details on the highly polyploid somatic macronucleus of B.americanum.

Ciliados (Protozoa) de sedimentos secos de una charca temporaria de la Argentina

Temporary ponds represent special environments that are inhabited by organisms adapted to changingenvironmental conditions. Ciliates are able to survive complete loss of water in these transient habitats through cyst formation. However, ciliates from the Neotropical region in general have been poorly studied with modern techniques. The main goal of this study is to describe the ciliates in dried sediments of a temporary pond from Buenos Aires Province, Argentina, through sampling efforts that were performed 2003-2005. Soil samples were obtained during drought phases and rewetted in laboratory to establish raw and enriched cultures. Ciliates were then studied both in vivo and after impregnation with protargol. In this study, we present 4 new records for Argentina (Gonostomum affine (Stein, 1859) Sterki, 1878, Stylonychia bifaria (Stokes, 1887) Berger, 1999, Pleurotricha lanceolata (Ehrenberg, 1835) Stein, 1859, Meseres corlissi Petz and Foissner, 1992), 1 for South America (Blepharisma americanum (Suzuki, 1954) Hirshfi eld, Isquith and Bhandary, 1965), and 2 for the Neotropical Realm (Gonostomum strenuum (Engelmann, 1862) Sterki, 1878, Stylonychia lemnae Ammermann and Schlegel, 1983).

Community convergence in a simple microbial food web

AbstractPrevious studies of communities implicate many potential mechanisms that can create alternate stable states. These include density‐dependent foraging behavior, size refuges reached by early colonists, environmental feedback following disturbance, and different initial densities of intraguild predators. Previous work shows that alternate states of varying stability can occur in food webs containing the intraguild predators Blepharisma americanum and Tetrahymena vorax. Differences in colonization history could create the alternate states, consisting of dominance by either Blepharisma or Tetrahymena, but it was unclear whether results depended on effects of initial density or only on changes in the resource base. We manipulated initial densities of both species to determine if density effects alone could create alternate stable states. Convergence of these communities over time indicated that differences in initial density did not create alternate stable states. By default, other factors influenced by colonization history, such as resource availability, may produce alternate states. Models of alternate stable‐state phenomena should incorporate differences in resource availability in addition to direct competitive and predatory interactions to provide a more complete depiction of the causes of differences in community composition in otherwise similar habitats.

COLONIZATION HISTORY DETERMINES ALTERNATE COMMUNITY STATES IN A FOOD WEB OF INTRAGUILD PREDATORS

Several recent studies suggest that community composition can take on alternate, apparently stable, states. For the most part, these studies are observational in nature and have not shown that alternate states arise from the order of species arrival during community assembly. Previous studies also provide little insight into the mechanisms that create alternate states, especially when direct species interactions, rather than environmental feedback, control their formation. We conducted experiments in identical physical environments to show that differences in the sequence of community assembly can create alternate species compositions that persist for many generations. Assembly sequence determined whether experimental food webs contained a single intraguild predator, or two intraguild predators. When the intraguild predator, Blepharisma americanum, became established in communities before the introduction of Tetrahymena vorax, combined effects of resource depletion and intraguild predation prevented the establishment of Tetrahymena. In contrast, when Tetrahymena preceded Blepharisma, both species frequently coexisted. Supporting experiments describe the exploitative competitive abilities of both intraguild predators, document the population dynamics of each species in less complex community modules, and determine the ability of Tetrahymena to invade Blepharisma cultures in the absence of competing prey. These results show that both competition and predation contribute to the formation of two alternate community states and confirm that one stable state resists transformation to the other.

