**Principle of Conservatism of Cellular Structures as the Basis for Construction of the Multikingdom System of the Organic Word**

Anatoliy L. Drozdov

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.68562

#### **Abstract**

This chapter describes the history of megasystematics (taxonomy of higher taxons) from Karl Linnaeus till the present day. Nowadays, the Whittaker's five-kingdom system of cellular organisms is the best known. This system has made monophyletic the kingdoms of plants, fungi, and animals but increased the heterogeneity of the kingdom Protoctista. There is one of the qualitative peculiarities of the subcellular level of the organization of living systems, which has been named "the principle of conservatism." We offer the multikingdom system of cellular organisms, based on this principle. In many ways, it can be done based on intuition. We promote the concept of three branches of cellular organisms that is accepted in megasystematics. It is proposed to give these branches of organic word the rank of domains Archaea, Bacteria, and Eucarya. The Empire Cellulata is divided into three domains, which, in turn, are divided into several kingdoms. Our system contains 26 kingdoms.

**Keywords:** history of megasystematics, multikingdom phylema, the most primitive eukaryotes

## **1. Introduction**

The history of megasystematics (taxonomy of higher taxons) dates back to the eighteenth century when a classification system of the living beings was created by the father of systematics (or taxonomy) Karl Linnaeus, which comprised two kingdoms—Vegetabilia and Animalia

(**Figure 1**). In the nineteenth century, the kingdom of fungi (Mycetoideum), on the one hand, and the kingdom of Protista or Protoctista, on the other, uniting unicellular or most of the lower organisms, were erected; however, most biologists continued adhering to the two-kingdom system.

The system of R.H. Whittaker is the most frequently adopted. He recognizes in his later work [1–4] the prokaryote as a kingdom Monera and divides the eukaryotes into three kingdoms higher kingdoms of plants, fungi, and animals, which as three stocks were transferred by him from the lower kingdom of Protista (**Figure 2**).

In that classification scheme, red and brown algae were placed near the base of the stock of plants, green algae were placed both in the protist kingdom (apparently, unicellular forms) and at the base of the plant kingdom, whereas myxomycetes were positioned near the base of the stock of fungi. This system is the most popular and in common use till date. The frequently adopted is Whittaker's five-kingdom system of cellular organisms modified by Lynn Margulis [5–8]. She thus made monophyletic the kingdoms of plants, fungi, and animals but increased the heterogeneity of the kingdom Protoctista. L. Margulis herself frankly admitted that "the

**Figure 1.** Two kingdoms of K. Linnaeus system (from Drozdov [5]).

Principle of Conservatism of Cellular Structures as the Basis for Construction of the Multikingdom... http://dx.doi.org/10.5772/intechopen.68562 5

**Figure 2.** Five kingdoms of R. Whittaker system (from Drozdov [5]).

protoctist kingdom becomes looking as if it were a dump." From these words of L. Margulis, it unambiguously follows that all schemes with few eukaryotic kingdoms (1–4) will err toward inadmissible polyphyly, as is confirmed by contemporary cytological and, especially, molecular biology data.

#### **2. History of megasystematics**

(**Figure 1**). In the nineteenth century, the kingdom of fungi (Mycetoideum), on the one hand, and the kingdom of Protista or Protoctista, on the other, uniting unicellular or most of the lower organisms, were erected; however, most biologists continued adhering to the two-king-

The system of R.H. Whittaker is the most frequently adopted. He recognizes in his later work [1–4] the prokaryote as a kingdom Monera and divides the eukaryotes into three kingdoms higher kingdoms of plants, fungi, and animals, which as three stocks were transferred by him

In that classification scheme, red and brown algae were placed near the base of the stock of plants, green algae were placed both in the protist kingdom (apparently, unicellular forms) and at the base of the plant kingdom, whereas myxomycetes were positioned near the base of the stock of fungi. This system is the most popular and in common use till date. The frequently adopted is Whittaker's five-kingdom system of cellular organisms modified by Lynn Margulis [5–8]. She thus made monophyletic the kingdoms of plants, fungi, and animals but increased the heterogeneity of the kingdom Protoctista. L. Margulis herself frankly admitted that "the

dom system.

4 Phylogenetics

from the lower kingdom of Protista (**Figure 2**).

**Figure 1.** Two kingdoms of K. Linnaeus system (from Drozdov [5]).

Whittaker's scheme was one of the last systems where adaptive features are interwoven with truly phylogenetic characteristics, that is, convergent similarity is claimed to be affinity. Being an ecologist, R.H. Whittaker himself pointed to the adaptive character of evolution of three higher kingdoms, which are connected with their feeding mode: plants are generally autotrophs, fungi feed by absorption, whereas animals are characterized by holozoic nutrition and digestion. R.H. Whittaker indicated this main trend in evolution with three arrows near each of the three higher kingdoms. As regards higher plants (development of the root, conductive system, orifices, reduction of gametophyte, and then loss of flagellate stage, appearance of seed and fruit), true fungi (loss of flagella), and higher vertebrates (appearance of amnion, egg enclosed by membranes, or viviparity), their progressive evolution is not related to feeding, but rather it is connected with adaptation for life on land and with the abandonment of whatever, even temporary, residence in the aquatic environment. It is no occasion that both the botanist Zernov [9] and the ecologist Odum [10, 11] considered Whittaker's scheme to be a functional, i.e. (*id est*), ecological, rather than a taxonomic one, and Y. Odum emphasized that his kingdoms, which are "functional kingdoms," should not be confused with taxonomic ones, although there are some parallels. It is therefore natural that L. Margulis, when revising Whittaker's system, replaced these arrows with another ones, indicating haplo-diploid nature of plants, diploid nature of animals, dikaryoid nature of fungi, and haploid nature of algae.

Other systems have been considered by us in details [12, 13]. In them [14–19], the eukaryotes are subdivided into 6–18 kingdoms. Thus, Edwards [16] proposed to distribute plants among seven kingdoms belonging to two subkingdoms of Prokaryota (kingdom of blue-green algae) and eukaryotic plants with six kingdoms: Erythrobionta with one division Rhodophyta, Chlorobionta embracing four divisions (Chlorophyta, Bryophyta, Tracheophyta, and Euglenophyta), Ochrobionta comprising four divisions (Phaeophyta, Chrysophyta, Cryptophyta, and Pyrrophyta), Myxobionta with four divisions (Myxogastriomycota, Dictyosteliomycota, Acrasiomycota, and Protosteliomycota), Fungi 1 with four divisions (Zygomycota, Ascomycota, Basidiomycota, and Chytridiomycota), and Fungi 2 comprising three divisions (Labyrinthulomycota, Hyphochytridiomycota, and Oomycota). One can concur with most of Edwards' kingdoms. Division of fungi into two kingdoms is wellgrounded. All divisions of Fungi 2 are now assigned to the same kingdom as Ochrobionta. Only Myxobionta are now removed from plants and distributed among two kingdoms.

After K. Linnaeus, the author of system, as well as J. Cuvie, who introduced the high-rank category of phylum, which also proved very useful, was the first case. The category of domain was set forth later.

It has become absolutely clear that the major high-rank taxonomic categories of Linnaeus are insufficient and new ones are needed. The simplest way is using additional categories such as subphylum, subkingdom, and superkingdom. Many scientists went this way, but, in doing so, they were compelled to introduce more categories such as "Uberreich," "Oberreich," "Unterreic" [19] or "Superkingdom," "Subphylum," "Infraphylum," and "Parvikingdom" [20–24].

Vorontsov [26–28] was the first who introduced into scientific usage a major taxonomic category higher in rank than kingdom, the empire. He recognized two empires: of precellular organisms in which he placed a single kingdom, that of viruses, and the empire of cellular organisms embracing two subempires: subempire of prenucleate organisms comprising bacteria and blue-green algae and the subempire of nucleate organisms (or eukaryotes). The introduction of such high-level taxonomic category as the empire is of much importance for taxonomy and quite a bold suggestion.

## **3. Principle of conservatism of subcellular structures**

and higher vertebrates (appearance of amnion, egg enclosed by membranes, or viviparity), their progressive evolution is not related to feeding, but rather it is connected with adaptation for life on land and with the abandonment of whatever, even temporary, residence in the aquatic environment. It is no occasion that both the botanist Zernov [9] and the ecologist Odum [10, 11] considered Whittaker's scheme to be a functional, i.e. (*id est*), ecological, rather than a taxonomic one, and Y. Odum emphasized that his kingdoms, which are "functional kingdoms," should not be confused with taxonomic ones, although there are some parallels. It is therefore natural that L. Margulis, when revising Whittaker's system, replaced these arrows with another ones, indicating haplo-diploid nature of plants, diploid nature of animals, dikaryoid nature of fungi, and haploid nature

Other systems have been considered by us in details [12, 13]. In them [14–19], the eukaryotes are subdivided into 6–18 kingdoms. Thus, Edwards [16] proposed to distribute plants among seven kingdoms belonging to two subkingdoms of Prokaryota (kingdom of blue-green algae) and eukaryotic plants with six kingdoms: Erythrobionta with one division Rhodophyta, Chlorobionta embracing four divisions (Chlorophyta, Bryophyta, Tracheophyta, and Euglenophyta), Ochrobionta comprising four divisions (Phaeophyta, Chrysophyta, Cryptophyta, and Pyrrophyta), Myxobionta with four divisions (Myxogastriomycota, Dictyosteliomycota, Acrasiomycota, and Protosteliomycota), Fungi 1 with four divisions (Zygomycota, Ascomycota, Basidiomycota, and Chytridiomycota), and Fungi 2 comprising three divisions (Labyrinthulomycota, Hyphochytridiomycota, and Oomycota). One can concur with most of Edwards' kingdoms. Division of fungi into two kingdoms is wellgrounded. All divisions of Fungi 2 are now assigned to the same kingdom as Ochrobionta. Only Myxobionta are now removed from plants and distributed among two kingdoms.

After K. Linnaeus, the author of system, as well as J. Cuvie, who introduced the high-rank category of phylum, which also proved very useful, was the first case. The category of domain

It has become absolutely clear that the major high-rank taxonomic categories of Linnaeus are insufficient and new ones are needed. The simplest way is using additional categories such as subphylum, subkingdom, and superkingdom. Many scientists went this way, but, in doing so, they were compelled to introduce more categories such as "Uberreich," "Oberreich," "Unterreic" [19] or "Superkingdom," "Subphylum," "Infraphylum," and "Parvikingdom"

Vorontsov [26–28] was the first who introduced into scientific usage a major taxonomic category higher in rank than kingdom, the empire. He recognized two empires: of precellular organisms in which he placed a single kingdom, that of viruses, and the empire of cellular organisms embracing two subempires: subempire of prenucleate organisms comprising bacteria and blue-green algae and the subempire of nucleate organisms (or eukaryotes). The introduction of such high-level taxonomic category as the empire is of much importance for

of algae.

6 Phylogenetics

was set forth later.

taxonomy and quite a bold suggestion.

[20–24].

The multikingdom system of the organic world was advanced by us [11, 12, 28–30]. It was based on the principle of conservatism of cellular structures formulated by Mashansky and Drozdov [31, 32]. There is a biological paradox: the subcellular structures are highly conservative.

While discussing the structural foundations of biological systems, we should not limit our attention by fixing it only to the correlation of various levels of the organization of living matter, to what we are used to call the problem of integration. We have to realize that every single level is unique in its qualitative specificity and particular features.

There is an enormous variety of cells. They differ in their morphology, functions, and their chemical structure. However, when we proceed to the next, the so-called subcellular level, we are confronted with the fact that the principal structure of basic cellular organelles, such as membranes, mitochondria, centrioles, filaments, ribosomes, endoplasmatic reticulum, and Golgi apparatus, remains unchanged in a wide variety of objects. In the hierarchical sequence of the organizational levels of biological systems, it is the subcellular level, and that merits ever greater attention for its most remarkable characteristic feature, namely its membranous structure—its supermolecular system of proteins, lipids, and polysaccharides of several types. We can be certain to expect some new properties to be discovered typical of the structure and functions of cellular organelles on account of the peculiarities of their level of organization.

There are only two variants of the ultrastructure of biological membranes (lipid bilayer in Eubacteria and Eukaryotes and single layer in Archaebacteria), two variants of ribosomes, six variants of ultrastructural organization of plastids, three variants of organization of mitochondria, three variants of organization of nuclear apparatus, and three variants of organization of kinetic apparatus.

A suitable object for a comparative morphological analysis is presented by mitochondria. For identifying of mitochondria can be taken the typical organization of their membranes. The lipoprotein nature of mitochondrial membranes does not cause any doubt, neither does the similarity of these membranes to the ones belonging to other organelles. Yet, there are data on the mitochondrial membrane testifying to its structural as well as functional uniqueness.

Mitochondria are remarkable for the great variety of their organizations. There are lamellar or tubular crysts that can exist singly or densely packed, or they can be either scattered or highly organized. There can be several small mitochondria in a cell, or a single one spreading over a large number of shoots: there can be one mitochondrion in a cell, or quite a number of them densely packed together. Despite such great variety, the general pattern of the structure of mitochondria invariably repeats itself—it is one and the same in mushrooms, algae, multicellular animals, and plants. There are four types of structures of the crysts of mitochondria—the lamellar, the tubular, the tubularly vesicular, and discoid one. The nature of mechanisms determining the morphology of mitochondrial crysts is unknown yet. Nevertheless, the functional peculiarities of the cells are of considerable significance. Thus, in the cells synthesizing steroid hormones, we find mitochondria with tubularly vesicular crysts. However, an injection of steroid hormones to lower invertebrate allows to transform the mitochondria of the neurons with typically lamellar crysts into those with tubularly vesicular ones [31].

