Protistology 7 (4), 218–237 (2012) Protistology

New views on the megaclassification of *

Stefan Luketa

Department of and , Faculty of Science, University of Novi Sad, Serbia

Summary

This paper provides a review of the existing literature about megaclassification of the living beings. Novel classification system is introduced, including 5 dominions and 17 kingdoms. In addition, a new nomenclature of higher taxa has been proposed with a premise that the dominion names end with a suffix “-biota”, subdominions with a suffix “-konta”, kingdoms with a suffix “-ida” and subkingdoms with a suffix “-ides”. Dominion Prionobiota comprises acellular without nucleic acid, acellular organisms with nucleic acid have been classified in dominion Virusobiota, dominion Bacteriobiota consists of typical prokaryotic organisms, and dominion Archaebiota comprises specific organisms of prokaryotic structure, whereas dominion Eukaryobiota entails all living beings with eukaryotic organization.

Key words: Bikonta, , nomenclature, priones, , Unikonta, viruses

Introduction 1981, 1988, 1998; Parker, 1982; Shipunov, 2009), and certain authors perceive biodiversity through As humankind grew accustomed to a great numerous kingdoms (e.g. Leedale, 1974; Drozdov, variety of the living beings, a need arose to assort 2003). Nevertheless, none of the up-to-date them. Principles of classification largely differ and classification systems assumes an existence of only three main schools of thought can be distinguished. two kingdoms. In addition, a taxonomic category Part of the authors follow Aristotle and acknowledge beyond a level of – dominion or existence of not more than two kingdoms of the – is being used by some researchers (e.g. Moore, living beings (e.g. Dillon, 1962; Hadži, 1970), some 1974; Traub, 1975; Woese et al., 1990). advocate a small number of kingdoms (e.g. Fries, History of megaclassification can be divided 1821; Hogg, 1860; Dobell, 1911; Copeland, 1956; into four periods. The first period is characterized Traub, 1963, 1964, 1971, 1975; Whittaker, 1969; by the use of predominantly external morphological Moore, 1971, 1974; Taylor, 1978; Cavalier-Smith, characteristics of the living beings. During this period, main instruments for the analysis of biological characteristics were binocular magnifier * This article is published as an “opinion in dispute”. and light . This period lasted until Members of the Editorial Board of “Protistology” do not the invention of an electron microscope. In the necessarily concur with the author. course of the second period, by means of electron

© 2012 The Author(s) Protistology © 2012 Protozoological Society Affiliated with RAS Protistology · 219 the common nature of the living beings considerable amount of trouble. After publication of progressively became more obvious on the cellular these treaties, the interest for systematics began to level. During the third period, biochemical and decline. This period brought experimental biology molecular characteristics of the living creatures (, embryology and ) into focus, were taken as basis for interpretation of evolutionary and the systematics remained in the shadow of these associations. At the beginning of the 21st century, biological disciplines until the eighth decade of the the question of life definition was re-posed, starting 20th century. the fourth period of megaclassification history. The Even though during the mid-20th century it landmark of this period is not only systematization became clear that no sharp boundary existed between and rather superficial interpretation of facts obtained and protozoans, some attempted to by instrumental methods, but also an attempt to distinguish between these two groups. Unusual and comprehend a multitude of discrete data in the most uncomplicated systematization of the living world general fashion, taking on elements of philosophy. was proposed by Dillon (1962). According to this Since Aristotle’s time, the living beings have author, all single-celled organisms represented a been traditionally divided into two groups – part of the kingdom and the kingdom (Vegetalia) and (Animalia). This division entailed only multicellular heterotrophic organisms. is based on the plainly visible differences in the Within his , Dillon placed the outer appearance and the structure of plants and animal kingdom between the green and , animals (Ranković, 1994). Plants are autotrophic but still closer to the brown algae. According to his organisms, capable of synthesizing necessary viewpoint, the animals were direct descendants of “building blocks” of their cells from simpler in- the brown algae. organic compounds, using sunlight as a source of In 1970, Hadži postulated a classification system energy. Animals are heterotrophic organisms which, of the living beings that acknowledged two kingdoms in order to satisfy their energy requirements, use – plants and animals. Hadži believed that the the available chemical energy bound in organic differentiation between plants and animals occurred compounds. Next to the mode of nutrition, other as a clean-cut bifurcation; therefore, the algae should important differences have been elaborated, such be grouped within the plants and the protozoans as mobility – plants are immobile, whereas animals within the animal kingdom. According to this school are mobile. Carl Linnaeus was the first to introduce of thought, the term Protista would not be perceived taxonomic categories into the classification of the as a systematic category. All the which are living beings and assign the status of a kingdom to able to perform , as well as the closely plants (Vegetabilia) and animals (Animalia). related organisms that secondarily switched to the The word “Biology” in the sense of science heterotrophic nutrition, ought to be considered that studied the living beings was independently as algae, i.e. plants. Accordingly, the slime molds mentioned in 1802 by two life researches – Trevira- should not be grouped among protozoans, but rather nus and Lamarck. Even though life scientists had among fungi, which this author held for plants. been engaged in studying living organisms before Hadži thought that the attempts of a switch from that, and enjoyed a status of two typical autotrophic towards the typical independent scientific disciplines. heterotrophic had not conceived a typical animal life Systematics reached its first peak upon publishing form. All of such changeovers would geologically be the work “Systema Naturae” by Linnaeus (1735). of a later date, occurring upon a former successful This work introduced order into the older systema- switch of the algae towards a protozoan state. tics of the living organisms. Since then, describing According to this school of thought, development of the new species and determining resemblances the animals that lived to see the recent age and gave between certain taxa as well as grouping them rise to multicellular animals occurred only once. So, into systematic categories became the main task animals are monophyletic; they originated not from of systematics. After Darwin (1859) published his fully developed plant predecessors, but rather from the work “On the Origin of species…” the systematics predecessors of a lower organization. Those primitive achieved its second peak. predecessors of algae and protozoans (e.g. plants and gave the systematics a new task – interpreting the animals) were named Probionta. According to such relationships between systematic categories. Shortly understanding, neither had animals developed from before the end of the 19th century, two international plants, nor plants from animals. treaties (Candolle, 1867; Blanchard et al., 1905) Aristotle’s classification of the living world settled the questions of biological nomenclature did not come into question until the 13th century, that had been giving the systematic biologists since Albert von Bolsted postulated that the fungi 220 · Stefan Luketa were an intermediate group between plants and been observed. During the vegetative phase of the life animals (von Bolsted, cited in Ranković, 1994). cycle, slime molds are amoeboid and phagotrophic, Nonetheless, it was only in 1821 that Fries put them nevertheless they form fruiting bodies which shape into a separate kingdom. Reasons for classifying and size are very similar to those of the true fungi Fungi into an independent high-rank taxon are (Vrbaški, 1993). It became clear that no straight their