A New Noncanonical Nuclear Genetic Code: Translation of UAA into Glutamate

Deviant genetic codes reported in ciliates share the same feature: one (UGA) or two (UAR) of the three canonical stop codons are translated into one particular amino acid. In many genera, such as Oxytricha, Paramecium, and Tetrahymena, UAR codons are translated into glutamine. UGA is translated into cysteine in Euplotes or into tryptophan in Colpoda inflata and Blepharisma americanum. Here, we show that three peritrich species (Vorticella microstoma, Opisthonecta henneguyi, and Opisthonecta matiensis) translate UAA into glutamate and that at least UAA in O. matiensis is decoded through a mutant suppressor-like tRNA. This kind of genetic code has never been reported for any living organism. Phylogenetic analysis with α-tubulin sequences corroborates that peritrichs, peniculines (Paramecium), and hymenostomates (Tetrahymena) form a monophyletic group (class Oligohymenophorea). The differential translation (glu/gln) of UAR codons, the monophyly of the Oligohymenophorea, and the common evolutionary origin of glutamate and glutamine suggest that deviant genetic codes of present-day oligohymenophoreans could have the same origin.

The molecular basis of nuclear genetic code change in ciliates.

The nuclear genetic code has changed in several lineages of ciliates. These changes, UAR to glutamine and UGA to cysteine, imply that eukaryotic release factor 1 (eRF1), the protein that recognizes stop codons and terminates translation, changes specificity. Here we test whether changes in eRF1 drive genetic code evolution. Database sequence analysis reveals numerous genetic code alterations in ciliates, including UGA --> tryptophan in Blepharisma americanum and the distantly related Colpoda. We sequenced eRF1 from four ciliates: B. americanum, a heterotrich that independently derived the same eRF1 specificity as Euplotes, and three spirotrichs, Stylonychia lemnae, S. mytilus, and Oxytricha trifallax, that independently derived the same genetic code as Tetrahymena (UAR --> glutamine). Distantly related ciliates with similar codes show characteristic changes in eRF1. We used a sliding window analysis to test associations between changes in specific eRF1 residues and changes in the genetic code. The regions of eRF1 that display convergent substitutions are identical to those identified in a recently reported nonsense suppression mutant screen in yeast. Genetic code change by stop codon reassignment is surprisingly frequent in ciliates, with UGA --> tryptophan occurring twice independently. This is the first description of this code, previously found only in bacteria and mitochondria, in a eukaryotic nuclear genome. eRF1 has evolved strikingly convergently in lineages with variant genetic codes. The strong concordance with biochemical data indicates that our methodology may be generally useful for detecting molecular determinants of biochemical changes in evolution.

Effects of intra‐ and interspecific interactions on species responses to environmental change

Abstract The extent of directional environmental change expected in the next century underscores the need to understand density‐dependent population regulation. Direct density dependence generated by intraspecific competition and/or predator–prey interactions should buffer environmentally produced changes in density‐independent growth rates (r). Density dependence generated by interspecific competition should magnify sensitivity to changes in r. We tested these predictions by assembling protist communities in laboratory microcosms and subjecting the communities to directional environmental change. Two experiments used pairs of competing protists (Colpidium striatum Stein and Tetrahymena thermophila Nanney &amp; McCoy, and Paramecium tetraurelia Sonneborn and Blepharisma americanum Suzuki) along with single‐species controls. Two experiments used predator–prey pairs (Didinium nasutum Müller or Euplotes patella Ehrenberg preying on Colpidium) and controls containing only prey. We grew replicates of each species combination in two temperature regimes (constant or slowly increasing temperature). Independent of these experiments, we quantified the temperature dependence of r (intrinsic rate of increase) for each species. Comparison of effects of temperature on r to effects on mean density revealed that intraspecific competition buffered species densities against temperature changes that increased r by over 200%. Interspecific interactions did not affect species responses to environmental change. Temperature change had weak effects on species densities, whether or not other species were present. The results suggest that natural populations, regulated by direct density dependence, may be buffered against directional environmental change. Environmental change will have large effects when populations experience little density dependence, and when environmental change has different effects on the vital rates of different species.

