Predator-Prey Interactions in Ciliated Protists

Protists appeared relatively early in evolution, about 1.8 billion years ago, soon after the first prokaryotic organisms. During this time period, most species developed a variety of behavioral, morphological, and physiological strategies intended to improve the ability to capture prey or to avoid predation. In this scenario, a key role was played by specialized ejectable membrane-bound organelles called extrusomes, which are capable of discharging their content to the outside of the cell in response to various stimuli. The aim of this chapter is to describe the two main strategies adopted in ciliate predator-prey interactions: (a) the first is mediated by mechanical mechanisms and involves, for example, extrusomes called trichocysts and (b) the second is mediated by toxic secondary metabolites and involves different kinds of chemical extrusomes

Purification and initial characterization of two pheromones from the marine Antartic cilate, Euplotes nobilii

Among a set of wild-type strains of Euplotes nobilii, every one derived asexually from one specimen isolated from Terra Nova Bay (Ross Sea, Antarctica), two were found to be representative of different mating types mutually capable of inducing each other to form mating pairs through pheromones constitutively secreted into the extracellular environment. Pheromones of strain AC-1 were purified to homogeneity and shown to be represented by two distinct proteins, that were denoted En-1 and En-2. En-1, secreted in amounts three-fold greater than En-2, was determined to have a molecular weight of 5617 and an asparagine at the N-terminus of its amino acid sequence, while En-2 has a molecular weight of 6290 and bears an asparctic acid at its N-terminus. The fact that En-1 and En-2 are coreleased by genetically identical cells of the same strain was taken to imply that they carry a heterozygotic combination of allelic pheromone genes and that these genes are regulated by relationships of co-dominance

Euplotes pheromones

Species of Euplotes secrete protein signals (pheromones) in relation with their mating-type mechanisms of self-nonself recognition. Significant numbers of these pheromones have been characterized to varied extents of structural complexity in E. raikovi, E. nobilii, and E. octocarinatus. In every case, they form species-specific families of structurally homologous, cysteine-rich proteins. In E. raikovi, these proteins have been shown to share a common architecture based on a three-helix bundle, that permits them to bind to cells in competition with one another and elicit varied, context-dependent responses

Blepharismins used for chemical defense in two ciliate species of the genus Blepharisma, B. stoltei and B. undulans (Ciliophora: Heterotrichida)

It is known that the freshwater heterotrich ciliate Blepharisma japonicum uses five pigments called blepharismins, stored in its extrusive pigment granules, for both light perception and chemical defense against predators. In this work we focused our attention on the defensive strategies of two additional pigmented species of Blepharisma, B. stoltei and B. undulans. In particular: (1) we observed the predator\u2013prey interactions of B. stoltei or B. undulans against one multicellular and two unicellular predators; (2) we clarified the nature of B. stoltei and B. undulans pigments by means of High Performance Liquid Chromatography (HPLC) and Liquid Chromatography Mass Spectrometry (LC-MS); and (3) we demonstrated and compared the toxicity of the purified pigments on a panel of ciliated protists, and against one metazoan predator. The results indicate that the chemical defense mechanism present in B. stoltei and B. undulans is mediated by the same five blepharismins previously characterized for B. japonicum, although produced in different proportions

An Experience on Environmental Education Based on Respect-Reduce-Reuse-Recycle in a Village Primary School in the Marche Region, in Italy

The importance of environmental education is widely recognized worldwide: an important objective is that the citizens are accustomed to apply the best available practices in their life, while asking the administration and the government to gradually enforce measures leading to a reduction of the environmental impact and to a more sustainable lifestyle. This requires a formation starting at an early age: however, in Italy the relevant indications from the National Curriculum are still limited and do not suggest explicitly the performance of any practical activity, tailored on the specific school setting. This inadequacy of the Curriculum becomes particularly apparent in cases where other concerns are present, for example the areas in the Marche region affected by the 2016 earthquake: here, the need to resume a day-by-day routine put a kind of “damper” on other questions, such as environmental issues. This had an effect also in children’s life: on this question, school needs to take a stand, which is the objective of this study.  This work concerns an approach to environmental education in a multi-classes primary school context (age 6-10) in a small village school, located in the Central Apennine in Italy. This is performed through an experience carried out by investigating the behavior towards waste production of the community, as perceived by children at school. It is therefore articulated through the application of 4R (Respect, which encompasses Reduce, Reuse and Recycle) applying and extending the indications from the Italian National Curriculum. The project leads also to an experience of self-production of objects by the use of waste, to improve their awareness of the possibilities included in their upcycling. Feedback on the experience is finally acquired from the children and discussed, especially in the sense of creating future citizens with a more sensible and reasonable behavior towards the environment and leading possibly to zero-waste strategies in their family and work setting. The data were analyzed using thematic analysis and different themes about waste production were singled out. Keywords: Environmental education, reduce-reuse-recycle, respect, DIY materials, good practices DOI: 10.7176/JEP/11-15-04 Publication date:May 31st 202

Structural characterization of En-1, a cold-adapted protein pheromone isolated from the Antarctic ciliate Euplotes nobilii.