specific characteristics. Fungi differ from the borders could be drawn between protozoans and plants by their nutritional mode, composition of algae. the , mechanism of their response to light, In order to overcome the difficulties of fitting hormonal structure and numerous other features. into the system of the living world, Fungi are , and their cell wall contains Hogg (1860) proposed the third kingdom, named chitin. They have pores in intercellular septs of Primigenum (formal name) or Protoctista (verna- their hyphae that enable cytoplasmatic and nuclear cular name), which included all microorganisms. communication and transfer. Intercellular nuclear Haeckel (1866) acknowledged the third kingdom migration is specific for fungi alone (Ingrouille and and named it Protista, classifying microorganisms Eddie, 2006). In plants, morphogenetic processes within this kingdom. The kingdom consisted main- are activated by light with a maximum wave length of ly of four groups: algae, protozoans, fungi, and 660 nm, whereas in fungi that maximum lies between . Despite numerous differences among the 320 and 480 nm (Ranković, 1994). Developmental representatives of the kingdom, it was observed cycle of the fungi contains a haploid and a dikaryotic that the majority of species belonged to the cate- stage. Alternation of generations occurs extremely gory of unicellular organisms, whereas the fewer rare in fungi, whereas it is regular in plants. On the multicellular representatives of the kingdom had other hand, the fungi display similarities to animals. very simple body structure. In 1911, Dobell intro- Presence of chitin, as well as pigment melanin and duced the term “Protistology” and redefined the a common cytochrome system juxtapose them with concept of unicellularity. By doing so, he reinter- animals. Despite numerous differences, fungi and preted the definition of Protista as “acellular” plants share common features such as indefinite instead of “unicellular” organisms. growth ability, cell wall, absorption through the Ivanovsky found the viruses in 1892, however enlarged outer surface, biosynthetic pathway for nothing was known about their structure during the producing terpenes, and production of similar following nearly half-century, until the elucidation secondary metabolites. Early attempts to consider of viral nucleoprotein properties (Bawden and fungi as plants were based on the idea that fungi had Pirie, 1937). Two years later, the first virus particle evolved from algae (Martin, 1955). was observed by means of electron microscopy Even though a third kingdom of life – Fungi – (Kausche et al., 1939). Martin (1922) described an was separated from the other two kingdoms already unusual potato pest that turned out to be caused by in 1821, it was not widely accepted until 1969. organisms of even more simple organization than Prior to discovery of microorganisms, kingdoms viruses, later named viroids. Yet, it was only much of plants and animals were clearly demarcated. later that Diener and Raymer (1967) defined the Fungi fitted well into the plant kingdom despite nature of viroids. Two other groups of acellular many differences. As the microorganisms were organisms were discovered at the beginning of the first discovered, it seemed that they could be easily eighth decade of the 20th century. Randles and co- distributed between the initial two kingdoms based authors (1981) found one of those two groups and on the differences in mobility and nutritive mode. named them virusoids. Although the diseases caused Later on it appeared that the protozoans were by the other group – the prions – were known difficult to perceive as unicellular animals. Whereas before 1982, it was only then that Prusiner (1982) representatives characterized by phagotrophic characterized the nature of these organisms and gave nutrition and absence of cellular wall could easily them the name. fit within the definition of the animal kingdom, The third boom of systematics took place in numerous other representatives could not. Detailed the mid-20th century as a result of the notable research of flagellated protozoans has revealed accumulation of ultrastructural data from electron their specific nature, both plant- and animal-like. microscopic studies that helped to recognize the Namely, these organisms are actively motile due to remarkable similarities of all cellular organisms their “whip-like tails”, flagella. Some of them feed on a cytological level. During this period of time, phagotrophically or osmotrophically, others are a discipline that dealt with the understanding of capable of photosynthesis. Numerous resemblances phylogenetic relations between higher taxa of the between slime molds and certain animal groups have living beings developed abruptly. This discipline was Protistology · 221 named megaclassification. It provided a new motive Mereschkowski (1905, 1909), who worked it to gather all the biological disciplines into a single out in more detail. Subsequently, Wallin (1923) integrative science: biology. In 1952, Hadži wrote: postulated that the mitochondria stemmed from the “With pleasure, we can assert that nowadays a much endosymbiotic bacteria as well. These hypotheses needed balance between different areas of biology could not develop any further without gaining a has been reached, hereby attaining peer disposition profound understanding of a detailed structure of for all of them”. these , which was achievable only by the Along with the improvement of microscopic means of electron microscopy. Only L. Margulis methods, knowledge on microorganisms became (Sagan, 1967; Margulis, 1981) interpreted the more complete. Further efforts of putting such available at that time data which offered strong classification systems together that would reflect evidence to corroborate the endosymbiotic theory. evolutionary trends resulted in grouping the According to her, other organelles (flagella, cilia and living beings into two categories: Prokaryotes and peroxisomes, etc.) had also been incorporated into Eukaryotes. This division was made by Chatton the cells through endosymbiosis. The importance (1925); however, it was only in 1971 that Moore of this theory for the megaclassification of the living grasped these two categories as superkingdoms. world lies in the fact that the eukaryotes came into In 1938, Copeland was the first to understand that consideration as organisms built from well inte- the Bacteria should be classified into a separate grated groups of prokaryotic cells. kingdom. Copeland (1956) demarcated the following In 1978, Taylor introduced utilization of four kingdoms: (prokaryotic organisms), mitochondrial cristae as a leading character in Protoctista (eukaryotic microorganisms and fungi), classification of Eukaryotes. Eukaryotes with Metaphyta (plants) and Metazoa (animals). The plate-like cristae were listed within the kingdom of five-kingdom system was proposed by Whittaker in Lamellicristata, whereas those with tubular cristae 1969, separating Fungi as a stand-alone kingdom. ended up in the kingdom of Tubulicristata. Shortly, This classification system has remained widely Cavalier-Smith (1981) suggested a classification acknowledged until the last decade of the 20th system that was primarily based on the characte- century. ristics of mitochondrial cristae. This system sepa- Traub (1963, 1964, 1971) found it the most rated Eukaryotes into nine kingdoms: Eufungi, acceptable to classify the living beings into two Ciliofungi, Animalia, Biliphyta, , superkingdoms: Cellularae and Acellularae. This , Cryptophyta, Chromophyta and author divided the Cellularae superkingdom into . Cavalier-Smith (1988, 1989a, 1993) two kingdoms – Procaryotae (with subkingdoms formed the group, which included all Autobacae and Heterobacae) and Eucaryotae (with the that did not display mitochondria. subkingdoms Plantae, Heteroplantae and Animalia). According to this author, the Archezoa affiliates Moore (1974) distinguished three large groups of the primary eukaryotes that had appeared earlier living organisms and gave them a dominion rank: than the process of mitochondrial formation started. Virus, Prokaryota and Eukaryota. After Moore had Patterson (1999a) considered the Archezoa as a introduced the dominion rank into classification, paraphyletic group within the taxon . Traub (1975) revised his system by raising the level In the eighth decade of the 20th century, of superkingdom to that of dominion, kingdom