Mitochondria can cardinally change their ultrastructure under the impact of alternating factors or training. This signifies high liability in mitochondria, the ultrastructure of which is determined by the function of the cells irrespective of the systematic position of the object. On the basis of presently available material on the ultrastructure of mitochondria of the cells of a great variety of tissues as well as the specificity of their responses to various alternating factors, it is possible to state that there are no convincing facts which might permit to fix any correlations between the level of phylogenetic position, or ontogeny and the ultrastructure of mitochondria in investigated species belonging to different realms of living organisms. All the observed differences in the ultrastructure of mitochondria can be accounted for by their functional peculiarities, their loads during a certain period of activity. There, evidently, lies one of the qualitative peculiarities of the subcellular level of the organization of living systems, which has been named "the principle of conservatism" [31, 32].

These facts demonstrating the lack of changes in cellular organelles, such as rather intricate in their organization mitochondria, during the long process of their evolution give a reason to conclude that already at the early stages of evolution, the structural as well as chemical organization of living systems was rather complicated and well developed. This fact calls for a discussion of the problems of the early stages of evolution, beginning with the appearance of life on Earth, which is currently widely discussed on various levels, and extreme views are being stated.

The uniformity of the structure of cellular organelles, such as mitochondria and, perhaps, even those of a more intricate organization, namely filaments, gives grounds to view them as structures formed on one single occasion. The structure of nucleic acids has a common origin in all living systems. This conclusion is prompted by the widely known uniformity of the code formed on four bases.

The above-mentioned conception should explain why mitochondria have a genetic code that differs very slightly from that of the nucleus as well as that of the prokaryotes. In fact, the code of mitochondria differs very little from the universal one. Only five codons have different meaning: methionine, isoleucine, tryptophan, and, also, a changed terminator. While analyzing these divergences, it is possible to see that the code of mitochondria is nearer to the quasidouble "ideal" one. This may testify the fact that the code of mitochondria is more ancient than the universal one. Possibly, there was a time when all cells had a code similar to that of present-day mitochondria. Then, some changes occurred in the general code, but in mitochondria, the code proved to be more stable. The reason for this may lie in the small size of the genome of mitochondria and so every mutation brought about such changes in the characteristics that proved lethal.

There is another important problem, namely, why the mitochondrial genetic system, once formed, survived in the evolution practically unchanged, and how could it preserve its independence in the cell. Mahler and coauthors [33] suggested the opinion that, as polypeptides coded by DNA and belonging to mitochondrial complexes are rather hydrophobic, they must be synthesized somewhere near the place of their inclusion into the mitochondrial membrane and cannot be transported through the cell. Probably, the preservation of mt-DNA throughout the evolution was due to it serving as a supplier of the functionally indispensable elements to mitochondria.

We believe that the most topical, fruitful, and perspective objective of megasystematics is the elaboration of multikingdom system consisting of monophyletic taxa, and we are aware of all difficulties of this task. One of the main difficulties is that now in a boiling cauldron of new information, one can hardly make a whatever stable system. There are many reasons for this. The ultrastructure of many protists has not yet been studied; the structure of many organisms is either very peculiar or unclear so that they cannot be classified with the existing taxa of even high rank; the degree of conservatism of cellular structures is being elucidated; and the techniques for demonstrating relatedness are being improved. It is, therefore, no accident that different authors recognize different number of kingdoms, and the authors themselves sometimes remake their systems too hastily. Thus, Cavalier-Smith [34] delineates seven kingdoms among the eukaryotes; 3 years later, he already recognized nine kingdoms, and later he reduced the number of kingdoms to six [21–24, 35–39].

## **4. Multikingdom systems of the organic world**

tional peculiarities of the cells are of considerable significance. Thus, in the cells synthesizing steroid hormones, we find mitochondria with tubularly vesicular crysts. However, an injection of steroid hormones to lower invertebrate allows to transform the mitochondria of the

Mitochondria can cardinally change their ultrastructure under the impact of alternating factors or training. This signifies high liability in mitochondria, the ultrastructure of which is determined by the function of the cells irrespective of the systematic position of the object. On the basis of presently available material on the ultrastructure of mitochondria of the cells of a great variety of tissues as well as the specificity of their responses to various alternating factors, it is possible to state that there are no convincing facts which might permit to fix any correlations between the level of phylogenetic position, or ontogeny and the ultrastructure of mitochondria in investigated species belonging to different realms of living organisms. All the observed differences in the ultrastructure of mitochondria can be accounted for by their functional peculiarities, their loads during a certain period of activity. There, evidently, lies one of the qualitative peculiarities of the subcellular level of the organization of living systems,

These facts demonstrating the lack of changes in cellular organelles, such as rather intricate in their organization mitochondria, during the long process of their evolution give a reason to conclude that already at the early stages of evolution, the structural as well as chemical organization of living systems was rather complicated and well developed. This fact calls for a discussion of the problems of the early stages of evolution, beginning with the appearance of life on Earth, which is currently widely discussed on various levels, and extreme views are being stated.

The uniformity of the structure of cellular organelles, such as mitochondria and, perhaps, even those of a more intricate organization, namely filaments, gives grounds to view them as structures formed on one single occasion. The structure of nucleic acids has a common origin in all living systems. This conclusion is prompted by the widely known uniformity of the code

The above-mentioned conception should explain why mitochondria have a genetic code that differs very slightly from that of the nucleus as well as that of the prokaryotes. In fact, the code of mitochondria differs very little from the universal one. Only five codons have different meaning: methionine, isoleucine, tryptophan, and, also, a changed terminator. While analyzing these divergences, it is possible to see that the code of mitochondria is nearer to the quasidouble "ideal" one. This may testify the fact that the code of mitochondria is more ancient than the universal one. Possibly, there was a time when all cells had a code similar to that of present-day mitochondria. Then, some changes occurred in the general code, but in mitochondria, the code proved to be more stable. The reason for this may lie in the small size of the genome of mitochondria and so every mutation brought about such changes in the

There is another important problem, namely, why the mitochondrial genetic system, once formed, survived in the evolution practically unchanged, and how could it preserve its independence in the cell. Mahler and coauthors [33] suggested the opinion that, as polypeptides

neurons with typically lamellar crysts into those with tubularly vesicular ones [31].

which has been named "the principle of conservatism" [31, 32].

formed on four bases.

8 Phylogenetics

characteristics that proved lethal.

Nevertheless, the adherents of monophyletic system have made tremendous progress. The kingdom Ochrobiontes (Chromobionta or Chromista) is distinctly delineated; along with a number of divisions of chlorophyll *C*-containing algae, it embraces some groups from the kingdoms Fungi and Protozoa. The kingdom Viridiplantae comprises all green algae Chlorophyta s. lato, bryophytes, and higher plants but no more; the kingdom Metazoa (but not Animalia!) is also monophyletic. Some kingdoms are not as clearly delineated as Euglenobiontes, Alveolates, Cryptobiontes, Prymnesiobiontes, etc. are. However, some groups, among them Foraminifera, Radiolaria s. lato, and others, have not yet been placed properly. Instead of being squeezed, without due grounds, into the existing kingdoms, these groups should rather be regarded as groups incertae sedis, as is done by many taxonomists.

When analyzing the old system, in which features of adaptive similarity and phylogenetic relatedness turned out to be intermingled, there is an increasing criticism from various investigators that many taxa are not monophyletic but rather ecomorphological notions. Shafranova [40] addressed this problem in her paper "Plant as a Life Form." Mirabdullaev [41, 42] correctly points out that the former system of protists was primarily the system of life forms (ecomorphs) rather than phylogenetic taxa and that similar structural patterns can arise convergently. Many foreign scientists are now coming to analogous conclusions. Here, the question arises: what should be done with out-dated, traditional notions that serve no longer as taxa, namely plants, protists, animals, heliozoans, flagellates, sporozoans, etc. To avoid extremely troublesome and even unnecessary rejection of old terminology, many researchers began using them not as taxa but as designations of ecomorphs or life forms [43, 44]. This does not necessarily imply that the existing terminology should be changed radically. Up to now, in botanical institutions, mycologists have successfully worked hand in hand with algologists, and both prokaryotic bacteria and eukaryotic fungi have been applied in microbiological industry.

Moreover, it has turned out that these terms can and must be used in the ecomorphological system or the system of life forms, which has long been a necessity. Teofrast's system was one of early attempts at constructing such a system. To date, a variety of such systems have been created at different levels. Unfortunately, the ecomorphological system was elaborated independently of the taxonomic one, which was thought of as if being something stable, and its terms were little used. The adoption and use of the terms that are well established in taxonomy were not appropriate for the new system. Thus, in his ecomorphological system, which is one of the better developed, for high-rank taxa, he retains the names "Kingdom," "Division," "Phylum," and "Class," which can cause only confusion. Barr's viewpoint seems to be more correct [44]. Only for fungi, he proposed two systems: a phylogenetic one, where fungi were distributed among three kingdoms—Eumycota, Chromista, and Protozoa—and an ecological one, in which fungi in the old sense constitute union 1 of Fungi.

The idea is to create, on the basis of the old system in which the genetic and ecomorphological criteria were intermingled, two parallel systems—the phylema or phylogenetic, taxonomic system and the ecomorphological system. The elaboration of the ecomorphological system is a very complicated task, although much has been done in this respect. Without doubt, many descriptive terms of traditional systematics will find their place in the new system.

At present a lot of biologists study the problems of megasystematics. Close with our megasystem was build up the system by Leontiev and Akulov [45]. But most of new systems limit themselves to study the sequence of nucleotide in ribosomal RNA. The molecular biologists studying rRNA work at different countries—in USA, Canada, Belgium, Japan, and different European countries like Russia. There are a few sites in Internet, where the phylogenetic trees are represented. A lot of such trees were published last years. Attention should be paid to the discussion of their systems as variant of five-kingdom system. Nevertheless, Cavalier-Smith [20–24, 34–39] already published the six to nine kingdoms systems. He comprises two empires—Prokaryota and Eukaryota.

Since the end of year 1970, the concept of three branches of cellular organisms is accepted in megasystematics [46]. It is proposed to give these branches of organic word the rank of domains Archaea, Bacteria, and Eucarya [47–51]. Therefore, the empire Cellulata is divided to three domains, which, in turn, are divided into several kingdoms [25, 39] (**Figures 3**–**7**).

The scheme reflects the great diversity of life forms of bacteria adapted to living in almost all ecological niches. Some of them such as *Ancalochloris* (1), *Aquaspirillum* (2), and *Chromatin* (3) live in water, whereas *Aquaspirillum* can use a chain of magnetized particles to find sediments, rich in nutrient agents. *Haloarcula* (4) are distributed in the saline marshes. *Pyrodictium* (5) prefers hot places; *Rhizobium* (6) settles in the roots of plants and produces nitrogen available to the host tissue form. Type of bacteria: *Escherichia* (7), *Streptococcus* (8), <sup>10</sup> Phylogenetics Principle of Conservatism of Cellular Structures as the Basis for Construction of the Multikingdom System of the... <sup>9</sup> Principle of Conservatism of Cellular Structures as the Basis for Construction of the Multikingdom... http://dx.doi.org/10.5772/intechopen.68562 11

**Figure 3.** The main bacteria morphotypes (from Kussakin, Drozdov [11]). Archaebacteria: 1, *Methanococcus*; 2, *Methanobacterium, Halobacterium*; 3, *Thermoplasma*; 4, *Methanospirillum*; 5, *Haloarcula*; 6, square bacteria; 7, *Sulfolobus*; 8, *Pyrodictium*. Gram-negative bacteria (Gracilicutes): 9, *Neisseria, Veillonella*; 10, *Gemmiger*; 11, *Escherichia*; 12, *Seliberia*; 13, *Vibrio, Bdellovibrio*; 14, *Mycrocyclus*; 15, *Spirillum*; 16, *Spirochaeta*; 17, *Angulomicrobium*; 18, *Stella*; 19, *Prosthecomicrobium*; 20, *Caulobacter*; 21, *Hyphomicrobium, Rhodomicrobium*; 22, *Mastigocoleus*; 23, *Simonsiella*; 24, *Oscillochloris, Oscillatoria*. Gram-positive bacteria (Firmicutes): 25, *Micrococcus*; 26, *Bacillus, Erysipelothrix*; 27 and 28, *Desulfotomaculum, Clostridium*; 29, *Mycobacterium*; 30, *Streptomyces*; 31, *Caryophanon, Oscillospira*. Mycoplasma (Tenericutes): 32, 33, and 35, *Mycoplasma*; 34, *Spiroplasma*.

