1. From the molecules of life, to the simpler organisms
Part I
Paula B. Matheus Carnevali

2. Outline The first organisms Classification systems
The major divisions of life
Phylogeny of bacteria
Prokaryotic cell structure and function
Gram negative vs. Gram positive

3. Cartoon of the tree of life
Rocks dating from the time of Earth’s formation do no exist on the planet’s surface, but radiometric dating of meteorites yields an estimated age for the solar system, and hence Earth, of 4.5 to 4.6 billion years. The newborn Earth remained inhospitable for at least a few hundred million years. By the best estimates, life arouse on Earth a bit less than 4 billion years ago. Although origins of life must be reconstructed using indirect evidence alone, it is thought that there must have been a primordial life form, or first living thing, represented by the red dot at the bottom of the figure. Among the descendants of the primordial form was the last common ancestor of all extant organisms (orange dot), known as the cenancestor or LUCA, for last universal common ancestor. The evolutionary history of LUCA’S descendants (blue branches) constitute the tree of life.
Figure 1. Last Universal Common Ancestor

4. The first cellular life whose descendants ultimately survived, appeared at least 2 billion years ago and probably much earlier
The origin of life has been under investigation, via observation and experimentation, for over 80 years. Biologists have made artificial cells and artificial cell membranes, and have zeroed in on chemical reactions that could have built cellular materials from nonliving sources. Early, in the course of these studies, a very important question arouse: which of its two most vital substances did life acquire first, proteins or DNA?
Figure 2. Fossilized bacteria. a) Archean Apex, 3.5 billion yrs old, b) Gloeodiniopsis, 1.5 billion yrs old, c) Palaeolyngbya, 950 million yrs old.
From Prescott et al., 2005

5. Which of its two most vital substances did life acquire first, proteins or DNA? Ribozymes
This chicken-and-egg problem was essentially resolved with the discovery of catalytic RNA or ribozymes. Because RNA has both a capacity for information storage and transmission and the ability to perform biological work, researchers now think that it preceded both proteins and DNA in the origin of life. But, how could RNA sequences of any kind arise in an abiotic environment? In other words, the primordial form was not made of RNA, but of something else that preceded RNA.
Figure 3. The mechanism of action of the ribozymes.
From Essential cell biology, 2/e, 2004, Garland Science

6. Characteristics of the first organisms
Self-replicating systems
Use of DNA to store heritable information
Use of proteins to express information
Cellular forms
Cellular membranes
Once self-replicating systems evolved on Earth, at least one of them adapted to the use of DNA to store heritable information and to the use of proteins to express information. This system eventually gave rise to all lineages of life on the planet today. We draw this conclusion because all the life forms (except some viruses) use DNA and proteins. Because another shared feature of all extant life is the existence of cells, we also infer that the common ancestor was a cellular form. Technically speaking, we need to say that all life has descended from a population of interbreeding cells, because if portions of the primitive genome could be readily swapped, the life today cannot trace its ancestry to a single organism. The advantages of cellular membranes, as well as internal organellar membranes would have been enormous.

7. Classification of organisms
Taxonomy is the science of biological classification.
Classification is the arrangement of organisms into groups or taxa (s. taxon) based on mutual similarity or evolutionary relatedness.
Nomenclature is the branch of taxonomy concerned with the assignment of names to taxonomic groups in agreement with published rules.
Identification is the process of determining that a particular isolate belongs to a recognized taxon.
Systematics is the scientific study of organisms with the ultimate object of characterizing and arranging them in an orderly manner.
Another way to study the ancestral lineage is to reconstruct the phylogeny of all living things. A universal phylogeny should allow us to infer additional characteristics of the earliest life forms, beyond just their cellular nature.

8. Some remarkable discoveries in microbiology
Aristotle first classified living things in
Plants and Animals
Bacteria were first observed by Anton van Leeuwenhoek in 1676 using a single-lens microscope of his own design.
The invention of the Electron Microscope allowed for the distinction between Prokaryotic and Eukaryotic cells
In the 1960s Robert Whittaker presented the five kingdoms system of classification including Fungi
Based on 16S rDNA Carl Woese and his collaborators suggested that prokaryotes were divided into two distinct groups very early on, and presented the Three domain system including Archaea
The first attempt to reconstruct the phylogeny of everything were based on the morphologies of organisms. Linnaeus developed the first natural classification, based largely on anatomical characteristics, in the middle of the eighteenth century. Following the publication in 1859 of Darwin’s On the Origin of the Species, biologist began trying to develop phylogenetic classification systems. These are systems based on evolutionary relationships rather than general resemblance.