an intensive consideration of biochemical and to superkingdom and subkingdom to kingdom. molecular biological traits made its way into the Obviously, this author did not form the Protista classification of the living world. Introduction of group, but rather observed Algae in the frame of such attributes into the megaclassification has led the Plantae kingdom and protozoans within the to an understanding that the living world is far more Animalia kingdom. Traub separated the kingdom diverse than previously believed. Due to that, most Animalia into two subkingdoms: Protozoae and authors postulated the existence of more than five Metazoae. It is worth noticing that the viruses were kingdoms. Nevertheless, some biologists proposed included in the classification systems of the sixties systems with fewer kingdoms, in attempt to decrease and seventies in the 20th century, whereas later on the in megaclassification. they were completely ignored. Parker (1982) divided all living beings into two Schimper (1883) noticed the similarity between superkingdoms: Prokaryotae and Eukaryotae. He and plant chloroplasts and con- integrated viruses (Virus) and bacteria (Monera) cluded that the chloroplasts stemmed from endo- as kingdoms of the first superkingdom. Within the symbiotic bacteria. However, this hypothesis second superkingdom, Parker also introduced two was not acknowledged until the publications of kingdoms: Plantae (algae, fungi and plants) and 222 · Stefan Luketa

Animalia (protozoa and animals). Clearly, this while composing classification systems of the living author did not recognize protists as formal taxon; world. nevertheless, he considered viruses as the living La Scola with co-authors (2003) described a beings. gigantic DNA virus named Mimivirus, with the Cavalier-Smith (1988) published a classification capsid diameter reaching 400 nm. This virus was system consisting of two large taxa with an empire the largest known virus of that time, and it was status – Bacteria and Eukaryota. Within the presumed that Mimivirus harbored an enormously Bacteria, two kingdoms were proposed – Eubacteria large genome. This presumption was confirmed as and Archaebacteria, whereas the empire Eukaryota the genome length of 1,182 kb had been ascertained was divided into two superkingdoms – Archezoa (Raoult et al., 2004). It is of interest that the (comprising only the Archezoa as a kingdom) and Mimivirus genome contained three genes for Metakaryota (composed of five kingdoms: Proto- aminoacyl-tRNA-synthetase (Raoult and Forterre, zoa, Plantae, Animalia, Fungi and ). 2008). In 1998, Cavalier-Smith proposed a six-kingdom Häring with co-authors (2008) described a classification system of the living beings – Animalia, novel virus with a hyperthermophylic prokaryotic Protozoa, Fungi, Plantae, Chromista and Bacteria. organism of the group as a host. This species According to Shipunov (2009), the living beings in acidic hot springs. The lemon-shaped virus were grouped into four kingdoms – Monera, that infects this species develops a filament at each Protista, Vegetabilia and Animalia. These authors of the two virion endings once it exits the host acknowledged the existence of paraphyletic groups. cell, which means that the virion is not necessarily Leedale (1974) made an exceptional, revolutio- inactive functionally all the time, as previously nary step by demarcating 19 kingdoms. However, believed. this system had never been widely acknowledged, Arslan with co-authors (2011) described a DNA since Whittaker’s system was a lot more pragmatic. virus akin to Mimivirus and named it Megavirus. Namely, Leedale (1974) segregated a large number The Megavirus virion reaches 520 nm in diameter; of insufficiently studied groups of microorganisms, its genome size is 1,259 kb, and it contains four whereas Whittaker (1969) integrated all eukaryotic genes for aminoacyl-tRNA synthetase. Lately, microorganisms into a single kingdom. Within the other authors also hold viruses for the living beings coming thirty years, large number of classification (La Scola et al., 2008; Pearson, 2008; Claverie and systems turned up, assuming a large number of Ogata, 2009). Certain authors (Koonin et al., 2006; kingdoms. For example, Drozdov (2003) published Forterre, 2010; Koonin, 2010) even communicated the system encompassing 26 kingdoms. that the complex DNA viruses (giant viruses) should Woese and Fox (1977) concluded their research be seceded into a separate domain. Yet, none of of prokaryotic organisms by denying the phylogene- them have made a clear standpoint or in a customary tic base of division into Prokaryotes and Eukaryotes. way suggested any novel classification system, These authors acknowledged three groups of the considering such action as premature. Boyer with living beings: Archae, Bacteria and Eucarya (Woese co-authors (2010) separated giant viruses into a et al., 1990). As it had already been clear that the discrete domain, named Nucleocytoplasmic Large living beings were extremely diverse, introducing DNA Viruses (NCLDV); however, these authors did less than three kingdoms was not seen as pragmatic, not give a Latin name to the fourth domain either. since such system would require large number of subsidiary categories between a kingdom and General considerations division. Therefore, Woese with co-authors (1990) introduced the term “domain”. The division of the The above introductory part is a brief overview living beings into three domains became nearly of the history of research in the field of classification commonly adopted at the end of the 20th century: of life. Clearly, considerable confusion does exist Bacteria, Archaea and Eukaryota. in this research area. In the further section, a novel At the beginning of the 21st century, the question view on the classification and nomenclature of the of life definition was again put forward in the center living world will be proposed. of scientific discussions. In particular, this question In order to set up a classification system, one first relates to the dilemma: whether viruses exist as of all needs to define life as a term. The nature of life, the living beings, or they are representing forms of however, is yet insufficiently grasped to generate a inorganic matter. Novel discoveries in the area of common direct and adequate definition. Nowadays, challenged the opinions of most authors we are aware that life represents the highest form of that had for more than two decades ignored viruses mobility of matter and that it is an essential feature Protistology · 223 of the living beings. Unlike abiogenic matter, the that a part of the cellular genetic information or living beings are subjects to the , and they its informational RNA, as a copy of the cellular are capable of reproducing. Still, the majority of DNA molecule, acquired a self-reproducing contemporary authors regard the cell as a common ability and began its development independently structural and functional unit of all living beings. of the cell. Presumably, viruses originated from Does the cell represent a sole, basic structural plasmids (naked, cyclic DNA molecules, which are unit of the living beings, or it is the most complex transferable between cells) or transposones (parts of elementary structural corpuscle of the living beings? DNA molecule capable of commuting within a cell This dilemma puts forward a question of structure as genome). According to this theory, transposones as the criteria for determining the notion of life. mobile genetic elements could be the predecessors Status of viruses is certainly the most questiona- of today’s viruses. ble issue in the classification of the living world. Upon the discovery of viruses and electron Many authors do not consider the viruses as living microscopy, even tinier acellular organisms were beings at all and for those reasons do not deal with observed and considered as sub-viral particles in them within the classification systems. Two stron- literature. Viroids, virusoids and prions have been gest arguments underpin the hypothesis according assorted within this group of organisms. Currently, to which viruses are regarded as non-living entities: the commonly accepted classification system divides the viral ability to form mineral-like crystals and the acellular organisms into six groups. Viruses their inability to self-sufficiently divide and grow. represent the first five groups, and the sixth one Other authors consider viruses alive