PRODUCTIVITY, INTRAGUILD PREDATION, AND POPULATION DYNAMICS IN EXPERIMENTAL FOOD WEBS

Theory predicts that productivity and the relative efficiency of resource utilization determine the outcome of interactions between intraguild predators and prey. Interactions between two freshwater protists, Colpidium striatum (the intraguild prey) and Blepharisma americanum (the intraguild predator), in laboratory microcosms support the predictions of intraguild predation theory. Colpidium competitively excluded Blepharisma when bacterial production was low, even though Blepharisma had a potential advantage conferred by its ability to consume Colpidium. In contrast to competitive exclusion at low productivity, Colpidium and Blepharisma coexisted for many generations at higher levels of bacterial production. Theory predicts a similar transition from competitive exclusion to coexistence as productivity increases, if intraguild predators use resources less efficiently than intraguild prey/competitors. Other experiments showed that Colpidium depresses bacterial densities more than Blepharisma, a result consistent with greater efficiency of resource utilization by Colpidium. These results indicate that simple mathematical models can provide important insights into the population dynamics of intraguild predators and prey under different levels of productivity.

Evolution of 16S-like ribosomal RNA genes in the ciliophoran taxa litostomatea and phyllopharyngea

Complete small subunit (16S-like rRNA) gene sequences were determined for two litostome (Homalozoon vermiculare and Loxophyllum utriculariae) and two phyllopharyngean ciliates (Discophrya collini and Trithigmostoma steini). Six of the eight ciliate classes are now represented in the 16S-like rRNA data base. Phylogenetic reconstructions identify Blepharisma americanum as the first ciliate branch but the heterotrichs B. americanum and Metopus palaeformis do not appear to be closely related. Later branching ciliate lineages are represented by the litostomes, spirotrichs and a heterogeneous group that includes colpodid, oligohymenophoran, and phyllopharyngean ciliates. The relatively simple cytostomal organization of litostomes and phyllopharyngeans should not be regarded as primitive states. Using the rRNA phylogenies as framework to understand the evolution of non-molecular features, the low complexity of their oral structures can be interpreted as a secondary adaptation — possibly caused by a shift towards macro-phagous feeding behavior.

Phylogenetic Relationships of Blepharisma Americanum and Colpoda Inflata Within the Phylum Ciliophora Inferred From Complete Small Subunit Rrna Gene Sequences

The complete small subunit rRNA gene sequences of the heterotrich Blepharisma americanum and the colpodid Colpoda inflata were determined to be 1719 and 1786 nucleotides respectively. The phylogeny produced by comparisons with other ciliates indicated that C. inflata is allied more closely with the nassophoreans and oligohymenophoreans than the spirotrichs. This is consistent with the placement of the colpodids in the Class Copodea. Blepharisma americanum was not grouped with the hypotrichs but instead was placed as the earliest branching ciliate. The distinct separation of B. americanum supports the elevation to class status given the heterotrichs based on morphological characters.

Can Ciliates Adjust Their Intermembranellar Spacing to Prey Size?