**Figure 4.** The different forms of Eubacteria (from Drozdov [5]).

began using them not as taxa but as designations of ecomorphs or life forms [43, 44]. This does not necessarily imply that the existing terminology should be changed radically. Up to now, in botanical institutions, mycologists have successfully worked hand in hand with algologists, and both prokaryotic bacteria and eukaryotic fungi have been applied in micro-

Moreover, it has turned out that these terms can and must be used in the ecomorphological system or the system of life forms, which has long been a necessity. Teofrast's system was one of early attempts at constructing such a system. To date, a variety of such systems have been created at different levels. Unfortunately, the ecomorphological system was elaborated independently of the taxonomic one, which was thought of as if being something stable, and its terms were little used. The adoption and use of the terms that are well established in taxonomy were not appropriate for the new system. Thus, in his ecomorphological system, which is one of the better developed, for high-rank taxa, he retains the names "Kingdom," "Division," "Phylum," and "Class," which can cause only confusion. Barr's viewpoint seems to be more correct [44]. Only for fungi, he proposed two systems: a phylogenetic one, where fungi were distributed among three kingdoms—Eumycota, Chromista, and Protozoa—and an ecological

The idea is to create, on the basis of the old system in which the genetic and ecomorphological criteria were intermingled, two parallel systems—the phylema or phylogenetic, taxonomic system and the ecomorphological system. The elaboration of the ecomorphological system is a very complicated task, although much has been done in this respect. Without doubt, many

At present a lot of biologists study the problems of megasystematics. Close with our megasystem was build up the system by Leontiev and Akulov [45]. But most of new systems limit themselves to study the sequence of nucleotide in ribosomal RNA. The molecular biologists studying rRNA work at different countries—in USA, Canada, Belgium, Japan, and different European countries like Russia. There are a few sites in Internet, where the phylogenetic trees are represented. A lot of such trees were published last years. Attention should be paid to the discussion of their systems as variant of five-kingdom system. Nevertheless, Cavalier-Smith [20–24, 34–39] already published the six to nine kingdoms systems. He comprises two

Since the end of year 1970, the concept of three branches of cellular organisms is accepted in megasystematics [46]. It is proposed to give these branches of organic word the rank of domains Archaea, Bacteria, and Eucarya [47–51]. Therefore, the empire Cellulata is divided to three domains, which, in turn, are divided into several kingdoms [25, 39] (**Figures 3**–**7**).

The scheme reflects the great diversity of life forms of bacteria adapted to living in almost all ecological niches. Some of them such as *Ancalochloris* (1), *Aquaspirillum* (2), and *Chromatin* (3) live in water, whereas *Aquaspirillum* can use a chain of magnetized particles to find sediments, rich in nutrient agents. *Haloarcula* (4) are distributed in the saline marshes. *Pyrodictium* (5) prefers hot places; *Rhizobium* (6) settles in the roots of plants and produces nitrogen available to the host tissue form. Type of bacteria: *Escherichia* (7), *Streptococcus* (8),

descriptive terms of traditional systematics will find their place in the new system.

one, in which fungi in the old sense constitute union 1 of Fungi.

empires—Prokaryota and Eukaryota.

biological industry.

**Figure 5.** Schematic representation of the major lines of prokaryotic descent (after Fox et al. [46]).

and *Treponema* (9) cause various diseases in humans. The metabolism requirements can combine incompatible species of bacteria: aerobic methane consumer *Methylococcus* (10)

**Figure 6.** Unrooted tree shows the three branch of organic word (after Woese [49]).

Principle of Conservatism of Cellular Structures as the Basis for Construction of the Multikingdom... http://dx.doi.org/10.5772/intechopen.68562 13

**Figure 7.** Kandler ring. The phylogenetic unrooted tree constructed on base of analysis rRNA and cell wall (for prokaryotes) (after Kandler [52]).

and *Treponema* (9) cause various diseases in humans. The metabolism requirements can combine incompatible species of bacteria: aerobic methane consumer *Methylococcus* (10)

**Figure 5.** Schematic representation of the major lines of prokaryotic descent (after Fox et al. [46]).

12 Phylogenetics

**Figure 6.** Unrooted tree shows the three branch of organic word (after Woese [49]).

draws *Methanosarcina* (11), and anaerobic producing methane *Desulfovibrio* (12), producing hydrogen sulfide—*Ancalochloris* (1), *Beggitoa* (13), and *Chromatium* (3)— requires hydrogen sulfide. Another group of bacteria, consuming hydrogen sulfide, *Thiobacillus* (14), is used for extraction of metals from ore. *Streptomyces* (15) secrete antibiotics. *Anabaena* (16) produces oxygen from water in the process of photosynthesis, whereas *Bdellovibrio* attacks many other bacteria (17).

We support this idea and propose to distinguish 4 kingdoms in Archaebacteria, 7 kingdoms in Eubacteria, and 15 kingdoms in Eukaryotes. Our system we represent as scheme (**Figure 8**) and as the table (**Table 2**). In **Table 1**, we propose the next ends for word of designations of taxa on levels kingdom, phylum, class and order.

**Figure8.** The multikingdom phylogenetic unrooted tree constructed on base of principle of conservatism (after Drozdov [5]). I, Virae; II, Prokaryotes: 1, Methanobacteriobiontes; 2, Halobacteriobiontes; 3, Thermoacidobacteriobiontes; 4, Archaetenericutobacteriobiontes; 5, Tenericotobacteriobiontes; 6, Actinobacteriobiontes; 7, Firmicutobacteriobiontes; 8, Spirochaetobacteriobiontes; 9, Scotobacteriobiontes; 10, Anoxyphotobacteriobiontes; 11, Oxyphotobacteriobiontes; III, Eukaryotes: 12, Rhodobiontes; 13, Cryptobiontes; 14, Chlorobiontes (a, Thallobionti; б, Embryobionti); 15, Parazoobiontes; 16, Metazoobiontes; 17, Mycobiontes; 18, Alveolatobiontes (a, Peridiniobionti; b, Parameciobionti); 19, Foraminiferobiontes; 20, Radiolariobiontes; 21, Myxobiontes; 22, Prymnesiobiontes; 23, Heterokontobiontes; 24, Euglenobiontes; 25, Archaemonadobiontes; 26, Microsporobiontes.


**Table 1.** Applicable ends for word of designations of taxa on kingdom, phylum, classis, and order taxonomic rank.

#### **Imperia Cellulata**

#### **Dominion Archaebacteria**

#### **I. Kingdom Thermoacidobacteriobiontes**


#### **II. Kingdom Archaetenericutobacteriobiontes**

3. Phylum Thermoplasmophyles

#### **III. Kingdom Halobacteriobiontes**


#### **IV. Kingdom Methanobacteriobiontes**

6. Phylum Methanobacteriophyles

#### **Dominion Eubacteria**

#### **Superkingdom Gracilicutobiontoi**

#### **V. Kingdom Cyanobiontes (Oxyphotobacteriobiontes)**


#### **VI. Kingdom Anoxyphotobacteriobiontes**


#### **VII. Kingdom Scotobacteriobiontes**


**Category Applicable ends Category Applicable ends**

**Figure8.** The multikingdom phylogenetic unrooted tree constructed on base of principle of conservatism (after Drozdov [5]). I, Virae; II, Prokaryotes: 1, Methanobacteriobiontes; 2, Halobacteriobiontes; 3, Thermoacidobacteriobiontes; 4, Archaetenericutobacteriobiontes; 5, Tenericotobacteriobiontes; 6, Actinobacteriobiontes; 7, Firmicutobacteriobiontes; 8, Spirochaetobacteriobiontes; 9, Scotobacteriobiontes; 10, Anoxyphotobacteriobiontes; 11, Oxyphotobacteriobiontes; III, Eukaryotes: 12, Rhodobiontes; 13, Cryptobiontes; 14, Chlorobiontes (a, Thallobionti; б, Embryobionti); 15, Parazoobiontes; 16, Metazoobiontes; 17, Mycobiontes; 18, Alveolatobiontes (a, Peridiniobionti; b, Parameciobionti); 19, Foraminiferobiontes; 20, Radiolariobiontes; 21, Myxobiontes; 22, Prymnesiobiontes; 23, Heterokontobiontes; 24,

**Table 1.** Applicable ends for word of designations of taxa on kingdom, phylum, classis, and order taxonomic rank.

Superkingdom -obiontoi Superclassis -idees Kingdom -obiontes Classis --indes Subkingdom -obiontoi Subclassis -iones Superphylum -ophylaces Superorder -iformi Phylum -ophylea Order -iformes Subphylum -ophylinea Suborder -oidei

Euglenobiontes; 25, Archaemonadobiontes; 26, Microsporobiontes.

14 Phylogenetics



Class Acrasilodes

Class Percolomonadiodes

Class Lyromonadioides

**VIII. Kingdom Spirochaetobacteriobiontes**

22. Phylum Spirochaetophyles **Superkingdom Firmicutobiontoi IX. Kingdom Actinobacteriobiontes** 23. Phylum Mycobacteriophyles 24. Phylum Corynebacteriophyles 25. Phylum Actinomycetophyles **X. Kingdom Eufirmicutobiontes** 26. Phylum Clostridiophyles 27. Phylum Bacillophyles 28. Phylum Lactobacillophyles 29. Phylum Micrococcophyles **XI. Kingdom Tenericutobiontes** 30. Phylum Mycoplasmophyles

16 Phylogenetics

**Dominion Eukaryota**

**XII. Kingdom Microsporobiontes** 31. Phylum Microsporidiophyles **XIII. Kingdom Archemonadobiontes Superphylum Archamoebophylacei**

32. Phylum Pelomyxophyles

**Superphylum Metamonadophylacei** 33. Phylum Retortomonadophyles 34. Phylum Hexamitophyles 35. Phylum Oxymonadophyles **Superphylum Parabasaliophylacei** 36. Phylum Trichomonadophyles

Class Pelornyxiodes Class Mastigamoeboides

Class Trichonymphiodes

**XIV. Kingdom Euglenobiontes Subkingdom Percolobionti** 37. Phylum Acrasiophyles Class Vahlkampfiiodes Class Acrasilodes

#### **Subkingdom Euglenobionti**

38. Phylum Stephanopogonophyles

39. Phylum Diplonemophyles

40. Phylum Bodonophyles

41. Phylum Euglenophyles

**XV. Kingdom Myxobiontes**

#### **Subkingdom Myxomycetobionti**

42. Phylum Cercomonadophyles

43. Phylum Dictyosteliophyles

44. Phylum Physarophyles

#### **Subkingdom Myxozoobionti**

45. Phylum Entamoebophyles

46. Phylum Haplosporophyles

47. Phylum Pararnyxiophyles

48. Phylum Myxidiophyles

#### **XVI. Kingdom Rhodobiontes**

49. Phylum Bangiophyles

**XVII. Kingdom Alveolatobiontes**

**Subkingdom Peridiniobionti**

**Superphylum Peridiniophylacei**

50. Phylum Peridiniophyles

#### **Superphylum Apicomplexophylacei**

51. Phylum Perkinsophyles

Class Colpodelliodes

Class Perkinsiodes

52. Phylum Gregarinophyles

#### **Subkingdom Parameciobionti**

53. Phylum Hemimastigophyles

54. Phylum Parameciophyles

#### **XVIII. Kingdom Heterokontobiontes**

55. Phylum Bicosoecophyles

56. Phylum Labyrinthulophyles


Class Pelagomonadiodes

#### **XIX. Kingdom Foraminiferobiontes**


#### **XX. Kingdom Radiolariobiontes**


#### **XXI. Kingdom Prymnesiobiontes (=Haptophyta)**

75. Phylum Prymnesiophyles (=Haptophyles)

#### **XXII. Kingdom Cryptobiontes**


#### **XXIII. Kingdom Chlorobiontes (=Viridiplantae)**

#### **Subkingdom Thallobionti**

78. Phylum (Division) Prasinophyles

79. Phylum (Division) Chlorophyles

80. Phylum (Division) Charophyles

57. Phylum Saprolegniophyles 58. Phylum Hyphochytriophyles 59. Phylum Diatomophyles 60. Phylum Triboneroatophyles

18 Phylogenetics

61. Phylum Fucophyles

62. Phylum Eustigmatophyles 63. Phylum Synurophyles 64. Phylum Chrysococcophyles 65. Phylum Raphidomonadophyles

66. Phylum Dictyochophyles 67. Phylum Pedinellophyles

Addition to Kingdom Heterokontobiontes

68. Phylum Psamminidophyles (=Xenophyophora)

71. Phylum Sphaerozoiophyles (=Polycystinea)

**XXI. Kingdom Prymnesiobiontes (=Haptophyta)** 75. Phylum Prymnesiophyles (=Haptophyles)

76. Tип Cryptomonadophyles (Cryptophycota) 77. Tип Centrochelidophyles (Acantocystidae) **XXIII. Kingdom Chlorobiontes (=Viridiplantae)**

**XIX. Kingdom Foraminiferobiontes**

69. Phylum Foraminiferophyles 70. Phylum Plasmodiophoreophyles **XX. Kingdom Radiolariobiontes**

72. Phylum Phaeodiniophyles 73. Phylum Acanthometriophyles 74. Phylum Sticholoncheiophyles

**XXII. Kingdom Cryptobiontes**

**Subkingdom Thallobionti**

78. Phylum (Division) Prasinophyles 79. Phylum (Division) Chlorophyles

Class Pedinelliodes Class Actinophryiodes Class Clathruliniodes

Class Pelagomonadiodes

#### **Subkingdom Embryobionti (= Cormobionti)**

81. Phylum (Division) Bryophyles

82. Phylum (Division) Rhyniophyles (= Psilophyles)

83. Phylum (Division) Psilotophyles

84. Phylum (Division) Lycopodiophyles

85. Phylum (Division) Equisetophyles (Sphenophyles)

86. Phylum (Division) Polypodiophyles (= Filicophyles)

87. Phylum (Division) Pinophyles (= Gymnospermae)

88. Phylum (Division) Magnoliophyles (= Angiospermae)

**XXIV. Kingdom Mycobiontes (= Fungi)**

#### **Subkingdom Opistomastigomycotobionti**

89. Phylum (Division) Chytridiomycotophyles

#### **Subkingdom Amastigomycotobionti (= Eufungi=Eumycota)**

90. Phylum (Division) Mucoromycotaphyles (= Zygomycota)

91. Phylum (Division) Trichomycotaphyles

92. Phylum (Division) Ascomycotaphyles

93. Phylum (Division) Basidiomycotaphyles

#### **XXV. Kingdom Parazoobiontes**

94. Phylum Choanoflagellata (= Crasperomonadia)

95. Phylum Spongia (= Porifera)

#### **XXVI. Kingdom Metazoobiontes**

96. Phylum Placozoa

97. Phylum Cnidaria

98. Phylum Ctenophora



**Table 2.** Multikingdom system of the cellular living beings.

## **5. The root phylogenetic tree**

The construction of a root phylogenetic tree based on the principle of conservatism is not simple. It is necessary to analyze the structure of the six systems of cellular organelles in each Protista group: surface apparatus (membranome), genetic apparatus (karyome), synthetic apparatus (syndetome), mitochondria, plastids, and kinetic apparatus (kinetome). In many ways, it can be done based on intuition. Of course, now the study of the building of phylema of the organic world focuses mainly on the genomic level. Nevertheless, we tried to present phylema of the organic world in a tree, where the kingdom is placed as the complexity of the systems of cellular organelles (**Figure 8**). The main complication is the allocation of the core group in the structure of the tree. It may seem that the problem is simple—the most primitive group includes cells of the simplest arrangement structure. Certainly, the simplest organisms are Microsporobiontes—eukaryotic unicellular intracellular parasites. They have only plasmatic membrane, nucleus, and ribosome. Moreover, their ribosome is closer to 70S-prokaryotic ribosome than to 80S-eukaryotic ribosome. The first molecular studies of ribosomal RNA sequence suggest that Microsporidia are extremely ancient eukaryotes [35, 53]. Later, biochemists discovered that phylogenomics supports Microsporidia as the earliest diverging clade of sequenced fungi [54–59]. Therefore, Microsporidia are secondarily simplified, during adaptation to intracellular anaerobic existence.