9. Prokaryotes vs. Eukaryotes
Table 1. Comparison of Prokaryotic and Eukaryotic cells
From Prescott et al., 2005

10. Prokaryotes vs. Eukaryotes
Figure 4. Comparison of Prokaryotic and Eukaryotic cell structure. (a) The prokaryote Bacillus megaterium, (b) The eukaryotic alga Chlamydomonas reinhardtii, a deflagellated cell. From Prescott et al., 2005

11. Whittaker’s 5-Kingdom system
It lacks distinction between Archaea and bacteria. The kingdom Protista also may be too diverse to be taxonomically useful. The boundaries between kingdoms are ill-defined
Organisms are placed into five kingdoms based on at least three major criteria: (1) cell type-prokaryotic or eukaryotic, (2) level of organization-solitary and colonial unicellular organization or multicellular, and (3) nutritional type. The kingdom Monera contains all prokaryotic organisms. The kingdom Protista is the least homogeneous and hardest to define. Protists are eukaryotes with unicellular organization, either in the form of solitary cells or colonies of cells lacking true tissues. They may have ingestive, absorptive or photoautotrophic nutrition, and they include most of the microorganisms known as algae, protozoa, and many simple fungi. The kingdom Fungi contains eukaryotic and predominately multinucleate organisms.
Figure 5. The five kingdom system proposed by Whittaker. From Prescott et al., 2005

12. Major characteristics used in taxonomy
Classical Characteristics:
Morphological characteristics
Physiological and metabolic characteristics
Ecological characteristics
Genetic analysis
Molecular characteristics:
Comparison of proteins
Nucleic acid base composition
Nucleic acid hybridization
Nucleic acid sequencing

13. Phylogeny of bacteria
In preparing a classification scheme, one places the microorganism within a small, homogeneous group that is itself a member of larger groups in a non-overlarpping hierarchichal arrangement. A category in any rank unites groups in the level below it based on shared properties. Microbial groups at each level or rank have names with endings or suffixes characteristic of that level. Microbiologist often use informal names in place of formal hierchachical ones. The basic taxonomic group in microbial taxonomy is the species. Taxonomists working with higher organisms define the term species differently than do microbiologists. Species of higher organisms are groups of interbreeding or potentially interbreeding natural populations that are reproductively isolated from other groups. This is a satisfactory definition for organisms capable of sexual reproduction but fails with the microorganisms because they do not reproduce sexually. Prokaryotic species are characterized by phenotypic and genotypic differences.
Figure 6 . Hierarchical arrangement in taxonomy. From Prescott et al. , 2005
A prokaryotic species is a collection of strains that share many stable properties and differ significantly from other groups or strains.
A species (genomospecies) is a collection of strains that have a similar G+C composition and 70% or greater similarity as judged by DNA hybridization experiments.

14. Woese’s three domain system
The tree is divided into three major branches representing the three primary groups. The archaea and bacteria first diverged, the the eukaryotes developed. These three primary groups are called domains and placed above the phylum and kingdoms levels (the traditional kingdoms are distributed among these three domains). The domains differ markedly from one another. Eukaryotic organisms with primarily glycerol fatty acyl diester membrane lipids and eukaryotic rRNA belong to the Eukarya. The domain Bacteria contains prokaryotic cells with bacterial rRNA and membrane lipids that are primarily diacyl glycerol diesters. Prokaryotes having isoprenoid glycerol diether or diglycerol tetraether lipids in their membranes and archaeal rRNA compose the third domain, Archaea.
Figure 7. Tree of life by the three domain system

15. Bacteria, Archaea and Eucarya
Table 2. Comparison of Bacteria, Archaea, and Eucarya
From Prescott et al., 2005
Figure 8a. Prokaryotic cell
Figure 8b.
Eukaryotic cell

16. Table 2 cont. Comparison of Bacteria, Archaea, and Eucarya

17. Eukaryotic cells arouse form prokaryotic cells
It appears likely that modern eukaryotic cells arose from prokaryotes about 1.4 billion years ago (some chemical evidence dates them as early as 2.7 billion years ago). It is not certain how this process occurred, and two hypothesis have been proposed. According to the first, nuclei, mitochondria, and chloroplasts arose by invagination of the plasma membrane to form double-membrane structures containing genetic material capable of further development and specialization. The similarities between chloroplasts, mitochondria, and modern bacteria are due to conservation of primitive prokaryotic features by the slowly changing organelles.
Figure 9. A schematic representation of the process of endosymbiosis