The most rapidly cleared particle size has been reported to increase with the body size of the ciliate, a relationship attributed to changes in the morphological features of the oral apparatus. This study examined the possibility that the spacing of the adoral zone of membranelles (AZM) in Blepharisma japonicum could change or adapt to different sizes of prey. To test this hypothesis, specimens of B. japonicum were cultured on the bacterium Proteus vulgaris, the small ciliate Tetrahymena pyriformis, and smaller individuals of B. japonicum, each of which represented a different size of prey. After completing the culture period, a portion of the AZM was viewed by scanning electron microscopy and measurements of intermembranellar spacing were made from the micrographs. Significant differences (P = 0.01) in intermembranellar spacing were found between treatments: the smaller the prey size, the greater the spacing between the cilia of adjacent membranelles. The relationship between body size and particle size in Blepharisma cannot be explained by the inverse relationship already noted. The possibility remains that other oral structures (e.g., the gullet) may govern this relationship. In natural waters, the range of food particle sizes often changes seasonally. Therefore, filter feeding organisms would benefit by being adaptable to different particle sizes. Copepods have been shown to shift their grazing pressure from one size of particle to another in order to take advantage of peak particle concentration (Poulet, 1974). Nival & Nival (1976) formulated an equation for filter feeding efficiency from a number of quantitative setal characteristics including setal spacing. They were able to show that the most efficiently filtered particle size increased throughout the maturation of the copepod. Richman et al. (1977) suggested that the flexibility in copepod feeding behavior cannot be explained solely by the mechanism of a fixed sieve, but they admitted not being able to offer an alternative explanation until the kinetics of filter feeding become more clearly understood. Ciliates, like copepods, are suspension (filter) feeders, but unlike copepods, possess cilia rather than setae for filtering. Fenchel (1980a,b) studied suspension feeding organelles and suspension feeding efficiency of ciliates in relation to particle size. He found: (1) that the free space between membranellar cilia determined the minimum size of particle retained; and (2) that the size of particle most rapidly cleared increased with the size class of Blepharisma americanum (see Fenchel, 1980b). He attributed the latter relationship to changes in the morphological properties of the oral apparatus. The oral apparatus of Blepharisma consists of both oral ciliature and cytopharyngeal struc1 We thank David Montagnes, Guy Gilron, and Stephen Wickham for their enthusiastic support and helpful criticism throughout this investigation. Research supported by the Department of Zoology and by N.S.E.R.C. Canada Operating Grant A-6544 awarded to D. Lynn. 2 Person to whom reprint requests should be addressed. TRANS. AM. MICROSC. SOC., 104(4): 333-340. 1985. ? Copyright, 1985, by the American Microscopical Society, Inc. This content downloaded from 157.55.39.27 on Sat, 26 Nov 2016 04:15:15 UTC All use subject to http://about.jstor.org/terms TRANS. AM. MICROSC. SOC. tures (Giese, 1973). The two major components of the oral ciliature are: (1) the adoral zone of membranelles (AZM), which functions to set up a vortex and draw in food particles from the surrounding water; and (2) the undulating membrane, which directs the particles into the cytostome (Giese, 1973; Sleigh & Barlow, 1982). Since the AZM functions to concentrate the food particles, it also may function in the selection of particle size (Fenchel, 1980a). In the present study, intermembranellar spacing was chosen as a possible site of long-term adaptation. The membranelles of the AZM are anchored by microtubular rootlets interconnected with microfilaments (Dembitzer & Hirshfield, 1966; Giese, 1973). By contraction of the filamentous components, the ciliate may be able to change intermembranellar spacing. However, Fenchel's (1980b) study assumed that intermembranellar spacing is more or less constant for a species. We assumed, then, for the purposes of this study, that the intermembranellar spacing is constant and does not change or adapt to prey size. Blepharisma were fed natural prey organisms to test for the possibility of adaptation of intermembranellar spacing to size of prey after several weeks of exposure to prey. Exposure to actual prey represents a more plausible ecological situation than using latex beads (Fenchel, 1980b) since the prey organisms chosen are known to support growth of Blepharisma (see Giese, 1973). Since a ciliate is more likely to encounter a monospecific patch of prey organisms in the natural environment, Blepharisma has been grown with one prey species at a time. MATERIALS AND METHODS

Determination of lethal dose in a protozoa: screening of contrast media toxicity.

Suitability of a protozoan culture for determination of contrast media (CM) toxicity was investigated. A culture of Blepharisma americanum was exposed to varying concentrations of CM and the time elapsed until 50% cell mortality was determined. The cultures were standardized using solutions of Na iothalamate and/or Metrizamide. A standardized culture responded reproducibly between days 6 and 10 of the culture age. Comparison of concentrations of CM needed to achieve LD50 at 7 minutes ranked CM in order of decreasing toxicity as follows: oral cholecystopaques, intravenous cholecystopaques, and urographic agents; ie ionic monomers, ionic dimers, monovalent dimers, nonionic monomers, nonionic dimers. Comparison of CM toxicity determined by protozoa LD50 at 7 minutes with i.v. LD50 in mice showed a good correlation, suggesting usefulness of the protozoan assay as a complementary or screening toxicologic method.