The second candidate for the most primitive Eukaryota is the Kingdom Archemonadobiontes with Pelomyxophyles, Retortomonadophyles, Hexamitophyles, Oxymonadophyles, and Trichomonadophyles. They are anaerobic organisms without mitochondria but have from two to numerous flagella. The problem is: had they originally no primary mitochondria or they lost them during adaptation to anaerobic environment? Most professionals concerned with megasystematics are inclined to consider anaerobic eukaryotes as the result of their secondary simplification: they have lost their mitochondria, adapting to obligate anaerobic metabolism.

Rhodobiontes (red algae) had no flagella originally or they have lost them? This is a problem, because they are marine algae only and flagella are necessary organelles in water environment.

## **6. Conclusion**

**5. The root phylogenetic tree**

Incertae sedis: Genera *Gyromitus*; Genera *Discocelis*; Genera *Jacoba*.

**Table 2.** Multikingdom system of the cellular living beings.

108. Phylum Vestimentifera 109. Phylum Tardigrada 110. Phylum Pentastomida 111. Phylum Onichophora 112. Phylum Arthropoda 113. Phylum Rotifera 114. Phylum Cycliophora 115. Phylum Acanthocephala

20 Phylogenetics

116. Phylum Dicyemataria (= Rhombozoa)

117. Phylum Nemathelminthes

118. Phylum Loricifera 119. Phylum Gastrotricha 120. Phylum Nematomorpha 121. Phylum Priapulida 122. Phylum Kinorhyncha 123. Phylum Chaetognatha 124. Phylum Phoronida 125. Phylum Bryozoa 126. Phylum Brachiopoda 127. Phylum Hemichordata 128. Phylum Echinodermata 129. Phylum Chordata

The construction of a root phylogenetic tree based on the principle of conservatism is not simple. It is necessary to analyze the structure of the six systems of cellular organelles in each Protista group: surface apparatus (membranome), genetic apparatus (karyome), synthetic apparatus (syndetome), mitochondria, plastids, and kinetic apparatus (kinetome). In many ways, it can be done based on intuition. Of course, now the study of the building of phylema of the organic world focuses mainly on the genomic level. Nevertheless, we tried to present phylema of the organic world in a tree, where the kingdom is placed as the complexity of the systems of cellular organelles (**Figure 8**). The main complication is the allocation Euglenoids that have all organelles (membrane with special cell wall, nucleus, 80S ribosome, mitochondria with discoid crista, plastids, and flagella), may be considered as most primitive Eukaryota. According to our system [12], Kingdom Euglenobiontes Leedale, 1974 (from the Greek eu -, in English "good," in compound words it means "well-developed," "authentic," consistent with the ideal and glene—the pupil of the eye) combines the heterotrophic or autotrophic green, usually unicellular monad often amoeboid, but usually with a monadic form in the cycle, rarely colonial organisms. They have and mitochondria with cristae that are usually flattened, rounded with a tapered base—discoid, rarely vesicular, or even less often ribbonlike tube; usually single-nucleus; mitosis in a closed intranuclear ortomitosis; reproduction by a longitudinal division; sexual process is unknown. This kingdom includes two subkingdoms: Euglenobionti and Percolobionti. Although acrasia and heterolobosea amoebas are combined into one common taxon usually called Heterolobosea, we prefer to give it the name from the type genus Acrasia-Acrasiophyles.

With this assumption, understanding of phylogenetics of Eukaryota has no problem. Eukaryota are divided into two branches: Tubulicristata (with mitochondria with tubular crista) and Lamellicristata (with mitochondria with lamellar crista). Cryptomonads occupy an intermediate position with riblike crista and nukleomorf in plastids (**Figure 9** and **Table 2**).

**Figure 9.** The multikingdom phylogenetic hierarchical tree constructed on base of principle of conservatism (after Drozdov [5]).

#### **Author details**

Anatoliy L. Drozdov

\*Address all correspondence to: anatoliyld@mail.ru

1 National Scientific Center of Marine Biology, Far Eastern Branch, Russian Academy of Sciences, A.V. Zhirmunsky Institute of Marine Biology FEB RAS, Vladivostok, Russia

2 Far Eastern Federal University, Vladivostok, Russia

#### **References**

[1] Whittaker RH. On the broad classification of organisms. The Quarterly Review of Biology. 1959;**34**:210-226


**Author details**

Drozdov [5]).

22 Phylogenetics

Anatoliy L. Drozdov

**References**

Biology. 1959;**34**:210-226

\*Address all correspondence to: anatoliyld@mail.ru

2 Far Eastern Federal University, Vladivostok, Russia

1 National Scientific Center of Marine Biology, Far Eastern Branch, Russian Academy of

**Figure 9.** The multikingdom phylogenetic hierarchical tree constructed on base of principle of conservatism (after

[1] Whittaker RH. On the broad classification of organisms. The Quarterly Review of

Sciences, A.V. Zhirmunsky Institute of Marine Biology FEB RAS, Vladivostok, Russia


[29] Drozdov AL, Kussakin OG. State of the art and problems of megasystematics. In: Kryukov AP, editor. Problems of Evolution. Collected papers. **5**. Vladivostok: Dalnauka; 2003. pp. 18-30

[17] Edwards P. A classification of plants into higher taxa based on cytological and biochemi-

[18] Ragan MA, Chapman A. Biochemical Phylogeny of the Protists. N.Y., San Francisco, L.: Academic Press; 1978. p. 317. Available from: https://books.google.ru/books?id=Bq qnNKmU8I8C&pg=PA257&dq=Ragan+MA,+Chapman+A.+Biochemical+phylogeny+o f+the+protists.+N.Y.,+San+Francisco,+L.&hl=ru&sa=X&ved=0ahUKEwja1JbS04HQAh UG8ywKHRZIClYQ6AEIVDAF#v=onepage&q=Ragan%20MA%2C%20Chapman%20 A.%20Biochemical%20phylogeny%20of%20the%20protists.%20N.Y.%2C%20San%20

[19] Starobogatov YI. The position of flagellated protists in the system of lower eukaryotes.

[20] Möhn E. System und Phylogenie der Lebewesen. 1. Physikalisch-chemische und biologische Evolution, Procaryonta, Eucaryonta (bis Ctenophora). Stuttgart: Schweizerbart'sche Verlagsbuchhandlung; 1984. p. 884 S. Available from: http://www.schweizerbart.de/ publications/detail/isbn/9783510651177/System\_und\_Phylogenie\_der\_Lebewesen

[21] Cavallier-Smith T. Kingdom protozoa and its 18 phyla. Microbiology Review. 1993;**57**:

[22] Cavallier-Smith T. Zooflagellate phylogeny and classification. Cytology (Цитoлoгия).

[23] Cavallier-Smith T. A revised six-kingdom system of life. Biological Reviews of Cambridge Philosophical Society 1998;**73**:203-266. Available from: https://books.google. ru/books?id=−GyARbyTobYC&pg=PA158&dq=Cavallier-Smith+T.+A+revised+six-kin gdom+system+of+life.+Biol.+Rev.+Comb.++Phylos.+Soc&hl=ru&sa=X&ved=0ahUKEw i1i\_jQ1YHQAhWIdCwKHRn4BY8Q6wEIHzAA#v=onepage&q=Cavallier-Smith%20 T.%20A%20revised%20six-kingdom%20system%20of%20life.%20Biol.%20Rev.%20

[24] Cavalier-Smith T. The phagotrophic origin of eukaryotes and phylogenetic classification of Protozoa. International Journal of Systematic and Evolutionary Microbiology.

[25] Cavalier-Smith T. The neomuran origin of archaebacteria, the negibacterial root of the universal tree and bacterial megaclassification. International Journal of Systematic and Evolutionary Microbiology. 2002;**52**:7-76. Available from: http://www.microbiologyresearch.org/docserver/fulltext/ijsem/52/1/0520007a.pdf?expires=1477802129&id=id&accn

[26] Vorontsov NN. Origin of life and diversity of its forms. Novosibirsk. 1965;55 (in Russian)

[27] Vorontsov NN. Systems of the organic world and the position of animals in them.

[28] Vorontsov NN. Systems of the organic world and the position of animals in them.

1. Russian Journal of Ecology. (Зooл. ж.). 1987;**66**:1668-1684 (in Russian)

2. Russian Journal of Ecology. (Зooл. ж.). 1987;**66**:1765-1774 (in Russian)

cal criteria. Taxon. 1976;**25**:529-542

Francisco%2C%20L.&f=false

953-994

24 Phylogenetics

1995;**37**:1010-1029

Cytology (Цитoлoгия). 1995;**37**:1030-1037

Comb.%20%20Phylos.%20Soc&f=false

2002;**52**:297-354. DOI: 10.1099/00207713-52-2-297

ame=guest&checksum=93DC34728A09636A6C799F9529E111FC


## **Phylogenetics for Wildlife Conservation**

## José Luis Fernández-García

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.69240

#### **Abstract**

[45] Leontiev DV, Akulov AY. Revolution in megataxonomy: Prerequisites and results.

[46] Cavalier-Smith T. The kingdom chromista. In: Green JC, Leadbeater BSC, Diver WC, editors. The Chromophyte Algae: Problems and Perspectives. Oxford: Oxford University

[47] Fox GE, Stackebrandt E, Hespel RB, Gibson J, Maniloff J, Dyer TA, Wolfe RS, Balch WE, Tanner RS, Magrum LJ, Zablen LB, Gupta BRR, Bonen L, Lewis BJ, Stahl DA, Luehrsen KR, Chen KN, Woese CR. The phylogeny of prokaryotes. Science. 1980;**209**:457-463 [48] Woese CR, Fox GE. Phylogenetic structure of the prokaryotic domain: the primary kingdoms. Proceedings of the National Academy of Sciences of the United States. 1977;**74**:

[50] Woese CR, Kandler O, Wheelis ML. Toward a natural system of organisms: Proposal for the domains Archaea, Bacteria, and Eucarya. Proceedings of the National Academy of

[51] Woese CR. A new biology for a new century. Microbiology and Molecular Biology

[52] Woese CR, Goldenfeld N. How the microbial world saved evolution from the scylla of molecular biology and the charybdis of the modern synthesis. Microbiology and

[53] Kandler O. Evolution of the systematic of bacteria. In: Schleifer KH, Stackebrand E. eds., Evolution of Prokaryotes. FEMS Symposium 29. L. etc: Academic Press; 1995. pp. 335-361

[54] Vossbrinck CR, Maddox JV, Friedman S, Debrunner-Vossbrinck BA, Woese CR. Ribosomal RNA sequence suggests microsporidia are extremely ancient eukaryotes.

[55] Keeling PL, Fast NM. Microsporidia: Biology and evolution of highly reduced intracellular parasites. Annual Review of Microbiology. 2002;**56**:93-116. DOI: 10.1146/annurev.

[56] Gill EE, Fast NM. Assessing the microsporidia-fungi relationship: Combined phyloge-

[57] Corradi N, Keeling PJ. Microsporidia: A journey through radical taxonomical revisions.