18. Endosymbiotic hypothesis
According to the more popular endosymbiotic hypothesis, the first event was nucleus formation in the pro-eukaryotic cell. The ancestral eukaryotic cell may have developed from a fusion of ancient bacteria and archaea. Possibly Gram-negative bacterial host cell that had lost its cell wall engulfed an archaeon to form an endosymbiotic association. The archaeon subsequently lost its wall and plasma membrane, while the host bacterium developed membrane infolds. Eventually the host genome was transferred to the original archaeon, and a nucleus and the endoplasmic reticulum was formed. Both bacterial and archaeal genes could be lost during the formation of the eukaryotic genome. Mitochondria and chloroplasts appear to have developed later. The free-living, fermenting ancestral eukaryote with its nucleus established a permanent symbiotic relationship with the photosynthetic bacteria, which the evolved chloroplasts. Cyanobacteria have been considered the most likely ancestors of chloroplasts. Mitochondria arose from and endosymbiotic relationship between the free-living primitive eukaryote and bacteria with aerobic respiration.
Figure 10. A schematic representation of the process of endosymbiosis

19. Phylogeny of bacteria
In 1923, David Bergey and four colleagues published a classification of bacteria that could be used for identification of bacterial species, the Bergey’s Manual of Determinative Bacteriology. This manual is now in its ninth edition. The characteristics used to define sections are normally features such as general shape and morphology, Gram-staining properties, oxygen relationship, motility, the presence of endospores, the mode of energy production, and so forth.
Figure 11. Phylogeny of Bacteria. The tree is based on 16S rRNA comparisons. From Prescott et al., 2005.

20. Purpose of the Gram Stain
To separate bacteria based upon their cell wall structure and to determine their morphology and possible cellular arrangement
Gram-staining properties play a singularly important role in this classification. It usually reflects fundamental differences in bacterial wall structure. Gram-staining properties also are correlated with many other properties of bacteria.
Gram-positive cocci in chains
Gram-negative rod
Figure 12. The Gram-Positive and Gram-Negative envelopes.From Prescott et al., 2005

21. Gram negative vs Gram positive
Table 3. Comparison between Gram-positive and Gram-negative bacteria
Modified from Prescott et al., 2005

22. Prokaryotic cell organization
Figure 13. A prokaryotic cell

23. Size, shape and arrangement
Although it is true that many prokaryotes are similar in morphology, there is a remarkable variation due to differences in genetics and ecology. Most commonly encountered bacteria have one of two shapes. Cocci (s. coccus) are roughly spherical cells. They can exist as individual cells, but also are associated in characteristics arrangements that are frequently useful in bacterial identification. Diploccoci arise when cocci divide and remain together to form pairs. Long chains of cocci result when cells adhere after repeated divisions in one plane. Others divide in random planes to generate irregular grapelike clumps. Divisions in two or three planes can produce symmetrical clusters of cocci. The can also divide in three planes producing cubical patterns of eight cells. The other common bacterial shape is that of a rod, often called bacillus (s. bacilli). Bacilli differ considerably in their length-to-width ratio. Although many rods do occur singly, they may remain together after division to form pairs or chains. A few of them, are curved to form distinctive commas or incomplete spirals.
Figure 14. Examples of bacterial shapes. From Prescott et al., 2005

24. Size, shape and arrangement
Bacteria vary in size as much as in shape. The smallest are about 0.3 μm in diameter, approximately the size of the largest viruses. However nonobacteria or ultramicrobacteria appear to range from around 0.2 μm to less than 0.05 μm in diameter, but some researchers think that they are artifacts. A few bacteria become fairly large reaching 7 μm in diameter.
Figure 15. Comparison between the size of a human cell, bacteria and viruses. From Prescott et al., 2005

25. Table 4. Prokaryotic structures
From Prescott et al., 2005

26. Plasma membrane
This diagram of the fluid mosaic model of bacterial membrane structure shows the integral proteins (blue) floating in a lipid bilayer. Peripheral proteins (purple) are associated loosely with the inner membrane surface. Small sphere represent the hydrophilic ends of membrane phospholipids and wiggly tails, the hydrophobic fatty acid chains. Other membrane lipids such as hopanoids (pinks) may be present. For the sake of the clarity, phospholipids are shown in proportionately much larger size than in real membranes.
Figure 16 . Plasma membrane structure. From Prescott et al., 2005