because they consists of subviral particles. contain nucleic acids. Understanding that the postulate about cells Today, three groups of theories exist that deal being the only elementary structural units had no with the origin of viruses. One says that there is no substantial ground brought about the nucleic acids doubt about certain organelles coming into living as criteria for differentiating the living creatures organism as intracellular symbionts. Namely, pre- from the non-living matter. Nevertheless, the decessors of contemporary eukaryotic cells had once prions as protein particles with no nucleic acid ingested free-living prokaryotes, which became content are capable of changing and reproducing. adapted to survival and proliferation inside the hosts No other natural body without nucleic acid is cell over time. Secondary plastids and other complex able to evolve or reproduce. Due to these features, structures originated in a similar way through prions may be considered as the living beings. It is successive rounds of endosymbiosis. Such way of life obvious that the viroids, virusoids and viruses share inevitably leads to structural simplification of what a number of common characteristics, whereas the were to become the organelles. Some authors suggest prions reproduce in an entirely different fashion. that the acellular organisms appeared in much Additionally, prion structure is uncommon to that of similar manner. Chlamydia, a rather simplified other acellular organisms that contain nucleic acid. intracellular parasite, is commonly presented as the It can therefore be assumed that the evolutionary evidence for such theory. Moreover, structure and origin of prions is not connected with the origin of of acellular organisms and Chlamydia other acellular organisms. largely differ, making it difficult to assume the Acellular organisms manage to maintain their mechanism of acellular organisms evolving from elementary traits at minimum energy requirement cells. This theory cannot explain the origin of RNA as a pre-requisite to survival and . I viruses, since both eukaryotic and prokaryotic cells believe that it is important to alter the definition of have DNA-based genome. The regressive viral life according to novel discoveries of the extreme life origin theory is able to explain the origin of the DNA diversity, but not reassess the life diversity according genome and the intracellular life of viruses; however, to life’s definition. For this reason, I propose the it fails to clarify the next step – appearance of the following definition: the living beings are natural viral envelope. This theory of the viral origin leaves entities that have the ability of reproduction. numerous questions unanswered. Since the current literature does not offer clearly According to another group of hypotheses, viruses define rules for the nomenclature of the higher taxa, represent acellular organisms labeled as protobionts. I am hereby proposing a simple system of rules to Authors that are advocating this view regard viruses as classify the tree of life. I suggest that the domain offsprings of the primal organisms that appeared on names for the living beings end with a suffix “-biota”, Earth in its ancient past. This theory cannot explain subdomains with “-konta”, kingdoms with “-ida” the origin of the cell from a virion. and subkingdoms with “-ides”. In addition, I share Third group of theories about viral origin claims the views of Dubois (2007) and Shipunov (2009), 224 · Stefan Luketa who consider the name of the author upon the name cellular membrane of neuronal and glial cells. Innate of the higher taxa unnecessary, since it would only structure of prions does not cause any disorders. The make the contemporary megaclassification even exact function of this protein is still unknown; it has more complicated. been assumed that it plays a role in maintenance of It is important to stress that the term “domain”, the cellular membrane integrity and in transmission suggested by Woese with co-authors (1990), in fact of neural impulses (Prusiner, 2004). Primary represents a synonym for the category of dominion structure of a pathogenic prion protein is identical to (lat. dominium), introduced earlier by Moore (1974). the nonpathogenic form. They differ in their tertiary Unlike Moore, Woese with co-authors (1990) did structure. Namely, pathogenic prion protein con- not suggest a Latin term for this category, which tains a higher percentage of the β-sheet on account represents a further argument supporting the of a lower frequency of its α-helix. Pathogenic prion accurately introduced term dominion. protein can occur via two routes – consequential to a Overall, the current classification system of the mutation of the prion-coding gene or upon digestion living beings requires modification. Differences of infected meat. Once in the cell, pathogenic prion between prions and the rest of acellular organisms protein binds the nonpathogenic protein forming are comparable to those between the nucleic-acid- a heterodimer, which causes a conformational containing acellular organisms and the cellular change of the nonpathogenic protein. Upon the life forms. Namely, three large groups of the living conformational change, a homodimer is formed, beings exist: acellular organisms without nucleic giving raise to two molecules of pathogenic protein. acid, acellular organisms with nucleic acid, and Two newly formed pathogenic prion proteins cellular organisms. Regarding these three groups bind the other two nonpathogenic prion proteins, as dominions would be logical; however, since the hereby increasing the ratio of pathogenic over the cellular organisms comprise an immense number nonpathogenic prion proteins in the cell. Once the of species, practical reasons speak in favour of the prions reach the critical concentration in the cell, five-dominion classification of the living beings: they begin forming rod-shaped aggregations, which Prionobiota (acellular organisms without nucleic accumulate in the infected , causing tissue acid), Virusobiota (acellular organisms with nucleic damage and host cell death (Prusiner, 2004). acid), Archaebiota (organisms with prokaryotic structure that show many differences in their Dominium Virusobiota biochemistry and in genome from all other forms of life), Bacteriobiota (typical prokaryotic organisms), The dominion of Virusobiota encompasses and Eukaryobiota (eukaryotic organisms). Domi- acellular organisms with nucleic acid content. nion names have been modified according to the Traditionally, viruses, viroids and viral satellites aforementioned nomenclature rules. are known as the acellular organisms with nucleic acid (Hull, 2009). I attributed a kingdom status to Dominium Prionobiota these groups, integrating all viral satellites within a common name virusoids. The names for these three The living beings without nucleic acid are ranked kingdoms have been assigned as aforementioned – within this dominion. Until now, they have been Virusoida, Viroida and Virusida. insufficiently studied, and only a modest number of species is known. Since majority of biologists do not Regnum Virusoida. In the course of researching regard prions as the living beings, their classification viroids, novel acellular organisms were discovered, has remained unedited. I have grouped all prions named viral satellites. A group of such organisms has into a single kingdom – Prionida. been named virusoides; therefore, I am proposing a derivative name of the kingdom based on the Regnum Prionida. Prions are infective protein group name. They are different from viroids by particles with an ability to multiply/replicate and structure and replication mode. Unlike viroid evolve. They live exclusively in eukaryotic cells RNA molecules, virusoid ones are smaller, single- and consist entirely of proteins without nucleic stranded and circular. Both virusoids and viroids acids. Prions are hereby deprived of any genetic lack a protein capsid (Strauss and Strauss, 2008). information in its classical sense. Genes necessary These organisms take part in pathogenesis of for prion replication are provided by the very host certain plant diseases. Viroids use innate cellular cell, named PrP (Prion Protein) genes (Prusiner, mechanisms to replicate, whereas virusoids replicate 2004). Healthy organisms comprise prions, which in the cytoplasm using cellular RNA-dependent are regarded as normal. They are located on the outer