The second of two diffusible cell signal proteins (pheromones) purified from a wild-type strain of the Antarctic ciliate, Euplotes nobilii, has been determined by automated Edman degradation of the whole molecule and peptides generated by its chymotryptic digestion. The proposed sequence of 52 amino acids of this new pheromone, designated En-1, is: NPEDWFTPDT10CAYGDSNTAW20TTCTTPGQTC30YTCCSSCFDV40VGEQACQMSA50QC. In common with the previously determined 60-amino-acid sequence of the other pheromone, En-2, it bears eight cysteines in conserved positions (presumably linked into four conserved intrachain disulfide bonds), and physicochemical features of potential significance for cold adaptation, such as a reduced hydrophobicity, an increased solvent accessibility, and an improved local backbone flexibility. However, En-1 diverges from En-2 for having evolved a threonine cluster in the place of a glycine cluster to apparently make more flexible a region that is likely functionally important

Structural characterization of a protein pheromone from a cold-adapted (Antarctic) single-cell eukaryote, the ciliate Euplotes nobilii

Free-living species of ciliated Protozoa control their vegetative (mitotic) proliferation and mating (sexual) processes by diffusible, cell type-specific protein signals (pheromones). One of these molecules, designated En-2, was isolated from a species, Euplotes nobilii, living in the stably cold marine waters of Antarctica, and its complete amino acid sequence of 60 residues was determined by automated Edman degradation of the whole protein and peptides generated by trypsin digestion. The proposed sequence is : DIEDFYTSETCPYKNDSQLA20- WDTCSGGTGNCGTVCCGQCF40SFPVSQSCAGMADSNDCPNA60. The En-2 structure appears to be characterized by an adaptive insertion of a glycine-rich motif potentially capable to confer more flexibility to a functionally critical region of the molecule

DNA binding and oxidative DNA damage induced by climacostol\u2013copper(II) complexes: Implications for anticancer properties

Climacostol is a natural toxin isolated from the freshwater ciliated protozoan Climacostomum virens and belongs to the group of resorcinolic lipids. Climacostol exerts a potent antimicrobial activity against a panel of bacterial and fungal pathogens. In addition it inhibits the growth of tumor cell lines in a dose-dependent manner by inducing programmed cell death via intrinsic pathway. In this work, we investigated the possibility that climacostol exerts a prooxidant effect, inducing plasmid DNA strand breakage and eukaryotic DNA damage in presence of Cu(II) ions. Inhibition of DNA breakage using SOD, catalase and neocuproine confirmed the involvement of reactive oxygen species and Cu(I) ions in the DNA damage. UV\u2013visible absorption changes and mass spectrometric analysis identified a product of reaction as a deprotonated form of climacostol. Study of the interaction with DNA, using fluorescence spectroscopic techniques, showed that climacostol binds with DNA. Given the structure\u2013activity relationship of this compound and the mechanism of its prooxidant effect, we propose that the Cu(II)-mediated oxidative DNA damage by climacostol could explain its antimicrobial and antiproliferative activity

Protists exhibition in zoological garden.

The characteristics and roles of zoological gardens are described by the law n. 73/2005 in implementation the Directive CE 1999/22, "to promote and implement education and awareness programs on biodiversity". The role of zoos is extremely important for communicating ecological goals and their objectives. In fact, zoos represent a channel for communicating biodiversity in general. The species kept in zoos always have two major constants: high educational and conservation value. For these reasons we have decided to create an exhibit dedicated to the protists. Indeed, the protists are one of the best models of single-celled organisms for use in teaching biology. We focused on heterotrophic protists, usually known as “protozoa”, in order to show the unicellular life. To better show unicellular organisms, it was thought to use colorful and large taxon, and to this purpose Blepharisma japonicum is certainly a valid species. The breeding of this species is not difficult, and cells can be kept alive by means of bacterial feeding (rice seed). In this case, it will be sufficient to attach a modest magnifying glass ( ̴10x) to see the cells swimming. We are currently also thinking of setting up a tank that hosts flagellate algae of the genus Chlorogonium, in this case it will be impossible to see the individual algae, but you will be just the green color. Each breeding tank will have its own explanation panel with text and images. The exhibition is designed without the use of electricity. On the website of the structure, it will be possible to deepen the topic with multimedia material. For the writing of the popular texts we have decided to assign a degree thesis to a student of "Primary Education"

The chemical defense/offense in ciliated protists by means of toxic secondary metabolites.

Several species of protists developed a number a different strategies intended to improve prey capture or avoid the predation. Particularly, in ciliated protist, the predator-prey interactions are often mediated by means some peculiar ejectable membrane-bound organelles generally called extrusomes. These organelles are usually localized in the cell cortex and attached to the cell membrane, sharing a common characteristic in discharging their contents to the outside of the cell in response to different stimuli. Some of these extrusomes are known to function as defensive organelles as the cortical granules in Climacostomum virens; others were demonstrated to have an offensive function (such as the toxicysts in raptorial ciliates, for example in Colpes hirtus), but the role of many other extrusomes remain unknown. Several of these extrusomes are known to produce toxic secondary metabolites that have been isolated from ciliates, and many of these molecules seem to be the result of evolutionary selection related to defense and/or predatory mechanisms. To date, only few of these molecules have been deep studied in function and biological activities, and include keronopsin isolated from Pseudokeronopsis rubra, euplotins from Euplotes species, blepharismins from Blepharisma japonicum, stentorin from Stentor coeruleus and climacostol from Climacostomum virens