27. Cell wall
The cell wall is the layer, usually fairly rigid, that lies just outside the plasma membrane. Except for the mycplasmas and some Archaea, most bacteria have strong walls that give them shape and protect them from osmotic lysis; wall shape and strength is primarily due to peptidoglycan. The cell wall of many pathogens have components that contribute to their pathogenicity. The wall can protect a cell from toxic substances and is the site of action of several antibiotics.
Figure 17. The cell wall. The Gram+ envelope is from Bacillus licheniformis, and the Gram- micrograph is of Aquaspirillum serpens. M=peptidoglycan or murein layer, OM=outer membrane, PM=plasma membrane, P=periplasmic space, W=Gram+ peptidoglycan wall. From Prescott et al., 2005

28. Gram positive cell walls
After Christian Gram developed the Gram stain in 1884, it soon became evident that bacteria could be divided into two major groups based on their response to the Gram-stain procedure. Gram-positive bacteria stained purple, whereas Gram-negative bacteria were colored pink or red by the technique. The true structural difference between these two groups became clear with the advent of the transmission electron microscope. The gram+ cell wall consist of 20 to 80 nm thick homogeneous peptidoglycan or murein layer outside the plasma membrane. Usually it also contains large amounts of teichoic acids, polymers of glycerol or ribitol joined by phosphate groups.
Figure 18. The Gram-Positive envelope. From Prescott et al., 2005

29. Gram negative cell walls
The Gram- cell wall is quite complex. It has 2 to 7 nm peptidoglycan layer surrounded by a 7 to 8 nm thick outer membrane. Frequently a space is seen between the plasma membrane and the outer membrane in electron micrographs of Gram- bacteria, and sometimes a similar but smaller gap may be observed between the plasma membrane and wall in Gram+ bacteria. This space is called the periplasmic space. The substance that occupies the periplasmic space is the periplasm, where many proteins that participate in nutrient acquisition, and binding proteins involved in transport of materials into the cell are found. The most abundant membrane protein is the Braun’s protein, a small lipoprotein covalently joined to the underlying peptidoglycan. Possibly the most unusual constituents of the OM are its lipopolysaccharides (LPSs) which consist of the lipid A, the core polysaccharide and the O side chain.
Figure 19. The Gram-Negative envelope. From Prescott et al., 2005

30. Bacterial nucleoid Figure 20. Some bacterial nucleoids.
Probably the most striking difference between Eukaryotes and Prokaryotes is the way in which their genetic material is packaged. Eukaryotic cells have two or more chromosomes contained within a membrane-delimited organelle, the nucleus. In contrast, prokaryotes lack a membrane-delimited nucleus. The prokaryotic chromosome is located in an irregularly shaped region called the nucleoid. It usually contains a single circle of double-stranded DNA, but some have linear DNA. In actively growing bacteria, the nucleoid has projections that extend into the cytoplasmic matrix. Presumably DNA that is being transcribed into mRNA. Careful electron microscopic studies often have shown the nucleoid in contact with either the mesosome of the plasma membrane. Chemical analysis reveals that they are composed of about 60 % DNA, 30 % RNA and 10 % protein by weight. Many bacteria posses plasmids in addition to their chromosomes.
Figure 20. Some bacterial nucleoids.
From Prescott et al., 2005
E. coli chromosome growing

31. Flagella and motility
Most motile bacteria move by use of flagella (s. flagellum), locomotor appendages extending outward from the plasma membrane and cell wall. The longest and most obvious portion is the filament, which extends from the cell surface to the tip. A basal body is embedded in the cell, and a short, curved segment, the hook, links the filament to its basal body and acts as a flexible coupling. The filament is a hollow, rigid cylinder constructed of a single protein called flagellin. The filament is in the shape of a rigid helix, and the bacterium moves when this helix rotates. Considerable evidence shows that flagella act just like propellers on a boat.
Figure 21. The ultrastructure of bacterial flagella. Flagellar basal bodies and hooks in (a) gram-negative and (b) gram-positive bacteria. From Prescott et al., 2005

32. Flagella and motility
Bacterial species often differ in their patterns of flagella distribution (monotrichous, polar flagellum, amphitrichous, lophotrichous, peritrichous).
Figure 22. The long flagella and the numerous shorter fimbriae on Proteous vulgaris. (a) Monotrichous polar, (b) Lophotricous, (c) Peritrichous.
From Prescott et al., 2005