RNA-polymerase or RNA-polymerase coded by a Protistology · 225 helper virus. Therefore, their replication depends by virions as carriers of genetic information, can be on a co-infection with a helper virus (Strauss and DNA or RNA, but both types never occur within Strauss, 2008). the same virion. Certain viruses contain both Regnum Viroida. Viroids are subviral particles, DNA and RNA, however never synchronously, smaller than viruses and characterized by a simpler but exclusively in separate phases of their life cycle structure (Hadidi et al., 2003). These organisms are (Hull, 2009). built solely of a single-strain RNA with molecular weight of 50,000 – 200,000 Da. RNA molecules can Dominium Bacteriobiota be linear or circular. Although the viroid RNA is single-stranded, complementary base matching may Traditionally, bacteria are divided into Gram- occur between certain segments, hereby forming a positive and Gram-negative. The division has double-strain secondary structure (Hadidi et al., been based on staining techniques and reaction 2003). This gives the viroid molecule a hairpin shape. of the cell wall to this procedure. Gram-negative It contains 250–400 bases with high guanine-cytosine bacteria have an additional outer content. Viroids are sensitive to ribonuclease activity containing in its outer leaflet and resistant to organic solvents, high temperature, and phospholipids in the inner leaflet. Gram- deoxiribonucleases and proteases. Since such a positive bacteria lack such a membrane, but have a few nucleotides cannot carry substantial genetic thicker layer. This layer is responsible information, replication of the viroid RNA rema- for ability of Gram-positive bacteria to retain crystal ined elusive for quite a long time. Namely, in order violet dye in Gram staining protocol. In contrast, to replicate, acellular organisms need a virus-specific Gram-negative bacteria can not retain this stain; polymerase – an enzyme which requires RNA of instead they take up the red counterstain. This is roughly 300,000 Da, or at least 1000 nucleotides in not the only difference between these two groups length (Hadidi et al., 2003). Viroid RNA composed of bacteria, but it is certainly the most important of several hundred nucleotides is unable to synthe- one. It has been found that Gram-positive bacteria size the polymerase, and a helper virus assisting viro- are more susceptible to antibiotics than Gram- id reproduction has not been observed. Therefore, negative bacteria. Classification system proposed it was assumed that viroids used a DNA-dependant by Margulis and Schwartz (1988) relies directly on RNA polymerase to replicate and that their genome this demarcation, except for naming Gram-positive did not function as mRNA. However, it has currently bacteria and Gram-negative bacteria been determined that viroids replicate in the host cell . However, the first classification system nucleus by means of the host RNA polymerase II, based on these criteria was proposed by Gibbons using viroid RNA as a template (Hadidi et al., 2003). and Murray (1978). They divided the kingdom Pathogenic mechanism of these organisms remaine Procaryotae into three divisions: Gracilicutes (“cells unclear. It has been assumed that viroids interfere that have a rigid or semirigid cell wall, containing with regulation of the host cell’s gene expression peptidoglycan, and in which the Gram reaction or that they prevent correct introne excision and is negative”), Firmacutes (“cells that have a rigid subsequent splicing. Terminal consequence of the or semirigid cell wall containing peptidoglycan viroid presence in the cell is inability to express genes and in which the Gram reaction is positive”), required for normal cellular functions. The way of and (“cells that do not have a rigid or their release from the cell is insufficiently conceived, semirigid cell wall”). Margulis and Schwartz (1998) as well as their transmission paths and mechanisms proposed classification of bacteria comprised of of the host cell infection (Hadidi et al., 2003). nine groups; Tudge (2000), Garrity with co-authors Regnum Virusida. Viruses are basically built of (2001, 2003) and Black (2002) suggested later some two components – nucleic acid and protein sheath. modifications to this system. For example, in the Infective viral particle outside the cell is called internationally recognized manual on systematic virion. The dimensions of a virion vary within 28– (Garrity et al., 2001) as many as 23 200 nm range. Virions are said to be filterable, since bacterial groups (“phyla”) have been recognized. they are capable of passing through bacteriological Since 2001, the number of has increased up to filters. Size of a viral particle can be measured 70 including possible candidates with unculturable by electron microscopy, ultracentrifugation in representatives (e.g. Rappé and Giovannoni, 2003; gradients, ultrafiltration, and by means of ionizing Pace, 2009). The phylogenetic analysis of bacteria irradiation. The inner structure and symmetry of is very specific. Horizontal gene transfer represents viruses can be explored by electron microscopy or the biggest problem, and many authors (e.g. Gupta, X-ray structural analysis. Nucleic acids, contained 2001; Gupta and Griffiths, 2002; Ciccarelli et al., 226 · Stefan Luketa

2006) have been investigating this issue. hot springs, sulphur basins, thermal ocean cavities, Later, Cavalier-Smith (2006) proposed the and similar extreme habitats (Inc Icon Group division of the prokaryotic kingdom into two International, 2008). These bacteria are dominant subkingdoms – Negibacteria and Unibacteria. in many terrestrial neutral-to-basal hot springs with Within the subkingdom Negibacteria, three clades water temperature exceeding 60°C. Aquificida are were distinguished – Glidobacteria (Chlorobacte- autotrophic organisms, representing the primary ria, Hadobacteria and Cyanobacteria), Gracilicutes carbon fixators in their habitats (Inc Icon Group (Spirochaetae, , and International, 2008). ) and Eurybacteria (Selenobacteria, and Togobacteria). Within this parti- Regnum Fusobacterida. Only 32 species belong tion, Chlorobacteria, Hadobacteria, Planctobac- to this kingdom (Battistuzzi and Hedges, 2009). teria, Spirochaetae, Sphingobacteria, Proteobac- These bacteria are anaerobic, sessile and spindle- teria, Planctobacteria and Eurybacteria have been shaped. Frequently, cells have a swelling in the granted the level of within the subkingdom middle part (Samaranayake, 2006). Negibacteria. Subkingdom Unibacteria is comprised of two clades – Posibacteria (Endobacteria and Regnum Thermotogida. Thirty species belong ) and Archaebacteria (Euryarchaeota to this kingdom (Battistuzzi and Hedges, 2009), and Crenarchaeota), both given the status of a representing a small, coherent group both in phylum. morphological and physiological sense. All the This paper essentially supports the classification representatives are extreme thermophyles and proposed by Battistuzzi and Hedges (2009). Accor- inhabit exclusively the anaerobic niches. An outer ding to these authors, the Bacteria encloses the sheath-like membrane (“toga”) is specific for following five clades: , Hydrobacteria, Thermotogida (Oren and Papke, 2010). Fusobacteria, and . In the proposed megaclassification system these clades are Dominium Archaebiota granted the status of a kingdom within the dominion Bacteriobiota. Woese with co-authors (1990) postulated two genetically clearly separated kingdoms within the Regnum Terrabacterida. This prokaryotic king- domain Archaea – Crenarchaeota and Euryarch- dom comprises more than 6100 species (Battistuzzi aeota. Upon that, a third, clearly separated evolu- and Hedges, 2009). They have been given this tionary line was discerned within this dominion name since the majority of them are terrestrial. – Korarchaeota (Barns et al., 1996). Huber with In the course of evolution, they have adapted to co-authors (2002) discovered a fourth group – varying environmental stresses of the land habitats Nanoarchaeota. Nevertheless, the initial sectio- – atmospheric impacts, ultraviolet radiation, etc. ning into two kingdoms, Crenarchaeota and Eury- Some species among these organisms are capable archaeota, was corroborated by recent molecular of oxidative photosynthesis. Both Gram-positive analysis (Elkins et al., 2008), which is acknowledged and Gram-negative representatives have been here. found within this kingdom (Battistuzzi and