By Yu. POLYANSKY, Corresponding Member of the RAS, President of the Society of Protozoology, RAS,

and T. BEYER, Dr . Sc. (Biol.), Leading Researcher, Institute of Cytology, RAS

The study of the objects of protozoology, the unicellular organisms or protozoa, was not possible until the microscope was invented. The honour of detection of protozoa belongs indeed to the Dutch microscopist of the 17th century Antoni van Leeuwenhoek, who can be justly regarded as the father of protozoology In 1675 he first observed a free living protozoon (presumably, a ciliate of the genus Vorticella), and a year before he saw coccidian oocysts in the rabbit liver.

For many years since Leeuwenhoek's days an ever-increasing number of unicellular living creatures were described and given a common name - "infusoria" and a lot of ridiculous descriptions of their structure regularly appeared and were discussed in literature.

Of great importance for a further understanding of the protozoa was the creation of the cell theory, first clearly expounded by T. Schwann in 1838-1839, according to which the cell was regarded as the unit of organization of the majority of plants and animals.

The biosphere around us is the home of large groups of organized living creatures that form numerous ecological systems. The most ancient are the prokaryotic microorganisms, whose hereditary information is written in the DNA, although not yet organized within a cell nucleus. The prokaryotic organisms may have appeared about four billion years ago. And two billion years passed before they could give the origin of the eukaryotic cell, provided with an organized nucleus. Compared to the prokaryotic organization, the eukaryotic one appeared highly progressive, building a real basis for emergence of numerous members of plants, animals and protists ^* during the 1.5-2 bln years of evolution that followed.

It is thought that the eukaryotic level of life could be presumably established on Earth in the form of unicellular organisms, or protists. It has been commonly believed that the present unicellular organisms are nothing but remains of that period when the entire life on Earth was still unicellular, and that part of these organisms may have persisted up to now, being able to adapt to life in certain ecological niches. Such a concept is now to be revised.

Analysis of the modem relevant data allows to think that the present-day protozoa are not merely the relics of the Earth remote history, partly conserved. These are peculiar organisms at a special level of organization of the living matter, and playing a significant part in the biosphere; however, these organisms are not yet completely investigated and still remain poorly understood.

The protozoa play a significant role in the Earth-crust creation. The World Ocean abounds in protozoa with mineral skeletons (lime, silicon), which after dying off settle down to the bottom of the oceans and seas to produce eventually the ooze layers many meters thick. Then, as a result of tectonic processes, the bottom becomes a dry land, and the accumulated sediments turn into rudimentary rocks to make up the main bulk of the surface layers in the Earth-crust: limestone, chalk, marble. Thus, the protozoa obviously take part in creating our Planet's picture, being an important link in chemical elements rotation (calcium, silicon, magnesium, strontium). The involvement of protozoa in this process was convincingly demonstrate by V. Xfemadsky, the founder of biosphere science.

The role of the protozoa as an appreciable component of the plankton and, partly of microbenthos (the bottom residing organisms) is still poorly known. Again, the protozoa are involved in many food chains between bacteria and zooplanktonic organisms. Without taking all this into consideration

^* The protists, or unicellular eukaryotic organisms, are comprised of two main subdivisions: unicellular algae (autotrophs, provided with chlorophyll), and protozoa (unicellular, heterotrophs), which are the subject of the present paper. (Author's Note).

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(alas, this often happens in some hydrobiological studies), the eventual pattern of reservoir ecosystems will appear not true. A vast evidence recently provided in this country confirmed the importance of protozoa for the World Ocean and other water systems. For example, the studies carried out in the upper part of the Volga, in the Rybinsk reservoir, have demonstrated that the food chains largely depend not only on the ciliates, but also on the flagellates, which were previously almost totally ignored by investigators (Institute of Inland Waters, Russian Academy of Sciences, Borok).

Another practical utilization of free-living protozoa is the possibility to obtain their abundant protein mass. The number of methods have been elaborated for growing protozoa in culture, where they can divide one to several times a day to give a rapid augmentation of biomass. This matter may serve a good food for fry, or some kind of protein additions to animal and human food (Institute of Biophysics, Siberian Branch of the RAS, Krasnoyarsk).

Studies of the fauna of free-living freshwater and marine protozoa yielded much new knowledge of theoretical and practical value. These organisms turned out to be active in creation of soil, and in sewage purification by facilitating the disintegration of organic substances. Due to their high sensitivity, even to the slightest changes in the environment, these protozoa are good indicators of water conditions, in particular, of the degree of water pollution (St. Petersburg Pedagogical University, Moscow University). A great number of protozoa have evolved to become parasites, i.e., to live at the expense of other organisms that serve as their hosts. This peculiar way of development is posing a lot of questions concerned with host-parasite relationships. Many parasitic protozoa are pathogenic for humans, animals, birds, fishes, insects.

According to the WHO records, malaria is one of the most widely spread human diseases. Hundreds of thousands of malaria cases are registered annually, a lot of them being lethal cases. The agents of human malaria (it also occurs in birds and reptiles) are protozoan parasites of the class Sporozoa, having complex life cycles, comprising alternation of hosts: intermediate (man) and final (mosquitoes of the genus Anopheles) ones, resp. The mosquito-vector takes blood from an infected vertebrate (man, bird, reptile) and simultaneously injects its own saliva containing infective stages (sporozoites).