[58] Corsaro D, Walochnik J, Venditti D, Steinmann J, Müller K-D, Michel R. Microsporidialike parasites of amoebae belong to the early fungal lineage Rozellomycota. Parasitology

[59] Capella-Gutiérrez S, Marcet-Houben M, Gabaldón T. Phylogenomics supports microsporidia as the earliest diverging clade of sequenced fungi. BMC Biology. 2012;**10**:47. Available from: http://www.biomedcentral.com/1741-7007/10/47. DOI: 10.1186/1741-7007-10-47

Research. 2014;**113**. DOI: 10.1007/s00436-014-3838-4 Source: PubMed

Sciences of the United States. 1990;**87**:4576-4579. DOI: 10.1073/pnas.87.12.4576

Molecular Biology Reviews. 2009;**73**(1):14-21. DOI: 10.1128/MMBR.00002-09

Journal of General Biology (Ж. oбщ. биoл.). 2002;**63**:168-186 (in Russian)

[49] Woese CR. Bacterial evolution. Microbiology Review. 1987;**51**:221-271

Reviews. 2004;**68**:173-186. DOI:10.1128/MMBR.68.2.173-186.2004

netic analysis of eight genes. Gene. 2006;**375**:103-109

Press; 1989. pp. 379-405

Nature. 1987;**326**:411-414

micro.56.012302.160854

Fungal Biology Review. 2009;**23**:1-8

5088-5090

26 Phylogenetics

Recent extinctions and the continuing threats to the survival of rare species will make conservation biology crucial in the twenty-first century. Conservation genetics for wildlife is an emerging challenge for humanity because it is accepted that a number of species and its populations are under oppression by a huge human expansion. Conservation genetics is the science that aims to minimize the risk of extinction. The International Union for Conservation of Nature and Natural Resources (IUCN) recognizes three hierarchical levels to conserve biodiversity: genetic diversity (populations), species (taxon ascertainment), and ecosystems (living organisms and their interactions). In view of the world's imminent biodiversity crisis, the risk of extinction at several biotic levels is nowadays unavoidable and requires urgent action. One prime conservation goal is focusing on preserving the genetic variation. The main reasons are: (1) to preserve a representation of past evolution and (2) to maintain raw material for future evolution, favoring the balance of ecosystems. Having these aims in mind, a new approach utilizes different metrics, such as phylogenetic diversity, split distance, and heightened evolutionary distinctiveness, which are being considered for immediate practical use to manage threat species and stocks submitted to new policies for conservation.

**Keywords:** distinctiveness metrics, extinction risk, genetics wildlife management, phylogenetic and conservation, species diversity

## **1. Introduction**

*"In the face of inevitable future losses to biodiversity, ranking species by conservation priority seems more than prudent. Setting conservation priorities within species (i.e., at the population level) may be critical as species ranges become fragmented and connectivity declines."* [1]

Ever since the revolutionary ideas put forward by Darwin, the evolutionary perspective of wildlife has played a fundamental role and has aimed to the efficient protection and preservation of

© 2017 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

biological diversity, which started off with an adequate, accurate or, at least, the best approached inventory of its current status. But the recent extinction or continuing threats to the disappearance of many species and populations has made conservation biology essential in the twenty-first century. The primary forces concerned with its long-term persistence of wildlife populations, may be ecological, political, economic, or other. Nowadays, these forces (or factors) use more objective genetics principles and related applications for conservation. In particular, the application of new molecular techniques, widely used in conservation research, has made genetics examination of endangered species feasible. Conservation genetics for wildlife is an emerging challenge for humanity because it is generally accepted that the extinction of present species, even some of its populations, were caused by the huge expansion of a sole species, the man (*Homo sapiens*). So, the number of studies based on genetic data aimed at understanding biological diversity patterns and processes has increased in recent years, partially, because biodiversity assessments made using species counts (e.g., total, endemic, threatened) may not be the most suitable metrics. In consequence, a more reliable approach has been proposed to improve the situation. On the one hand, using genetic data and phylogenetic analysis to adequately represent the processes that gave rise to the observed patterns of diversity and, on the other hand, allowing conservation efforts to apply not only to threatened species, but also to other particularly interesting populations. The metrics to be employed is yet under debate and an agreement needs to be reached.

As we said above, conservation genetics is the science that aims to minimize the risk of extinction from genetic factors [2]. Conservation genetics has flourished over the last 20 years and has shown that there are many ways genetic knowledge can help to conserve biodiversity, ranging from identifying the concerned populations to resolving taxonomic uncertainties, or understanding the biology of a focal taxon. The International Union for Conservation of Nature and Natural Resources (IUCN) is also focused on these ideas and recognizes three hierarchical levels to conserve biodiversity: genetic diversity (populations), species (taxon ascertainment), and ecosystems (living organisms and their interactions).

Although it is reasoned that endangered species have deserved a noteworthy attention on conservation research [3], less concerned species are also of research and sometimes conservation interest (e.g., European red deer [4, 5]). So, every species are important especially the distribution of their particular isolated populations when they are genetically distinct, although by not well known reasons. In this last case (including minor concern species), it reaches relevant importance to those inferior levels of taxonomic arrangement as subspecies, an historical nomination concept that is being replaced by evolutionary significant units (ESU), management units (MUs), and distinct population segments (DPS). In this way of thinking, the intraspecific diversity is officially recognized as one of three levels of biodiversity. This level of diversity, coupled with ecosystems and whole genetic diversity is worthy of protection [6] but often require more adequate information [7] about concerned species, ESUs, MU, or DPS [8].

In view of the world's imminent biodiversity crisis, referred to, by some people as the 'sixth mass extinction' but different from the five previous ones, "the next extinctions will be due to human impact", which are now unavoidable and need urgent actions to prevent it. Nowadays, optimistic scenarios predict significant changes in biodiversity around 2100, with most of the loss starting with isolated populations of whichever wild species.

A large portion of the conservation genetics is dealing with the genetic conclusions about the causes and consequences of isolated small populations characterized by its low effective populations size (Ne), and simultaneously, the genetic drift effect because it causes a random change from generation to generation of gene pool. Whichever the case, they are both relevant issues associated to inbreeding under no random mating. The long-term effect of inbreeding leads to loss of genetic variability until reduced adaptability and ecosystem function, too [9].

Below the species level, it has been advocated the identification of populations that deserve long-term conservation or are derived from a recent rank fragmentation [10–12]. Although populations' relationships are being represented by bifurcating trees, it is known that bifurcating trees often fail to show everything and/or complex relationships, a major shortcoming if populations do need to be prioritized for conservation [1]. In this way of thinking, several studies have shown how measuring and maximizing phylogenetic diversity can be performed using phylogenetic networks and evolutionary isolation indices adapted for populations within species ([1, references therein). The new approach utilizes different metrics, like phylogenetic diversity (PD), split distance (SD) [13], or Shapley metric (SH) [14], and heightened evolutionary distinctiveness (HED) (refined by [15]) to assess not only from the species level, but also to population differentiations within each other. These metric might be of immediate practical use to manage discrete populations within species with several degrees of threat and stocks submitted to new policies for conservation triage [16].

## **2. The phylogenetic context**

biological diversity, which started off with an adequate, accurate or, at least, the best approached inventory of its current status. But the recent extinction or continuing threats to the disappearance of many species and populations has made conservation biology essential in the twenty-first century. The primary forces concerned with its long-term persistence of wildlife populations, may be ecological, political, economic, or other. Nowadays, these forces (or factors) use more objective genetics principles and related applications for conservation. In particular, the application of new molecular techniques, widely used in conservation research, has made genetics examination of endangered species feasible. Conservation genetics for wildlife is an emerging challenge for humanity because it is generally accepted that the extinction of present species, even some of its populations, were caused by the huge expansion of a sole species, the man (*Homo sapiens*). So, the number of studies based on genetic data aimed at understanding biological diversity patterns and processes has increased in recent years, partially, because biodiversity assessments made using species counts (e.g., total, endemic, threatened) may not be the most suitable metrics. In consequence, a more reliable approach has been proposed to improve the situation. On the one hand, using genetic data and phylogenetic analysis to adequately represent the processes that gave rise to the observed patterns of diversity and, on the other hand, allowing conservation efforts to apply not only to threatened species, but also to other particularly interesting populations. The metrics to be employed is yet under debate and an agreement needs to be reached.

28 Phylogenetics

As we said above, conservation genetics is the science that aims to minimize the risk of extinction from genetic factors [2]. Conservation genetics has flourished over the last 20 years and has shown that there are many ways genetic knowledge can help to conserve biodiversity, ranging from identifying the concerned populations to resolving taxonomic uncertainties, or understanding the biology of a focal taxon. The International Union for Conservation of Nature and Natural Resources (IUCN) is also focused on these ideas and recognizes three hierarchical levels to conserve biodiversity: genetic diversity (populations), species (taxon

Although it is reasoned that endangered species have deserved a noteworthy attention on conservation research [3], less concerned species are also of research and sometimes conservation interest (e.g., European red deer [4, 5]). So, every species are important especially the distribution of their particular isolated populations when they are genetically distinct, although by not well known reasons. In this last case (including minor concern species), it reaches relevant importance to those inferior levels of taxonomic arrangement as subspecies, an historical nomination concept that is being replaced by evolutionary significant units (ESU), management units (MUs), and distinct population segments (DPS). In this way of thinking, the intraspecific diversity is officially recognized as one of three levels of biodiversity. This level of diversity, coupled with ecosystems and whole genetic diversity is worthy of protection [6] but often require more adequate information [7] about concerned species, ESUs, MU, or DPS [8]. In view of the world's imminent biodiversity crisis, referred to, by some people as the 'sixth mass extinction' but different from the five previous ones, "the next extinctions will be due to human impact", which are now unavoidable and need urgent actions to prevent it. Nowadays, optimistic scenarios predict significant changes in biodiversity around 2100, with most of the

ascertainment), and ecosystems (living organisms and their interactions).

loss starting with isolated populations of whichever wild species.

The concepts of taxonomy are familiar for every biologist because they have spent a long time studying species names and retrospectively their order into genus, family, …, kingdoms. Such a classification recalls a scenario like ancestor-descendent relationships among taxa (phylogeny), which result in a scheme describing an evolutionary relationship that could not be subject to critical analysis. Recently, modern phylogenetic science captures, as empirically as possible, the relatedness among similar taxa using the most orderly manner for mapping the path of evolution that leads to and represents the true ancestry relating the upstream organisms. The resultant classification must be reasonably and objectively assumed by worldwide biologists, undoubtedly. In this way, groups of species or its populations are essentially related by a set of both, morphological and molecular characteristics but, more importantly yet, these should be matched by properties such as its ecological abilities.

Firstly, phylogenetic studies have been proven to be of utility, of course, but in a research-oriented framework. In this way, a simple data research can provide guidelines to find gaps and strengthen interpretations to ensure management affirmations. So, multi-locus phylogenies can be used to infer the species tree whose nodes represent the actual separation between species, thus providing essential information about their evolutionary history or helping analyzes of species delimitation, gene flow, and genetic differentiation within species [17]. As an example, now adequate markers are available by extracting intron information from genomes of human, chimpanzee, macaque, cow, and dog (three mammalian orders) searching for the ENSEMBL database. This analysis led to a final list of 224 intron markers randomly distributed along the genome for six mammals species, which can be useful to gather genetic markers with unambiguous phylogenetic signals (see [17] for details and design) (**Figure 1**).

Secondly, the use of phylogenetic diversity is of current interest in view of its objective metrics for conservation in evolution history (the past), genetic status of species (the present), and

**Figure 1.** Steps for intron extractions and filtering processes. Adapted from [17].

management for conservation in geographically split species (the future).The first two may be of general interest on research, but within a practical approach the last issue is of plentiful applicability to wildlife population management. The phylogenetic ramifications reflect more than simple systematic classifications. The molecular information and its association with other kinds of data can be an objective measure to identify species or population groups with different or similar vital aptitude such as habitat use among taxa or similar facts. A straightforward example has been pointed out in the case of strong associations between habitats and morphology in shorebirds, ducks, and other water bird species. However, supposedly described subspecies differentiation (e.g., the specimens of the whole geographic Iberian range was pooled as a single genetic population instead of delimiting them as lineage clusters) based on morphological information has been seen to fail, probably due to the mixing of genetic lineages. After a molecular survey of the Iberian desman (*Galemys pyrenaicus*), the data set suggested two main phylogenetic clusters delimited by mitochondrial DNA (**Figure 2**) in this emblematic species. Because of a strong geographic splitting in type localities of this species and the absence of clear morphological discrimination with nowadays data, its populations may easily be regrouped in two big clades that would correspond to two nominal subspecies *Galemys pyrenaicus rufulus* (clade A) and *Galemys pyrenaicus pyrenaicus* (clade B) [18]. Consequently, it has recently been suggested to treat these outstanding lineages as separated groups in the wildlife management contexts.

**Figure 2.** Main lineages in *Galemys pyrenaicus*. Adapted from [18].

ENSEMBL database. This analysis led to a final list of 224 intron markers randomly distributed along the genome for six mammals species, which can be useful to gather genetic markers with unambiguous phylogenetic signals (see [17] for details and design) (**Figure 1**).

30 Phylogenetics

Secondly, the use of phylogenetic diversity is of current interest in view of its objective metrics for conservation in evolution history (the past), genetic status of species (the present), and

**Figure 1.** Steps for intron extractions and filtering processes. Adapted from [17].

Thirdly, however, is the issue of hybridization: a cause for debate. Hybridizations have occurred for long and they are well known by managers and scientists around the world. The main question about hybridization is which, the species or its hybrid, should be prioritized and valued. The concept of hybridization understood to mean mating between different species has been extended to mating between two genetically distinct populations that produce offspring (F1 to several backcross; **Figure 3**), regardless of its fertility.

Two competing effects of such introgression are assumed but with different final results on species diversity: (1) **a negative view** is a feeling of concern when human activity is the main cause of the introgression [19] and (2) **a positive view** is when nature is the main responsible of admixture among populations but with a long-term component [20] because, at present, man intervention is in everything, so, consequently, the first view is the one that is considered of most concern.