Hedges, 2009). Regnum Euryarchaeida. This kingdom entails relatively few species which cells come in largely Regnum Hydrobacterida. This bacterial king- differing forms. It has been determined that the shape dom contains over 3200 species (Battistuzzi and is a relatively stable feature and it could be of great Hedges, 2009). Hydrobacterida are the exceptionally importance for correct determination. Within this ancient and divergent prokaryotic group, difficult group, eight clearly defined morphological forms of for any common characterization. The cells are found: rod-like, coccoid, irregularly coccoid, clade is also classified within this group, characte- lanceted, spiral, discoid, triangle-shaped and cubical. rized by long, spiral cellular shape. Many of the Certain representatives are Gram-positive, and species are free-living, some are commensalls, others are Gram-negative (Garrity and Holt, 2001). and a large number of spirochaetes are parasites Physiologically, the group is highly heterogeneous. (Battistuzzi and Hedges, 2009). Metanogenic forms occur as well as extreme halophile and hyperthermophile species. Some of Regnum Aquificida. This kingdom encompasses these organisms inhabit niches with temperatures as 22 species of extremophylic bacteria (Battistuzzi high as 100°C (Pommerville, 2010). and Hedges, 2009). So far, they have been found in Regnum Crenarchaeida. This kingdom consists Protistology · 227 of hyperthermophilic prokaryotic organisms, which amoeboid shape (Saville-Kent, 1880). The jelly-like inhabit warm areas, rich in sulphur. For these mass resembles mesoglea of the and reasons, they are mostly found in the ocean depths in the transformation of the collar cells into amoeboid vicinity of geothermal releases. Most representatives cells may explain the origin of amoeboid cells in the are optimally adapted to 80°C water temperature mesoglea (Hadži, 1970). At first, the critics (Pommerville, 2010). have taken the fact that Proterospongia haeckeli was a freshwater organism as a main argument against Dominium Eukaryobiota this theory, since the sponges are marine animals and therefore could not stem from the freshwater Eukaryotes are divided into several clades: life forms. This argument was dismissed when the , Opisthokonta, , Archae- marine representatives of these organisms were plastida, and Excavata (Adl et al., found (Nielsen, 2001). Shortly after this discovery, 2005). The clade Opisthokonta encompasses ani- Brusca and Brusca (2003) formulated a hypothesis mals and fungi, as well as a number of smaller clades: that perceived Choanoflagellata as the drastically Choanoflagellata, Mesomycetozoa, , reduced sponges. Corallochytrea, and Adjacent to Opisthokonta, within the Unikonta (Adl et al., 2005). Small group of protozoans supergroup, another group was distinguished, which named Choanoflagellata has been classified into was composed of amoeboid protists and slime the Zooflagellata group by numerous authors in molds – Amoebozoa. Traditionally all protists with the second half of the 20th century (Margulis and pseudopods were treated together as the Rhizopoda Schwartz, 1988, 1998; Buck, 1990). However, some or Sarcodina (e.g. Goldfuss, 1817; Honigberg et authors have noted that this is an unusual group of al., 1964; Levine et al., 1980). Cavalier-Smith protozoans that are associated with different groups (1993) proposed seven parvikingdoms within the of eukaryotic microorganisms. Because of that, this infrakingdom Neozoa. The representatives of group is paid more attention here than the other three of these seven parvikingdoms are protists groups of eukaryotes. with : Actinopoda, Neosarcodina and Sleigh with co-authors (1984) proposed alloca- Entamoebia. Cavalier-Smith (1998) proposed four tion of Choanoflagellata into a separate group. phyla within the infrakingdom Sarcomastigota. Taylor (1976), Lee (1980) and Sze (1986) postulated The three representative groups of these phyla are a hypothesis that these organisms were related amoeboid protists: , and to the chrysophytes. Modern genetic research Amoebozoa. The monophyly of Amoebozoa has showed that Choanoflagellata are a specific group, been proposed by many authors (e.g. Baldauf et al., phylogenetically close to the representatives of 2000; Bolivar et al., 2001; Milyutina et al., 2001; Filasterea and Corallochytrea. Cladification system Arisue et al., 2002; Bapteste et al., 2002; Forget proposed by Adl with co-authors (2005) groups et al., 2002; Baldauf, 2003; Fahrni et al., 2003; Choanoflagellata together with Metazoa, Fungi Keeling, 2004; Nikolaev et al., 2004; Adl et al., and Mesomycetozoa. According to these authors, 2005, 2012). representatives of Filasterea and Corallochytrea are Cavalier-Smith (2003) stated that ranked into Mesomycetozoa. were phylogenetically closer to Bikonta rather than Already in late 19th century, Metschnikoff to Unikonta. This view was supported in several (1886) postulated that sponges originated from subsequent papers (Berney et al., 2004; Nikolaev et Choanoflagellata. This hypothesis based on the al., 2004). Cavalier-Smith and Chao (2003) postu- great structural resemblance of Choanoflagellata lated that Unikonta was a paraphyletic group, and choanocytes, which form the lining of the branching near the base of the evolutionary tree sponge paragastral cavity. Such point of view of all eukaryotes. The first phase in the evolution was corroborated by the fact that the cells with a of eukaryotes had been the formation of a single and a collar were found only in sponges flagellum, which contained microtubules inside and Choanoflagellata. Second argument which its membrane sheath. It was only later that the supported this hypothesis was the existence of mitochondria had been formed endosymbiotically. the species Proterospongia haeckeli, discovered by Two groups have stemmed from these early euka- Saville-Kent (1880). Colonies of this species are ryotic organisms – one with the flat mitochondrial 100 µm-wide spheres, build of jelly-like mass with cristae (Opisthokonta) and the other with the tubular collar cells protruding outwards. Upon a phase of cristae (Anterokonta). The Anterokonta group intensive feeding, some of these cells move into gave rise to biflagellated eukaryotes (Bikonta). Of the inner part of the jelly-like mass and attain an all Anterokonta, only organisms from the clade 228 · Stefan Luketa

Amoebozoa have persisted to the present days. between Chromophyta and Alveolata; later on, Organisms similar to the present Apusozoa are the close relationships of these two groups were thought to be the intermediate forms between confirmed by other researchers (Patterson, 1999b; Anterokonta and Bikonta. is a polyphyletic Taylor, 1999; Baldauf, 2003; Keeling, 2004; Harper assemblage of diverse unrelated groups, all of which et al., 2005). are concentrated in the part of the global Representatives of the clade eukaryotic tree. According to Kim and co-authors had predecessors that displayed symbiotic rela- (2006), Apusozoa is phylogenetically close to tionship with a prokaryotic organism capable of Opisthokonta. photosynthesis (Cavalier-Smith, 1982, 2003). Within the Rhizaria clade, the morphologically Contemporary organisms of this clade have photo- heterogeneous representatives have been grouped synthetically active plastids bordered by two together basing on their molecular features. The membranes – the inner one has originated from Rhizaria group was proposed by Cavalier-Smith the photosynthetic and the outer (2002b). In addition, numerous more recent surveys membrane has stemmed from a phagocytotic vesicle (Baldauf, 2003; Keeling, 2004; Nikolaev et al., that once had trapped the prokaryote during the 2004) confirmed that it was a monophyletic group. phagocytotic process (Cavalier-Smith, 2003). All Burki with co-authors (2007) and Hackett with photosynthetic organisms with plastids outlined co-authors (2007) stated that Rhizaria has close with three membranes have come into the cell phylogenetic relationships with Chromalveolata. by symbiotically engaging with representatives of Baldauf (2008) named this group the “RAS-group” Archaeplastida (Cavalier-Smith, 2002b; Keeling, (Rhizaria-Alveolatae-Stramenopilae group). This 2004; Adl et al., 2005). group united all photoautotrophic eukaryotic According to Cavalier-Smith (2002a), the organisms. Cavalier-Smith (2002b) distinguished first eukaryotic organisms were phagotrophs by two clades within this group: (Foraminifera nutrition and amoeboflagellates by morphology. and ) and Cercozoa (Cercomonada, They either scaled or slided along solid substrates Haplosporidia and ). Cavalier- and have developed into two major evolutionary Smith and Chao (2003) supported this opinion, lines – Unikonta and Bikonta. These two groups whereas Nikolaev with co-authors (2004) did not differ primarily by their cytoskeleton structure and endorse such ranking. the development of the flagellar apparatus. Roger Patterson (1999b) was among the first researchers and Simpson (2009) acknowledged the division of who observed close phylogenetic relationships eukaryotes into Unikonta and Bikonta. However, between the , Euglenophyta, and Cavalier-Smith (2010b) later on set out an opinion Heterolobosea. He pointed out the swollen or about Bikonta being a paraphyletic group. In the discoid form of the mitochondrial cristae as well present paper, these two groups are considered as as major resemblance in the structure of their two subdominions. flagellar apparatus. The following analyses showed a monophyletic nature of Subdominium Unikonta this group (Simpson et al., 2002, 2006; Simpson, 2003; Breglia et al., 2007; Leander et al., 2007; Subdominium Unikonta was proposed by Rodriguez-Ezpeleta et al., 2007; Burki et al., 2008). Cavalier-Smith (2002a). Adl with co-authors Later on, some other related groups were added to (2012) proposed name for this group. the one discussed earlier, and all together they were Representatives of this group are characterized by united within the taxon Excavata (Cavalier-Smith, mono-flagellar cells. In certain groups, flagellum 2002b). has become reduced in the course of evolution. Pascher with co-authors (1925) grouped a large Most representatives of Unikonta have a single number of photosynthetic and non-photosyn- centriole in their protoplast. Some species have two thetic organisms into an informal taxon, named centrioles, but their origin is different from the one Chromophyta, basing on the characteristics of supposed for . Presumably, this was a result their flagellar apparatus. It was Cristensen (1962) of convergent evolution (Cavalier-Smith, 2002a). I who ranked this group as a division Chromophyta. demarcate four kingdoms within the subdominion Cavalier-Smith (1989b) ranked this group as a Unikonta: Amoebozoida, Apusozoida, Fungida kingdom and included all the current representatives and Animalioida. In the course of evolution of this of Heterokontae and in it. Cavalier-Smith subdominion, Amoebozoida have branched earlier (1981) was the first to observe phylogenetic relations than Apusozoida (Kim et al., 2006). After branching Protistology · 229

Subregnum Amoebozoides. Representatives of this subkingdom move by means of pseudopodia, usually of the lobopodia type. Some flagellated representatives of this group are also known (Smirnov et al., 2011). Mitochondrial cristae are tubular, often branched. Certain species are characterized by a locomotory stage propelled by a single flagellum, stemming from a single basal body (Adl et al., 2005). Subregnum Breviatides. anathema (Minge et al., 2009) and Subulatomonas tetraspora (Katz et al., 2011) are two unicellular, free-living anaerobic species which form this group. They possess amoeboid cell body and a single flagellum. One basal body presents a base for the flagellum; therefore it has been named flagellatic. Next to it, at least one other basal body exists, named non . This second basal body was observed only in (Roger and Simpson, 2009). Fig. 1. Phylogenetic relationships between king- B. anathema is considered to be amitochondriate doms of the subdominion Unikonta. Phylogenetic B. anathema tree was constructed based on the conclusions (lacking typical aerobic mitochondria), but it drawn in the present paper. harbors a kind of degenerate mitochondria-like organelles bounded by double membranes. of the above-mentioned taxa, a bifurcation took Regnum Apusozoida. Doflein (1916) established a place which gave rise to two kingdoms: Fungida and new family Spironemidae. Foissner with co-authors Animalioida (Fig. 1). (1988) proposed allocation of this family into a new protistan phylum, the . The order Regnum Amoebozoida. This kingdom contains was established by Karpov and heterotrophic amoeboid protists. Most of them Mylnikov (1989) and originally included two genera are unicellular inhabitants of the soil and aquatic – and Amastigomonas. Cavalier-Smith . There are also multicellular (as certain (2002b) created the phylum Apusozoa including stages of life cycle) and multinuclear macroscopic these two genera plus the representatives of phylum representatives. Walker with co-authors (2006) Hemimastigophora. Subsequently, many new taxa described an amoeboflagellate Breviata anathema, have been described (e.g. Cavalier-Smith et al., previously thought to be an amoebozoan (“Mastig- 2008; Cavalier-Smith and Chao, 2010; Glücksman invertens”). Minge with co-authors (2009) et al., 2011; Yabuki et al., 2012). This kingdom observed phylogenetic relationships between comprises few free-living, heterotrophic species. Breviata and Amoebozoa. Shipunov (2009) divided They move by means of the two or more flagella. the superphylum Sarcobionta into two phyla: Apusozoida are cosmopolitan and can be found both Amoebozoa and Breviatozoa (included only in marine and fresh water ecosystems, as well as in Breviata). Katz with co-authors (2011) described terrestrial habitats. Nevertheless, as they occur at a new species Subulatomonas tetraspora and deter- low population densities, they are often overlooked mined its close phylogenetic relationship to Breviata in the samples (Karpov, 2007/8). anathema. Although the exact place of the genera Subulatomonas and Breviata within the dominion Regnum Fungida. I have ranked three subking- Eukaryota is not known (Katz et al., 2011), I doms within the kingdom Fungida on the basis of propose grouping them together with Amoebozoa their close phylogenetic relations. Fungida consists because the morphology of representatives of these of unicellular, filamentous and even very large two groups is similar. I propose two subkingdoms multicellular organisms of a relatively complex within the kingdom Amoebozoida: Amoebozoides structure. The most numerous subkingdom is (all representatives of the group Amoebozoa) and Fungides, comprised of fungi, whereas the other Breviatides (Breviata anathema and Subulatomonas two subkingdoms have so far remained scarce in tetraspora). species. They do not share many common features 230 · Stefan Luketa and the group was formed foremost based on the Tabrizi et al., 2008). The group Mesomycetozoa close phylogenetic relationships, that have been was introduced into the literature as a DRIP-clade confirmed by contemporary studies. Two other by Ragan with co-authors (1996). Ichthyosporea, groups, which I have ranked as subkingdoms, are Corallochytrea and Filasterea, together with the closely related to Fungi – Nucleariides (Steenkamp group Nucleariida were ranked into Mesomycetozoa et al., 2006; Baldauf, 2008) and Microsporides (Adl et al., 2005). In this paper, the Nucleariida group (Fischer and Palmer, 2005; Vossbrinck and Debrun- is ranked into the kingdom Fungida, in accordance ner-Vossbrinck, 2005; Gill and Fast, 2006; Liu et with the previously cited studies. Due to many al., 2006; Lee et al., 2008). common features of sponges and Choanoflagellata Subregnum Nucleariides. This subkingdom and the major differences between sponges and encompasses a limited number of species characte- the rest of the animal kingdom species, I proposes rized by their amoeboid appearance. Body of these grouping of the clades Porifera and Choanoflagellatea organisms is spherical or plate-shaped and filopodia into a common subkingdom named Choanozoides. protrude from their surface (Minelli, 2009). Nuclea- Based on the presented perception and in agreement riides have flat mitochondrial cristae (Adl et al., with the earlier defined nomenclature, I propose a 2005, 2012; Minelli, 2009). separation of the following three subkingdoms within Subregnum Microsporides. Approximately 1200 the kingdom Animalioida – Mesomycetozoides, unicellular, spore-forming species constitute this Choanozoides and Animalioides. subkingdom (Lee et al., 2008). They are intracellular Subregnum Mesomycetozoides. The representa- animal parasites. Most of the known species are tives of this group do not share many common parasites of