Induction of Macrostome Formation in Blepharisma americanum (Suzuki, 1954) by Alpha-Tocopheryl Succinate

LENNARTZ, D. & BOVEE, E. 1980. Induction of macrostome formation in Blepharisma americanum (Suzuki, 1954) by alpha-tocopheryl succinate. Trans. Amer. Micros. Soc., 99: 310-317. The transformation of the heterotrich ciliate, Blepharisma americanum (Suzuki, 1954), from a bactivore to a carnivore, or cannibal giant is described. This transformation process is induced by exogenous d-alpha-tocopheryl acid succinate present at 10-6 to 10-4 M. In the presence of such concentrations, B. americanum undergoes a sequential alteration of morphology, especially with respect to the oral ciliature and buccal cavity. The length and volume of the organism also are increased greatly. Over a 6-10-h period following the addition of the vitamin, Blepharisma americanum develops an enlarged undulating membrane and adoral zone of membranelles due to the elongation of the component cilia. At 10-4 M tocopheryl, some monsterism is observed in the form of L-shaped or greatly elongated organisms after 3.5 h. At lower concentrations, transformation takes 8-15 h, and fewer individuals attain giant size. When B. americanum is removed from the vitamin solution, washed, and placed in sterile Chalkley's solution at pH 6.8, the ciliates revert to a normal bactivorous form by means of numerous cell divisions over a 3-day period. The results obtained in this study are essentially the same as the results of similar research on forms found in declining cultures of B. americanum, and bear considerable resemblance to the stomatin-induced transformations of Tetrahymena vorax. However, in the present work, a defined substance was used to induce transformation.

Regeneration Rate of Blepharisma with Special Reference to the Effect of Temperature*

SYNOPSIS. The regeneration of three species of Blepharisma—americanum, undulans and japonicum (grown in conslant yellow light in a constant‐temperature room at 25d̀C) from specimens of which the hypostome had been removed, was studied under a variety of conditions. The reliability of the methods was checked by an extensive series of experiments with B. americanum and B. japonicum under normal conditions. The time of regeneration of 50% of the individuals was taken as a convenient means for comparing action of various agents. No diurnal rhythm of rate of regeneration was observed. Regeneration is readily affected by the state of nutrition of the cells, starved ones regenerating more slowly than well‐fed ones. Regeneration is accelerated by maintaining the cells at 30d̀ but retarded at 35d̀. Even brief exposures to 40d̀ may retard regeneration. Regeneration is slowed by maintenance at temperatures below 25d̀, and stopped at 10d̀. Cut Blepharisma placed at 5d̀ lose pigment in a “shock” reaction and many die. The injury is more marked in lettuce medium than in balanced salt solution. Between 13 and 30d̀ the rate of regeneration increases with rise in temperature, the Q10′s for the ranges 13 to 18d̀, 18 to 25d̀ and 25 to 30d̀ being, respectively, 6.92, 3.93 and 1.56. The effect of temperature upon division of Blepharisma was found to be similarly sensitive to temperature, the Q10's for the ranges 13 to 18d̀, 18 to 25d̀ and 25 to 30d̀ being 3.87, 2.92 and 1.59, respectively. High and low temperature limits for division were also found to be similar to those for regeneration. Preliminary evidence indicates some degree of thermal adaptation in Blepharisma to both higher and lower temperatures than those usually tolerated. The studies indicate a general similarity in sensitivity of regeneration and division to environmental influences in the various races of the species studied, with some individual variations.