One well-studied example about the negative effect of human impact on hybridization in wildlife in nonthreatened species is the European red deer. During the last century (past and currently also), there has been an extensive arbitrary trading of European red deer aimed at breeding improved trophies for hunting on extinct or nearly extinct autochthonous populations [21]. The direct consequence of the restocking and the action of introducing geneticallydistinct populations has had various types of negative effects. On the one hand, hybridization with introduced animals has impaired the phylogenetic boundaries between former and natural populations, contributing to blurring true genetic history and confounding future researches. Worldwide allochthonous and indigenous red deer have been admixed (and are) through several Europe countries. It is believed that the scarce documentation about this fact is opposite to the true dimension of human impact, which should have been huge instead. Because of a generalized worldwide impact of anthropic action, a mixture of phylogenetic scenarios would probably be expected (**Figure 4**). Accordingly, though genetic variation is supposedly structured hierarchically, some exceptions occurred under hybridization associated to human activity. To overcome this drawback, an effective sampling strategy according to the specific problem should be design based on knowledge. In the European red deer example, due to the arbitrariness of admixture, these scenarios caused different effects. One of them may be the presence of mixture allochthonous lineages as in Val di Susa (Italy) being genetically similar to Bulgarian red deer. Although the origin of Val di Susa red deer was

**Figure 3.** The most probable distribution of hybrid and their backcross in a natural framework of admixture.

Thirdly, however, is the issue of hybridization: a cause for debate. Hybridizations have occurred for long and they are well known by managers and scientists around the world. The main question about hybridization is which, the species or its hybrid, should be prioritized and valued. The concept of hybridization understood to mean mating between different species has been extended to mating between two genetically distinct populations that produce

Two competing effects of such introgression are assumed but with different final results on species diversity: (1) **a negative view** is a feeling of concern when human activity is the main cause of the introgression [19] and (2) **a positive view** is when nature is the main responsible of admixture among populations but with a long-term component [20] because, at present, man intervention is in everything, so, consequently, the first view is the one that is considered

One well-studied example about the negative effect of human impact on hybridization in wildlife in nonthreatened species is the European red deer. During the last century (past and currently also), there has been an extensive arbitrary trading of European red deer aimed at breeding improved trophies for hunting on extinct or nearly extinct autochthonous populations [21]. The direct consequence of the restocking and the action of introducing geneticallydistinct populations has had various types of negative effects. On the one hand, hybridization with introduced animals has impaired the phylogenetic boundaries between former and natural populations, contributing to blurring true genetic history and confounding future researches. Worldwide allochthonous and indigenous red deer have been admixed (and are) through several Europe countries. It is believed that the scarce documentation about this fact is opposite to the true dimension of human impact, which should have been huge instead. Because of a generalized worldwide impact of anthropic action, a mixture of phylogenetic scenarios would probably be expected (**Figure 4**). Accordingly, though genetic variation is supposedly structured hierarchically, some exceptions occurred under hybridization associated to human activity. To overcome this drawback, an effective sampling strategy according to the specific problem should be design based on knowledge. In the European red deer example, due to the arbitrariness of admixture, these scenarios caused different effects. One of them may be the presence of mixture allochthonous lineages as in Val di Susa (Italy) being genetically similar to Bulgarian red deer. Although the origin of Val di Susa red deer was

**Figure 3.** The most probable distribution of hybrid and their backcross in a natural framework of admixture.

offspring (F1 to several backcross; **Figure 3**), regardless of its fertility.

of most concern.

32 Phylogenetics

**Figure 4.** Phylogeography scenario of European red deer lineages. A = "Western European red deer" lineages, B = C-BRD "Corsican and Barbary red deer" lineages, and C = ERRD "Eastern European red deer" lineages. Map showing natural geography distribution of lineages. Network showed some restocked lineages into different areas of Spain and Europe (yellow quadrate).

Slovenia and Bulgaria; only Bulgarian blood survived, probably attributed to genetic drift. But in this case, the population could be easily qualified as allochthonous. On the other hand, outbreeding depression of the hybrid offspring due to lower reproductive success or survival of either parent has also been found. In our example, translocation of Wapitis and Asian red deer (today regarded as different species from European red deer) was unsuccessful by far, as a way to result in antler-size improvement. This failure was partially due to the **lack of adaptation** to local environmental factors or high susceptibility to local diseases. This example suggested outbreeding depression in hybrid populations [21]. Two mechanisms have been proposed for the outbreeding depression. An **intrinsic mechanism** upholds a reduced fitness of hybrids due to interactions between genes originating in different evolutionary taxa. Conversely, **extrinsic mechanisms** advocated for loss of adaptation to local environment with unsuccessful reproduction. Also, the interaction genotype-environment may be assessed.

Moreover, hybridized populations or species may consist on a hybrids swarm in which all individuals are to various degrees of admixture. In this respect, an important role in transferring or restocking species or populations to the wild is being played by enclosures (in zoos or collections), which serve as reservoirs of different populations and subspecies. Sometimes, these reservoirs have acted as the origin of feral populations of many different exotic species and subspecies contaminating autochthonous stocks. This was the case of the Woburn red deer from Bedfordshire [21] or the Mesopotamian fallow deer at the Opel Zoo [22]. In the latter case, phylogenetic studies can be used to assay the presence of hybridization in the Persian fallow deer from the Israeli Reintroduction Program started in 1996 and thus dispel all doubts.

The positive side of hybridization is more related to speciation. Hybridization occurs more frequently than previously recognized and is an important source of speciation. Hybridization leading to a new taxon, distinct from both parent species, is called (when homoploid) hybrid speciation or recombinational speciation [23]. Almost 50% of plant species originated from the hybridization of different species. For example, 10% of bird species are believed to hybridize with another species naturally. This sort of speciation promoted adaptive divergence and increased reproductive isolation. But introgressed genetic variation can also enhance the ability to coexist and promote invasiveness [24] enlarging the range of a hybrid populations. Moreover, a positive feedback between hybridization and speciation may exist [25]. So, hybridization may increase (1) the rate of speciation, (2) diversity of closely related species, and (3) adaptive radiation by incorporation into populations of selectively favored alleles or combinations of them; providing the basis for adaptive evolution and having important implications for the origin of new species.

The frequency of hybridization as a source of adaptive variation for speciation may be summarized as follows: firstly, hybridization among species occur about 10–30% of multicellular species regularly on a per-species basis but less frequently on a per-individual basis, the latter more frequently driven by humans (as the case of *Dama dama mesopotamica* described in [22]). Secondly, mutations are rare, around 10−8 to 10−9 per generation per base pair, that is, a considerable time for novel adaptations to appear but depending also on the population size. So, hybridization among species can act as a source of adaptive genetic variation rather than mutation [26–30]. For example, 'New additive genetic variance introduced by hybridization in Darwin's finches, which has been estimated to be two to three orders of magnitude greater than that introduced by mutation' [26], despite initial hybridization itself, which is unlikely to be adaptive because there is often evidence of selected against. Last but not least, adaptation is thought to be the most important process driving divergence during speciation [31–33] and divergence in ecology occurs almost exclusively under selection. Moreover, closely related species tend to hybridize more often. Species in rapidly diversifying adaptive radiations could especially be prone to hybridization [25, 34, 35].

#### **3. Conservation genetics**

Conservation genetics was born in the last third of the twentieth century integrating empirical and theoretical studies based on population genetic data, which were incorporated to the Conservation Biology doctrine giving rise to the discipline "Conservation Genetics" with a spectacular growth. The conceptual framework included all "genetics" issues that are phylogenetic, quantitative, evolutionary, ecological, and population genetics themes.

Nowadays, conservation genetics is being applied for practical conservation and wildlife management as a major paradigm. At first, the conservation of species was evaluated by indirect and phenotypic data but powerful advances on DNA technology resulted in a huge amount of genetic data more easily achieved, and also helped by an emerging sophisticated statistical procedures. Now, it is possible to gather the objective information coded long ago into genomes of every organism. Thereafter, the conservation genetic discipline raised its interest when people became aware of the growing rate of human population and its unavoidable effect on planet biodiversity. The IUCN (World Conservation Union, formerly International Union for Conservation of Nature diversity either ecosystems or species) recognized three main levels worthy of protection and conservation: genetic diversity within species, species in themselves, and either local or global ecosystems. However, the first goal in the mind of conservation geneticist is the assessments of genetic variability in threatened and unthreatened organisms as a metric to trace the well-being of the planet.

#### **3.1. Relevant items in conservation genetics: wildlife scenario from top to bottom**

The positive side of hybridization is more related to speciation. Hybridization occurs more frequently than previously recognized and is an important source of speciation. Hybridization leading to a new taxon, distinct from both parent species, is called (when homoploid) hybrid speciation or recombinational speciation [23]. Almost 50% of plant species originated from the hybridization of different species. For example, 10% of bird species are believed to hybridize with another species naturally. This sort of speciation promoted adaptive divergence and increased reproductive isolation. But introgressed genetic variation can also enhance the ability to coexist and promote invasiveness [24] enlarging the range of a hybrid populations. Moreover, a positive feedback between hybridization and speciation may exist [25]. So, hybridization may increase (1) the rate of speciation, (2) diversity of closely related species, and (3) adaptive radiation by incorporation into populations of selectively favored alleles or combinations of them; providing the basis for adaptive evolution and having important

The frequency of hybridization as a source of adaptive variation for speciation may be summarized as follows: firstly, hybridization among species occur about 10–30% of multicellular species regularly on a per-species basis but less frequently on a per-individual basis, the latter more frequently driven by humans (as the case of *Dama dama mesopotamica* described in [22]). Secondly, mutations are rare, around 10−8 to 10−9 per generation per base pair, that is, a considerable time for novel adaptations to appear but depending also on the population size. So, hybridization among species can act as a source of adaptive genetic variation rather than mutation [26–30]. For example, 'New additive genetic variance introduced by hybridization in Darwin's finches, which has been estimated to be two to three orders of magnitude greater than that introduced by mutation' [26], despite initial hybridization itself, which is unlikely to be adaptive because there is often evidence of selected against. Last but not least, adaptation is thought to be the most important process driving divergence during speciation [31–33] and divergence in ecology occurs almost exclusively under selection. Moreover, closely related species tend to hybridize more often. Species in rapidly diversifying adaptive radiations

Conservation genetics was born in the last third of the twentieth century integrating empirical and theoretical studies based on population genetic data, which were incorporated to the Conservation Biology doctrine giving rise to the discipline "Conservation Genetics" with a spectacular growth. The conceptual framework included all "genetics" issues that are phylo-

Nowadays, conservation genetics is being applied for practical conservation and wildlife management as a major paradigm. At first, the conservation of species was evaluated by indirect and phenotypic data but powerful advances on DNA technology resulted in a huge amount of genetic data more easily achieved, and also helped by an emerging sophisticated statistical procedures. Now, it is possible to gather the objective information coded long ago into genomes of every organism. Thereafter, the conservation genetic discipline raised its interest

genetic, quantitative, evolutionary, ecological, and population genetics themes.

implications for the origin of new species.

34 Phylogenetics

could especially be prone to hybridization [25, 34, 35].

**3. Conservation genetics**

#### *3.1.1. Kinship and genetic variation for within population conservation (population genetics)*

The loss of genetic variation due to inbreeding (as a result of mating among genetically related individuals) was (and is yet) the main issue regarding captive and natural populations of small size. Whichever the case, despite great scientific attention received by the deleterious effects arising from inbreeding depression; no less important are parentage, kinship, sex identification, and demographic history of population. Since a general scientific acknowledgement regarding inbreeding depression related to small captive populations and natural isolated populations as well, a preference position has been granted to those studies focused on inbreeding depression. The assessing of inbreeding depression has been the former issue in the design of conservation programs, formerly applied to domestic animals and plants, but today it has been extended to wildlife, both in captive breeding programs and in the management of natural isolated populations. An interesting case is the Pyrenean desman (*G*. *pyrenaicus*), which is annotated as vulnerable by the IUCN red list. However, the southernmost population in the Iberian peninsula (at the mountain place of the central system: green dashes in **Figure 2**) is listed as "endangered" with high extinction risk by the main Spanish government authority (MAGRAMA, that is, Ministry of Agriculture, Food and Environment) due to its almost null genetic variation (mtDNA studies suggested they carried a clonal lineage in several populations) and high level of anthropic threat but without possibility of implementing captive breeding programs [18].

Regarding population variations in wildlife, it is important to assess local kinship as offspring parentage, mating systems, sex determination, or lineages identification. The main field of study is the application of empirical data to be compared with theoretical assumptions as in the case of diploid lethal equivalents estimation to juvenile survival [36]. New DNA technologies are addressing molecular procedures to gather high informative loci as microsatellites and single nucleotide polymorphisms (SNPs) to finely estimate relatedness coefficient at several degrees of relatives, not only to parents-offspring pairs. An underestimate of the total impact of inbreeding has been declared and Ne/N bias between nonbreed and unmanaged wild population has been claimed after assuming statistical distribution of family size (Poisson distribution). The relative importance of the analysis of local kinships has several issues as follows: (i) isolated populations differ by drift and inbreeding but the first is more related to random sampling than specifically mating of relatives; (ii) balance among family sizes can be calculated by molecular procedures using as many genetic markers as possible in local or isolated populations; (iii) local populations exhibited correlations between diversity and family sizes but unbalances in this last one may influence minimum viable populations size(MVPs) (number of individuals needed for long-term persistence of populations with high probability), which assist scientific and wildlife managers in population viability analysis (PVA). However, some discrepancies arose between theoretical and empirical studies comparison about the deleterious effect of inbreeding, suggesting a case-by-case analysis in wild species due to strong species specific conditionings: lifestyle, demographic history, genetics, and more. Other significant assessment related to conservation is heterozygosity. It may be useful to understand a species life history. This type of analysis allows us to give a retrospective look at the past to make current comparisons and to perform realistic predictions about the future.