As early as in the 1940s, the Russian scientists B. N. Nikolaev, who studied one of the four species of human malaria parasites, revealed two subspecies of Plasmodium vivax, differing in the duration of incubation period, i.e., the time before patency becomes obvious in the patient. This pioneering discovery allowed, thereafter, to establish in the life cycles of Plasmodium species some earlier unknown developmental stages (later recognized as "hypnozoites") able to persist in the patient's liver for an indefinite time. The hypnozoites proved to be responsible for numerous cases of remote relapses characteristic of the tertian fever, of P. vivax -malaria, which was a many years puzzle for scientists all over the world.

In their studies in the field of malaria research Sh.D. Moshkovsky and A.Ya. Lysenko (Moscow) demonstrated that sporozoites of P. vivax differ in the duration of incubation period of their progeny This character is controlled presumably by

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special genes. When a mosquito bites a man it injects its saliva, containing a mixture of sporozoites of two different groups: the tachy- and bradysporozoites. The former starts to develop immediately causing disease in a short time, whereas the latter exhibits a long incubation time, as if after some hibernation. Realization of the above biological peculiarities of the pathogen allowed to find new approaches and means of malaria control.

In animal husbandry and poultry breeding enormous damage is caused by the Coccidia, i.e. organisms systematically close to the malaria parasites. In the warm regions cattle often succumb to diseases caused by blood parasites: piroplasmids and flagellates. Protozoologists from Moscow and St. Petersburg have much contributed to detailed studies on babesia and theilieria often causing heavy fever in cattle as well as on trypanosomes and lieshmania, the latter causing cutaneous and visceral diseases in man and animals.

To prevent or control protozoan diseases, it is important not only to recognize the pathogenes but also to study details of their structure and host-parasite relationship. This proved to be a truly delicate work to be carried out at the cellular, subcellular and molecular levels (Institute of Cytology, RAS, Institute of Evolutionary Physiology and Biochemistry RAS , Institute of Plant Protection, St. Petersburg; Moscow State University).

Some parasitic cells (e.g., in the Coccidid) appeared to be much more complex than any typical eukaryotic cell. They have an additional complex of special organelles, which not only favor the parasite's penetration into the host cell but also largely contribute to its intracellular growth and development. The intracellular parasites do not kill their host cells at once but modify them to make eventually a suitable habitat. In result, a temporarily dynamic hostparasite interaction is established, with preponderance favor of the parasite.

Biochemical studies allowed to find out that the protozoan parasites use selectively substances of the host cell for their own needs. The knowledge of the molecular and biochemical mechanisms, underlying the intracellular parasitism, opened up new possibilities for synthesizing certain active anticoccidian drugs, such as coccidine, iramine, butoxol, chimcoccid-7, etc., which are actively involved in coccidial control in animal and poultry farms.

Of much importance are recent studies of protozoologists from St. Petersburg on fish parasites (myxosporidia) and parasites of insects (microsporidia; these parasites being presumably dangerous also for man). Newly proposed systems of these pathogenes are largely based on detailed knowledge of their structure and metabolism, in addition to patterns of their ecology This enabled new proposals to be made on prevention and control of the above protozoa.

On describing the diversity and multiplicity of protozoa (almost 100,000 species have been investigated), their different habitats and complex life cycles, it is worth emphasizing a close association between protozoology and some other branches of biological sciences, such as zoology and botany, cell biology, including cytology and molecular biology. Protozoology it closely associated with problems of genetics and evolutionary theory, but solves these questions at a qualitatively peculiar level, which obviously differs from that in the multicellular organism (Metazoa).

It has been established that the morpho-physiological regularities of protozoan evolution are largely determined by the polymerization phenomenon, which means a progressive increase in the number of cell components, whereas the opposite process (oligomerization) is ruling the evolution of Metazoa. These important theoretical elaborations, started still by V.A. Dogies, the world recognized Russian protozoologist, were extended to include studies on the evolution of the nucleus apparatus in protozoa, and a recently advanced theory of nuclear dualism (Institute of Cytology, RAS).

This remarkable biological phenomenon had been found, in particular, in ciliates and is presumably associated with evolutionary peculiarities of the apparatus of storage, multiplication and materialization of hereditary information (DNA) represented by two morphologically distinct nuclei: macro- and micronuclei. The former is the storage of hereditary information throughout the number of asexual reproduction in ciliates, whereas the former assumes the function of materialization of hereditary information via the synthesis of information RNA and proteins.

Close to these studies is another series of experiments performed in St. Petersburg University A peculiar kind of symbiosis was established between bacteria (prokaryotic organisms) and ciliates of the genus Paramecium. This allowed to elaborate an original "symbiotic method" useful for studying the genetic apparatus in protozoa.

Geneticists-protozoologists of St. Petersburg contributed a good deal to the problem of variation in protozoa. Experimental evidence has been provided on the existence and importance of long-lasting modifications in the ciliated associated with their adaptation to the environment. These modifications arise as a response to some new environmental conditions to persist throughout the number of asexual generation, but not affecting the genome molecular structure.

Another interesting problem is epigenomic variation, i.e., an inheritable form also not associated with the molecular transformation of genome, but merely related to the genome expression. In these studies methods of "microsurgery" were employed for transplantation of cell nuclei. This technique allowed also to demonstrate the key role of the nucleus in heredity, followed throughout the asexual reproduction in protozoa. Simultaneously, some previously unknown phenomena were detected: nuclear interaction in heterokaryons (cells with two or more nuclei); nuclear-cytoplasmic, incompatibility; hereditary instability (Institute of Cytology RAS).

The facts and regularities, reported above, may be of much interest not only for studying the protozoa, but also for working out some general problems of genetics, since they are unveiling ever new mysteries of development of the living matter and contribute much to gaining a better insight into the evolution processes.

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