insects and fish, but it is assumed that distinctive characteristics. They live as parasites in they can infect members of virtually all animal the body of fish, birds, mammals, and crustaceans. taxa as well. Many species have their specific host, Some species of this subkingdom are free-living which makes Microsporides the good candidates forms. Their mitochondrial cristae are usually plate- for biological control of pest insects and ticks. like (Adl et al., 2005). Shape of the body is mostly irregular in certain life Subregnum Choanozoides. This subkingdom cycle stages. The spores contain a mononuclear consists of more than 5000 species. They are mostly or bi-nuclear (diplokaria) sporoplasm, extrusion unicellular or colonial organisms, which move by apparatus, and a polar tube. The spores are formed means of a single flagellum. Sessile forms have also from a single cell (Lee et al., 2008). been distinguished within this group. Mitochondrial Subregnum Fungides. Fungi are a highly specific cristae are plate-like. Most Choanozoides feed group of spore-forming eukaryotic organisms. as phagotrophs and a few species have green Fungal cells are equipped with a cell wall during chromoplasts that enable them to photosynthesize. most, if not all phases of the life cycle (Ranković, The protoplast contains a single nucleus. All 1994). Fungi are heterotrophs and they absorb their representatives are free-living forms; they inhabit food. They are the most widely spread organisms on freshwater and marine ecosystems. Earth. To the present day, roughly 100,000 species Subregnum Animalioides. These organisms have been defined and existence of about 1.5 million are multicellular and heterotrophic; they usually of species was assumed (Ranković, 1994). Vegetative swallow their food and digest it inside the body. body of the fungi is called somatic body, occurring Most representatives of this kingdom are mobile as cellular, non-micellic and micellic entities. Too organisms. They occur in all biogeographic areas, tiny or well hidden (due to their mode of life) soma- occupying various ecological niches and displaying tic body of the fungi is difficult to observe in nature. highly variable morphological and metabolic Fungi can also metamorphose into specific large features. They do not have a cell wall; molecules of macroscopic formations. collagen, proteoglycan and adhesive glycoproteins are found in their extracellular matrix. Their cells can Regnum Animalioida. I have ranked three move during certain ontogenic developmental stages. subkingdoms within this kingdom on the base of Common characteristics have been observed in the close phylogenetic relations. This highly divergent course of development of a zygote. Namely, in the group consists of unicellular, colonial and even very course of cleavage, a blastula is formed, to become large multicellular organisms of a complex structure. gastrula by germ layer formation in the most. Animals The most numerous group is Metazoa, which I have have a gastric cavity with one or two openings. In case grouped into a subkingdom Animalidae, with related of two openings, one is frontal – the mouth, and the representatives of Ichthyosporea, Corallochytrea, other opening is situated posteriorly and it is called Filasterea and Choanoflagellatea (Shalchian- anal opening. Protistology · 231

Subdominium Bikonta Names have been modified in agreement with the previously defined nomenclature. This taxonomic group was proposed by Cavalier- Subregnum Archaeplastides. This is a rather large Smith (2002b). The main feature of the group is group of organisms with differing morphological the presence of bi-flagellated cells. Even though physiological, ultrastructural and ecological fea- certain representatives do not have a flagellum at tures. The most important common characteristic all, the concept suggests that the predecessors of of these organisms and a base for their monophyletic this group had two flagella. Bikonta are divided into ranking is the presence of plastids with two mem- two groups: Excavata and the group that consists of branes. This fact implies that the plastids of these Archaeplasida, and , organisms evolved from symbiotic blue green Stramenopiles, Alveolata and Rhizaria (Roger and prokaryotes (Adl et al., 2005, 2012). Plastids of Simpson, 2009). The second group was named all other groups are surrounded by three or four Corticates (Cavalier-Smith, 2010b). The clade membranes. that consists of Cryptomonads and Haptophytes, Subregnum Cryptophytides. This subkingdom together with a few minor groups, was named consists of autotrophic, phagotrophic and hetero- Hacrobia (Okamoto et al., 2009). However, the most trophic organisms, which inhabit both marine and recent phylogenomic analysis doubts the common freshwater environment. They are all unicellular origin of haptophytes and cryptophytes (Burki et al., organisms, mobile by the means of flagella or 2012). In this paper, I have given the Excavata and pseudopodia and have ejectisomes. Corticates a status of kingdom, and I have corrected Subregnum Haptophytides. This subkingdom their names in agreement with the above defined consists of both autotrophic and phagotrophic nomenclature – Excavatida and Corticatida. organisms. They are unicellular, colonial or filamen- tous. Motile cells often possess a haptonema. All the Regnum Excavatida. The kingdom Excavatida is representatives are free-living forms; they inhabit relatively small and consists of organisms that differ freshwater and marine ecosystems. from each other in their morphology, ultrastructure Subregnum Rhizarides. This subkingdom en- and ecology. Therefore, autotrophic, heterotrophic, compasses a large number of unicellular species. parasitic and symbiotic species are found among These are ultrastructurally complex organisms. Excavata. This kingdom includes some parasites Morphologically, they differ from each other; still, that are of a high medical importance, such as most species have amoeboid appearance and move representatives of genera , Leishmania, by the means of filopodia, reticulopodia, lobopodia Trichomonas and Trypanosoma (Adl et al., 2005, and axopodia (Adl et al., 2005). Many species form 2012). Since many species do not have typical shells around their protoplasm that largely vary in mitochondria, some biologists considered Excavata shape and chemical composition; they can be highly as a very old eukaryotic group that had appeared complex in structure. Due to their hard shells, many before the origin of these endosymbiosis-derived of the species are known as microfossils. Almost organelles. Cavalier-Smith (1983) postulated a all representatives have mitochondria with tubular hypothesis of Excavata being a common predecessor cristae. for all other eukaryotes. Plesiomorphic characte- Subregnum Stramenopilides. Many species of ristic of these organisms is the presence of an this group are mobile cells with two flagella (Adl et excavate type , that has been secondarily al., 2005, 2012). The flagella are usually lost in certain representatives (Adl et al., 2005, – one flagellum carries numerous mastigonemes 2012). Still, most of the species contain highly and the other one does not. Mitochondria are altered mitochondria (Simpson, 2003). Most of the characterized by tubular cristae (Adl et al., 2005, representatives with mitochondria are characterized 2012). Most representatives are autotrophic, and by tubular or discoid cristae, whereas fewer have fewer are heterotrophic. laminar cristae. Most of the representatives have Subregnum Alveolatides. This group typically has two, four or more flagella (Simpson, 2003). cortical alveoles, although they can be secondarily absent. Cortical alveoles are flattened vesicles, Regnum Corticatida. Basing on the phylogenetic lined up to form a continuous layer beneath the tree proposed by Burki and co-authors (2012) (Fig. 2), cell membrane. Mitochondria are characterized by I propose dividing this kingdom into six subkingdoms: tubular or ampular cristae (Adl et al., 2005, 2012). Archaeplastides, Cryptophytides, Haptophytides, Both heterotrophs and are found among Rhizarides, Stramenopilides and Alveolatides. these organisms. 232 · Stefan Luketa

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Address for correspondence: Stefan Luketa. Department of Biology and Ecology, Faculty of Science, Uni- versity of Novi Sad, Trg Dositeja Obradovića 2, 21000 Novi Sad, Serbia; e-mail: [email protected]