#### *3.1.2. Conservation genetic of geographic variation*

Ecological and evolutionary sources of genetic variation, at the intra-specific level or higher, are also worth of being considered for conservation purposes. However, the two main areas of work in this broad field of study have its top representatives in phylogeography and genetics of populations. These two approaches are being used currently, one based on allelic frequencies (unordered polymorphism for population genetics but recently also phylogeography using e.g., network-net methods) and the other one based on mitochondrial DNA sequences (ordered polymorphism for phylogeography also using networks as in **Figure 4**). Both approaches are utilized because they can easily represent the pattern of spatial distribution of genetic variation for species and allocate the most genetically isolated populations and connectedness degree if any. Moreover, there is a current tendency to rejoin historical genealogical information plus contemporary forces modeling populations because it is believed to provide a larger resolution to illuminate the causes and consequences of such spatial pattern in nature. Moreover, practical biodiversity conservation is interested in conserving as many species or relevant populations within them allocated inside emblematic or unique places following the "species' genetic richness" concept.

At least, three competing concepts that connect researchers on conservation genetics and conservation managers, also needing to be delimited in the conservation biology context, are those that follow: evolutionary significant units (ESUs), managements units (MUs), and phylogenetic diversity (PD) of taxa as a way to estimate distinct population segments (DPS) [1]. Today, these three concepts are fully applicable for wildlife analysis and to take relevant decisions. The idea of MUs should be seen regardless of how recent the prior genetic history connections was, providing that exchange of individuals is so small as to be demographically independent units. By contrast, ESUs must imply a long historical separation of its populations. However, approaches based on demography and connectedness between populations can treat species or populations unequally. Consequently, a new appraisal introduces evaluations of phylogenetic trees connecting species (or populations within species) under a study approach called "Phylogenetic Diversity of taxa" (PDs). This approach has into account the edge length distances of the tree. Edge lengths depict the optimal number of features uniquely shared by all descending taxa below this edge and using a root. The set of taxa (populations) that maximizes the PD (normally less than the total populations considered) could be utilized in two types of projects. It has been employed to identify taxa and/or populations prone for conservation purposes. On the other hand, it is also a way to identify important taxa or geographically isolated for sequencing projects.

#### *3.1.3. Biodiversity of species*

and family sizes but unbalances in this last one may influence minimum viable populations size(MVPs) (number of individuals needed for long-term persistence of populations with high probability), which assist scientific and wildlife managers in population viability analysis (PVA). However, some discrepancies arose between theoretical and empirical studies comparison about the deleterious effect of inbreeding, suggesting a case-by-case analysis in wild species due to strong species specific conditionings: lifestyle, demographic history, genetics, and more. Other significant assessment related to conservation is heterozygosity. It may be useful to understand a species life history. This type of analysis allows us to give a retrospective look at the past to make current comparisons and to perform realistic predictions about

Ecological and evolutionary sources of genetic variation, at the intra-specific level or higher, are also worth of being considered for conservation purposes. However, the two main areas of work in this broad field of study have its top representatives in phylogeography and genetics of populations. These two approaches are being used currently, one based on allelic frequencies (unordered polymorphism for population genetics but recently also phylogeography using e.g., network-net methods) and the other one based on mitochondrial DNA sequences (ordered polymorphism for phylogeography also using networks as in **Figure 4**). Both approaches are utilized because they can easily represent the pattern of spatial distribution of genetic variation for species and allocate the most genetically isolated populations and connectedness degree if any. Moreover, there is a current tendency to rejoin historical genealogical information plus contemporary forces modeling populations because it is believed to provide a larger resolution to illuminate the causes and consequences of such spatial pattern in nature. Moreover, practical biodiversity conservation is interested in conserving as many species or relevant populations within them allocated inside emblematic or unique places fol-

At least, three competing concepts that connect researchers on conservation genetics and conservation managers, also needing to be delimited in the conservation biology context, are those that follow: evolutionary significant units (ESUs), managements units (MUs), and phylogenetic diversity (PD) of taxa as a way to estimate distinct population segments (DPS) [1]. Today, these three concepts are fully applicable for wildlife analysis and to take relevant decisions. The idea of MUs should be seen regardless of how recent the prior genetic history connections was, providing that exchange of individuals is so small as to be demographically independent units. By contrast, ESUs must imply a long historical separation of its populations. However, approaches based on demography and connectedness between populations can treat species or populations unequally. Consequently, a new appraisal introduces evaluations of phylogenetic trees connecting species (or populations within species) under a study approach called "Phylogenetic Diversity of taxa" (PDs). This approach has into account the edge length distances of the tree. Edge lengths depict the optimal number of features uniquely shared by all descending taxa below this edge and using a root. The set of taxa (populations) that maximizes the PD (normally less than the total populations considered) could be utilized in two types of projects. It has been employed to identify taxa and/or populations prone for

the future.

36 Phylogenetics

*3.1.2. Conservation genetic of geographic variation*

lowing the "species' genetic richness" concept.

In this section, the basic idea is that unique evolutionary lineages may contribute largely to overall genetics diversity. Their extinction would constitute a far great loss of diversity than would the extinction of species that have extant close relative. Although under discussion, phylogenetic distinctiveness is dealing with resolution of taxonomic issues due to its recognized role as measurement of taxon worthy of investing conservation resources. It is generally admitted that the importance of research to delineate the influence of introgression and hybridization on species diversity. It is a topic that is reaching great relevance at the interspecific level but also at the inter-subspecific level as a way of **silent extinction** due to human domestication of current wild species and random translocation of their products, overriding yet hidden evolutionary pathways unexpectedly by introgressive extinction (as for red deer subspecies, **Figure 4**).

In this state of things, the systematic evaluation focusing on elevating differentiated populations such as true species assessed by informative genetic loci of split populations. This is an important issue for wildlife conservation and for making management decisions. Several subspecies gathered the rank of species (e.g., historical nominal subspecies as Wapiti but today elevated to the species level: *Cervus canadensis* instead of *Cervus elaphus canadensis*) or at least will be considered from this point worth of deepest studies as Mesopotamian fallow deer (*Dama dama mesopotamica*) or Barbary red deer (*Cervus elaphus barbarus*), which are currently included in their respective conservation programs or even herdbooks. Hopefully, conservation programs and the creation of herdbooks to manage the most endangered species should be treated as a nonnegligible new Genetic discipline: "Domestic" wildlife issue. Nevertheless, it should not be obviate that the rate of speciation, diversity of closely related species and adaptive radiation by incorporation into populations of selectively favorable alleles or combinations of them may be increased by hybridization, providing thus, the basis for adaptive evolution and having important implications for the origin of species, as mentioned previously.

#### *3.1.4. Wildlife forensic: the case of Pyrenean desman in ecological studies*

Forensic identification by advanced DNA technology is also important for wildlife studies. But forensic analysis has several distinct fields of application. Firstly, free-ranging wildlife species, especially those endangered, where noninvasive methods are recommended to detect elusive or sensitive to human management species as sampling strategies (e.g., Pyrenean desman (*G*. *pyrenaicus*)). On the other hand, wildlife products from specimens under strict police management due to them are imperiled (e.g., rhinoceros horns).

Finally, the biology and ecology of species with elusive or with hidden activity, which are still poorly known. As an example, the nature of trophic interactions is a fundamental issue in ecology and has aroused the attention of biologists for decades. This knowledge is particularly important in endangered species such as the Pyrenean desman. Using DNA from feces of the Pyrenean desman, it is possible to identify 19 prey species by next generation sequencing methods like the DNA minibarcode (133 bp) of the COI gene barcoding. This tool is able to simultaneously perform screening of species at large-scale because sometimes feces could be difficult to identify directly. Despite potential pitfalls in this methodology, it is based on one or a few genes at present state, each new genome incorporated into the data bank increases the validity of it. Consequently, more and more literature is arising in recent times.

## **4. Biodiversity analysis by integrating phylogeny and conservation**

Quantification of biodiversity using phylogenetic analyzes has been proposed to provide a more objective framework to make conservation decisions. Three collaborative efforts among ecologists, evolutionary biologists, paleontologists, systematists, and conservation biologists from the USA, Canada, Australia, and England are driving these aims thorough the 'Tree of Life' project attempting to integrate phylogenetic and conservation biology. They are based on two complementary facts: (1) surprising amounts of phylogenetic diversity might remain even under high rates of extinction (random) and (2) it is feasible to detect current extinction events through missing phylogenetic diversity as it is mentioned in [37].

Three issues are being examined in the integrative framework as follow:

#### **4.1. Selectivity or random extinctions questions: species or upper taxonomic level**

The start viewpoint of New and May's was that simulated extinction occurred at random with respect to phylogeny [37]. However, phylogeny and conservation working groups ('phylogeny and conservation' working group sponsored by the National Center for Ecological Analysis and Synthesis (NCEAS) in Santa Barbara, CA, USA) reasoned that in this context randomness is not realistic due to extinctions and invasions tend to be strongly clumped for the most diverse taxonomic groups, for example, mammals or birds. After testing several statistics by simulation, the Moran's *I* index showed the high performance for detecting selectivity accurately, independent of tree size (i.e., number of species), tree shape (i.e., nodes with equal size groups in the tree), or prevalence of the desired trait (e.g., proportion of endangered or invasive species). As a result, it has been recognized that taxonomic selectivity is the main way to extinction and could be quantified, but hopefully selectivity varies across a wide variety of taxonomic groups, across geographical regions, between 'higher' and 'lower' taxonomic units, and extinction is related to selectivity for invasion within taxonomic groups.

#### **4.2. Levels below species**

A long-standing problem is how to designate conservation units below the species level: Subspecies, ESUs, MUs, and more. With the advent of molecular technologies, those historical concepts as "subspecies" fell in disuse. However, an overload of genetic information can lead to the designation of many small and isolated subunits hampering the standard delimitation of, for example, the ESU and MUs concepts. A survey of the recent literature revealed that most studies follow the guidelines of Moritz, which advocate a purely genetic definition of ESUs. Nevertheless, a large fraction of conservation decisions require both genetic and ecological evidence.

methods like the DNA minibarcode (133 bp) of the COI gene barcoding. This tool is able to simultaneously perform screening of species at large-scale because sometimes feces could be difficult to identify directly. Despite potential pitfalls in this methodology, it is based on one or a few genes at present state, each new genome incorporated into the data bank increases

Quantification of biodiversity using phylogenetic analyzes has been proposed to provide a more objective framework to make conservation decisions. Three collaborative efforts among ecologists, evolutionary biologists, paleontologists, systematists, and conservation biologists from the USA, Canada, Australia, and England are driving these aims thorough the 'Tree of Life' project attempting to integrate phylogenetic and conservation biology. They are based on two complementary facts: (1) surprising amounts of phylogenetic diversity might remain even under high rates of extinction (random) and (2) it is feasible to detect current extinction

the validity of it. Consequently, more and more literature is arising in recent times.

**4. Biodiversity analysis by integrating phylogeny and conservation**

events through missing phylogenetic diversity as it is mentioned in [37]. Three issues are being examined in the integrative framework as follow:

nomic groups.

38 Phylogenetics

**4.2. Levels below species**

**4.1. Selectivity or random extinctions questions: species or upper taxonomic level**

The start viewpoint of New and May's was that simulated extinction occurred at random with respect to phylogeny [37]. However, phylogeny and conservation working groups ('phylogeny and conservation' working group sponsored by the National Center for Ecological Analysis and Synthesis (NCEAS) in Santa Barbara, CA, USA) reasoned that in this context randomness is not realistic due to extinctions and invasions tend to be strongly clumped for the most diverse taxonomic groups, for example, mammals or birds. After testing several statistics by simulation, the Moran's *I* index showed the high performance for detecting selectivity accurately, independent of tree size (i.e., number of species), tree shape (i.e., nodes with equal size groups in the tree), or prevalence of the desired trait (e.g., proportion of endangered or invasive species). As a result, it has been recognized that taxonomic selectivity is the main way to extinction and could be quantified, but hopefully selectivity varies across a wide variety of taxonomic groups, across geographical regions, between 'higher' and 'lower' taxonomic units, and extinction is related to selectivity for invasion within taxo-

A long-standing problem is how to designate conservation units below the species level: Subspecies, ESUs, MUs, and more. With the advent of molecular technologies, those historical concepts as "subspecies" fell in disuse. However, an overload of genetic information can lead to the designation of many small and isolated subunits hampering the standard delimitation of, for example, the ESU and MUs concepts. A survey of the recent literature revealed that most studies The guidelines of Moritz admitted that ESUs should show significant divergence and reciprocal monophyly for mtDNA and significant divergence of allele frequencies at nuclear loci. This is straightforward because it requires to examine historical and recent restrictions to gene flow, that is, evidence for long-term divergence that continued in the mtDNA and nuclear loci (free from selection) where mutations accumulate relatively more slowly or very rapidly, respectively. Therefore, this molecular discrepancy is useful to evaluate restrictions to gene flow at different times or even detecting genetic distinctiveness but no adaptive potential. However, Moritz's definition no-longer mentions the ecological distinctness because ecological divergence may or may not be necessarily associated to genetic divergence. Crandall et al. [8] proposed the "cross-hair analysis" to have into account the four important scenarios to decide whether ESUs or not is present (**Figure 5**).

Consequently, there is a worldwide agreement that decisions should be based on both genetic and ecological evidence but in the context of ecological and genetic exchangeability. Ecology together with an examination of recent and historical processes provide a more fine-grained, and therefore, more flexible categorization than the current system to be employed to diverse set "case studies" as red wolf (*Canis rufus*), dusky seaside sparrow (*Ammodramus maritimus nigrescens*), Florida panther (*Puma concolor coryi*), and Gila topminnow (*Poeciliopsis occidentalis occidentalis*) or Pyrenean desman (*G*. *pyrenaicus*).

#### **4.3. Areas with distinct population segments (DPS): hotspots places**

There are many criteria to determine the relative conservation value of different areas (e.g., species, threatened species or large numbers of species across different groups), but now below species level as areas containing DPS received attention since the late 1980s mainly for economic-important taxa. The problem is how we can quantify DPS value. Population-based management is being a necessary task for scientists and managers due to climate change and habitat degradation associated to growing human demands ensuring continued speciesrange fragmentation, which will be expected during this century. In order to address this work, phylogenetic diversity (PD) is being used as a measure of at least three stuffs to choose important areas to protect with accountability incorporating phylogenetic information.

Firstly, the exploration of the relations between PD and the spatial distribution of biodiversity would permit to get insight into the population structure complementary to the current statistical assessment of differentiation employed by MUs and DPS. Moreover, under this perspective, it is feasible, when constrained, to choose only a limited number of areas for conservation, to develop appropriate protocols to assess the complementarity predictions to preserve future biodiversity. Secondly, PD is being extended to simulations aimed to find taxonomically nonrandom extinction risk. Current threat scenarios are tested by comparing the spatial distribution of PD both before and after projected extinction. Finally, the predictions that suggest rapid environmental change leads to explore whether phylogenetic patterns of threat could predict the amount of ecological disturbance in a region.


**Figure 5.** Cross-hair analysis for management recommendations (adapted from [8]).

## **5. Prioritizing populations for conservation using phylogenetic distances from networks: split diversity (SD)**

According to the "species richness" concept [38], practical biodiversity conservation has the aim to preserve as many species as possible. However, as previously said, such an approach has the hurdle of treating all species equally [39]. However, neither all species nor genetic lineages are equally important, with more isolated lineages providing more important contribution to total variation, that is, the base for identifying populations worthy of protection in law. Genetic variation is depicted perfectly onto a rooted phylogenetic tree, where the edge length represents the number of features uniquely shared by all descending taxa, say populations. Importantly, ESUs concept assumes that the relationships among populations can be represented by a bifurcating tree. However, these sort of phylogenetic trees often fail to capture complete genetic information among populations. Moreover, more complex interrelationships are expected for DPSs and MUs. So, it would seem a shortcoming could occur if populations do need to be prioritized for conservation on the basis of tree-based prioritization schemes. However, the prioritization approaches for trees can also be adapted for populations by using algorithms developed for network under the denomination "Neighbor-Net" procedures [1], where PD could be optimized via computing a circular split system. Optimal PD could be obtained by morphological and molecular data. Using PD, Faith [40] proposed a taxa selection once having a phylogenetic tree of n taxa by identifying the set of k taxa that maximizes the PD, where k < n. The optimal set is tested yet to determine taxa that are of interest for sequencing projects in wildlife [41]. Although mathematical formulations exceeded our aims, following [13] we summarize the example in the paper of these authors to show how it works.

In **Figure 6**, we show a network graph ordered in a circular format A to E taxon. Each split could be weighted according to edge distances from each bisecting taxon (arrows in **Figure 6**) to the rest of taxa. As an example, the procedure to get an optimal PD distance (PD is equivalent to SD in Ref. [13]) form circular taxon order of A–E will be constructed for an optimal three-set of split taxon as follows:



(iii) Index matrix to trace back the optimum. The 3-Path taxon maximizing DP (in blue an example: the ABC maximum 3-path DP; see **Figure 6**).


#### (iv) Compute the longest ordered two-path using L2 = duv (as in second line at formulae).


(v) Derived L<sup>3</sup> from L2 (as in second line at formulae) but only three taxa.

Example A to C = (3+2+4+2) + (2+6+4) = 23 (B features two times¡)

Example B to E = (2+4+6+4+5) + (5+2+4) = 32

**5. Prioritizing populations for conservation using phylogenetic** 

According to the "species richness" concept [38], practical biodiversity conservation has the aim to preserve as many species as possible. However, as previously said, such an approach has the hurdle of treating all species equally [39]. However, neither all species nor genetic lineages are equally important, with more isolated lineages providing more important contribution to total variation, that is, the base for identifying populations worthy of protection in law. Genetic variation is depicted perfectly onto a rooted phylogenetic tree, where the edge length represents the number of features uniquely shared by all descending taxa, say populations. Importantly, ESUs concept assumes that the relationships among populations can be represented by a bifurcating tree. However, these sort of phylogenetic trees often fail to capture complete genetic information among populations. Moreover, more complex interrelationships are expected for DPSs and MUs. So, it would seem a shortcoming could occur if populations do need to be prioritized for conservation on the basis of tree-based prioritization schemes. However, the prioritization approaches for trees can also be adapted for populations by using algorithms developed for network under the denomination "Neighbor-Net" procedures [1], where PD could be optimized via computing a circular split system. Optimal PD could be obtained by morphological and molecular data. Using PD, Faith [40] proposed a taxa selection once having a phylogenetic tree of n taxa by identifying the set of k taxa that maximizes the PD, where k < n. The optimal set is tested yet to determine taxa that are of interest for sequencing projects in wildlife [41]. Although mathematical formulations exceeded our aims, following [13] we summarize the example in the paper of these authors to show

In **Figure 6**, we show a network graph ordered in a circular format A to E taxon. Each split could be weighted according to edge distances from each bisecting taxon (arrows in **Figure 6**) to the rest of taxa. As an example, the procedure to get an optimal PD distance (PD is equivalent to SD in Ref. [13]) form circular taxon order of A–E will be constructed for an optimal

**distances from networks: split diversity (SD)**

**Figure 5.** Cross-hair analysis for management recommendations (adapted from [8]).

how it works.

40 Phylogenetics

three-set of split taxon as follows:

(i) Formulae to be used (n°4 in [13]).

(ii) Compute the pairwise distance matrix duv (distance count).



(vi) Calculate L<sup>3</sup> + L2 for the longest ordered three-path.


Several phylogenetic diversity measures have been adapted for nontree-like population genetic data. However, these methods could be conditioned to change when natural or artificial (human mediated) extinction alters the network structure. Given both the stochastic and/or selective nature of extinction, different metrics, like split diversity (SD; similar to PD) from [13] or Shapley metric (SH [14]), and heightened evolutionary distinctiveness (HED [15]) offer general ranking systems useful to wildlife managers rather than those based only on the present structure of a phylogenetic network trees. However, SH and HED rankings have been stated as able to allow lengthening or shortening the list of taxa to conserve in the event that resources become more or less available, which may give potential relevant frameworks or schemes for preserving future biodiversity [1].

**Figure 6.** Split graph and its split systems (adapted from [13]).

Nowadays, the most recent, more inexpensive, and robust advances in molecular techniques make of the genetic sampling of populations a standard component of conservation planning. Moreover, there are views that value phylogenetic network approach because it offers insight into a species' population structure complementary to the current statistical assessments of differentiation employed by MUs and DPSs [11, 12]. Genotyping at multiple informative loci and networks will provide population genetic studies aimed at giving advice to conservation agencies, to do more informative and accurate estimates of population differentiation and of conservation-relevant processes, mainly those important onto genetic isolation and their effects on diversity [42].

## **6. Conclusion**

(vi) Calculate L<sup>3</sup>

42 Phylogenetics

L3 + L2

is an element SD<sup>3</sup>

D E

schemes for preserving future biodiversity [1].

**Figure 6.** Split graph and its split systems (adapted from [13]).

+ L2

for the longest ordered three-path.

(vii) Determine maximal scores SD max (three-circular tour) as (L<sup>3</sup>

(viii)Determine the longest ordered three-path from A to D using *αuv*

with the highest scores for PD<sup>3</sup>

Several phylogenetic diversity measures have been adapted for nontree-like population genetic data. However, these methods could be conditioned to change when natural or artificial (human mediated) extinction alters the network structure. Given both the stochastic and/or selective nature of extinction, different metrics, like split diversity (SD; similar to PD) from [13] or Shapley metric (SH [14]), and heightened evolutionary distinctiveness (HED [15]) offer general ranking systems useful to wildlife managers rather than those based only on the present structure of a phylogenetic network trees. However, SH and HED rankings have been stated as able to allow lengthening or shortening the list of taxa to conserve in the event that resources become more or less available, which may give potential relevant frameworks or

+ L2

= 28.

**A B C D E**

A 42 56 54 B 50 54

= C 46

)max =56/2.

<sup>3</sup> . As a result, the set ACD

Conservation genetics for wildlife is a recent challenge for humanity because biodiversity at several biotic levels need to be preserved to maintain desirable genetic variation for future generations. As a result, understanding biological diversity patterns and processes has increased the interest for phylogenetic analysis, remaining relevant all species. Nowadays, the imminent biodiversity crisis predicts significant new scenarios of biodiversity at the beginning of the twenty-second century for whichever wild species, which motivates to the geneticists in deal with preserve "all the gene pool". However, two faced situations are clearly involved in the context of conservation decisions. On the one hand, the identification of small populations harbors any significant genetic relevance worthy of conservation. On the other hand, identification of natural hybridized populations or species, although do not lack detractors when artificially promoted, due to it is believed to be an important process causing divergence in speciation and enhances the ability for survive. So, practical biodiversity conservation has the aim to preserve as many species (populations) as possible, but the relative importance of species or its genetic lineages should be carefully studied for to be prioritized. Phylogenetic diversity measures have been adapted to offer potential relevant frameworks or schemes for preserving future biodiversity based on accurate estimates of population differentiation and conservation processes.

## **Author details**

José Luis Fernández-García

Address all correspondence to: pepelufe@unex.es

Genetic and Animal Breeding, Animal Production Department. Faculty of Veterinary. Universidad de Extremadura, Cáceres, Spain

## **References**

[1] Volkmann L, Martyn I, Moulton V, Spillner A, Mooers AO. Prioritizing populations for conservation using phylogenetic networks. PLoS One. 2014;**9**(2):e88945. DOI: 10.1371/ journal.pone.0088945


[17] Igea J, Juste J, Castresana J. Novel intron markers to study the phylogeny of closely related mammalian species. BMC Evolutionary Biology. 2010;**10**:369. DOI: 10.1186/ 1471-2148-10-369

[2] Pemberton J. Introduction to conservation genetics. In: Frankham R, Ballou JD, Briscoe DA. Genetical Research. Cambridge University Press. 2004;**83**(3):221-222. DOI: 10.1017/

[3] Hedrick PW. Conservation genetics: Where are we now?. Trends in Ecology and Evolution.

[4] Fernández-García JL, Carranza J, Martínez JG, Randi E. Mitochondrial D-loop phylogeny signals two native Iberian red deer (*Cervus elaphus*) Lineages genetically different to Western and Eastern European red deer and infers human-mediated translocations. Biodiversity and Conservation. 2014;**23**(3): 537-554. DOI: 10.1007/s10531-013-0585-2

[5] Zachos FE, Frantz AC, Kuehn R, Bertouille S, Colyn M, et al. Genetic structure and effective population sizes in European red deer (*Cervus elaphus*) at a Continental scale: Insights from microsatellite DNA. Journal of Heredity. 2016;**107**(4):318-326. DOI: 10.1093/jhered/esw011

[6] Zachos FE. Gene trees and species trees—Mutual influences and interdependences of population genetics and systematics. Journal of Zoological Systematics and Evolutionary

[7] Soulé ME. What is conservation biology? BioScience. 1985;**35**:727-734. DOI: 10.2307/

[8] Crandall KA, Bininda-Emonds OR, Mace GM, Wayne RK. Considering evolutionary processes in conservation biology. Trends in Ecology and Evolution. 2000;**15**(7):290-295.

[9] Redding DW, Mooers AØ. Can systematists help decide the relative worth of bits of

[10] Ryder OA. Species conservation and systematics: The dilemma of subspecies. Trends in

[11] Moritz C. Defining 'evolutionarily significant units' for conservation. Trends in Ecology

[12] Waples RS. Pacific salmon, *Oncorhynchus* spp., and the definition of 'species' under the

[13] Minh BQ, Klaere S, von Haeseler A. Taxon selection under split diversity. Systematic

[14] Shapley LS. A value for n-person games. In: Kuhn HW, Tucker AW, editors. Contributions to the Theory of Games. Vol. II. Princeton: Princeton University Press; 1953. pp. 307-317

[15] Steel M, Mimoto A, Mooers AØ. Hedging our bets: The expected contribution of species to future phylogenetic diversity. Evolution Bioinformatic Online. 2007;**3**:237-244

[16] Bottrill MC, Joseph LN, Carwardine J, Bode M, Cook C, et al. Is conservation triage just smart decision making? Trends in Ecology and Evolution. 2008;**23**:649-654. DOI:

Ecology and Evolution. 1986;**1**:9-10. DOI: 10.1016/0169-5347(86)90059-5

and Evolution. 1994;**9**:373-375. DOI: 10.1111/j.1365-294X.1994.tb00080.x

endangered species act. Marine Fisheries Review. 1991;**53**:11-22

Biology. 2009;**57**:586-594. DOI: 10.1093/sysbio/syp058

Research. 2009;**47**:209-218. DOI: 10.1111/j.1439-0469.2009.00541.x

2001;**16**:629-636. DOI: 10.1016/S0169-5347(01)02282-0

S0016672304216913

44 Phylogenetics

1310054

DOI: 10.1016/S0169-5347(00)01876-0

biodiversity? Systematist. 2010;**32**:4-8

10.1016/j.tree.2008.07.007

