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Ivan Kushkevych
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BACTERIAL
PHYSIOLOGY AND BIOCHEMISTRY
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Progress in Biochemistry and Biotechnology
BACTERIAL
Ivan Kushkevych
Department of Experimental Biology,
Faculty of Science, Masaryk University,
Brno, Czech Republic
Edited by
Josef Jampílek
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CONTENTS
ACKNOWLEDGEMENTS...................................................................................................... ix
ABBREVIATIONS.................................................................................................................. xi
FOREWORD.......................................................................................................................... xiii
CHAPTER 1. INTRODUCTION INTO BACTERIAL PHYSIOLOGY

AND BIOCHEMISTRY .................................................................................... 1
1.1. Subject of study ........................................................................................ 1
1.2. Bacteria in the phylogeny of living organisms
and diversity of cell shapes ...................................................................... 3
1.3. Bacterial evolution ...................................................................................7
1.4. Methods of studies of bacterial properties ............................................. 11
CHAPTER 2. BACTERIAL CHEMICAL COMPOSITION AND FUNCTIONAL

CELL STRUCTURES ..................................................................................... 23
2.1. Elemental composition ...........................................................................23
2.2. Compounds composition .......................................................................24
2.3. Bacterial nucleus .................................................................................... 33
2.4. Cytoplasm ...............................................................................................36
2.5. Plasma membrane ..................................................................................44
2.6. Cell wall .................................................................................................50
2.7. Flagella, pilus, and fimbria ..................................................................... 64
2.8. Bacterial capsule ....................................................................................72
2.9. Endospores .............................................................................................77
2.10. Pigments ................................................................................................. 85
CHAPTER 3. BACTERIAL NUTRITION AND GROWTH................................................. 91

3.1. Basic sources of nutrition........................................................................91
3.2. Sources of carbon.................................................................................... 92
3.3. Sources of nitrogen.................................................................................. 94
3.4. Mineral nutrition.....................................................................................95
3.5. Growth factors......................................................................................... 98
v
contents
3.6. Sources of energy.................................................................................. 108
3.7. Transport of compounds across the plasma membrane......................... 110
3.8. Passive transport.................................................................................... 111
3.9. Active transport.....................................................................................112
3.10. Transport of iron and its regulation.......................................................116
3.11. Transport of proteins ............................................................................124
3.12. Group translocation...............................................................................126
3.13. Bacterial growth and multiplication .....................................................127
3.13.1. Growth under conditions of static cultivation.......................... 130
3.13.2. Growth constants...................................................................... 133
3.13.3. Deviations from normal growth curve..................................... 136
3.14. Multiplication of microorganisms under conditions
of continuous (dynamic) cultivation......................................................138
3.15. Synchronous multiplication . ................................................................ 145
3.16. Bacterial cell cycle................................................................................ 146
CHAPTER 4. PROCESSES OF CELL DIFFERENTIATION.............................................. 151

4.1. Characteristics of differentiation processes........................................... 151
4.2. Polar differentiation in species of Caulobacter genus........................... 153
4.3. Differentiation of photosynthetic membranes
in facultative phototrophic bacteria....................................................... 156
4.4. Formation of heterocyst in cyanobacteria
under bound nitrogen deficiency........................................................... 157 t
CHAPTER 5. BACTERIAL METABOLISM....................................................................... 159

5.1. Energy of biochemical reactions........................................................... 160
5.2. Carriers of hydrogen.............................................................................. 163
5.3. Role of ATP and its formation in bacterial cells.................................... 166
5.4. Types of phosphorylation ..................................................................... 173
5.5. Processes of catabolism.........................................................................176
5.5.1. Catabolism of carbon compounds and fermentation..................177
5.5.1.1. Ethanol fermentation...................................................179
5.5.1.2. Lactic acid fermentation.............................................183
5.5.1.3. Pentose sugars fermentation....................................... 185
5.5.1.4. Propionic acid fermentation pathway......................... 188
5.5.1.5. Butyric acid fermentation and solvent formation....... 190
5.5.1.6. Mixed acid fermentation pathway.............................. 195
5.5.1.7. Fermentation of sugars and polysaccharides..............198
vi
contents
5.5.2. Anaerobic respiration................................................................. 199
5.5.2.1. Nitrate reduction and denitrification...........................199
5.5.2.2. Sulfate reduction (desulfurization)............................. 202
5.5.2.3. Carbon (IV) oxide reduction to methane....................208
5.5.3. Aerobic respiration in chemolithotrophic bacteria..................... 210
5.5.3.1. Oxidation of ammonia................................................ 211
5.5.3.2. Oxidation of reduced sulfur compounds..................... 213
5.5.3.3. Oxidation of iron compounds.....................................216
5.5.3.4. Oxidation of hydrogen................................................217
5.5.3.5. Oxidation of methane.................................................. 218
5.5.4. Aerobic respiration in chemoorganotrophic bacteria................. 218
5.5.4.1. Incomplete oxidation of substrate...............................219
5.5.4.2. Complete oxidation of substrate.................................220
5.5.4.3. Oxidation of saccharides and polysaccharides...........225
5.5.4.4. Oxidation of lipids......................................................227
5.5.4.5. Oxidation of hydrocarbons.........................................229
5.5.5. Catabolism of nitrogenous compounds...................................... 233
5.5.5.1. Dissimilation of proteins and amino acids.................. 233
5.5.5.1.1. Anaerobic degradation
(amino acid fermentation).......................... 233
5.5.5.1.2. Aerobic (oxidative) catabolism
of amino acids............................................240
5.5.6. Catabolism of heterocyclic compounds..................................... 246
5.5.6.1. Fermentation of heterocyclic compounds................... 246
5.5.6.2. Oxidation of heterocyclic compounds........................249
5.6. Processes of anabolism (biosynthesis)..................................................251
5.6.1. Biosynthesis of saccharides....................................................... 251
5.6.2. Biosynthesis of lipids.................................................................259
5.6.3. Consumption of CO2 by heterotrophic bacteria.........................265
5.6.4. Fixation of molecular nitrogen...................................................268
5.6.5. Biosynthesis of amino acids.......................................................269
5.6.6. Biosynthesis of nucleotides........................................................278
5.6.7. Biosynthesis of nucleic acids..................................................... 282
5.6.8. Biosynthesis of proteins.............................................................285
5.7. Regulation of metabolism process........................................................291
5.7.1. Regulation of enzyme synthesis................................................. 292
5.7.2. Regulation of enzymatic activity...............................................297
5.7.3. Specifics of regulation mechanisms...........................................298
5.7.4. Regulation of energetic metabolism...........................................300
vii
contents
5.8. Metabolism of phototrophic bacteria.................................................... 301
5.8.1. Photolithotrophs......................................................................... 301
5.8.2. Photoorganotrophs..................................................................... 305
CHAPTER 6. GROWTH OF MICROORGANISMS IN NATURE ....................................311

6.1. Microorganisms as part of the ecosystem............................................. 311
6.2. Physiological role of microorganisms in ecosystems............................ 313
6.3. Intercellular and internal population interactions
and quorum-sensing regulation of gene expression..............................315
6.4. Luminescent bacteria and bioluminescence.......................................... 319
CHAPTER 7. EFFECT OF ENVIRONMENTAL FACTORS

ON BACTERIAL CELLS.............................................................................. 325
7.1. Effect of external factors on bacteria.....................................................325
7.2. Mechanisms of the effect of environmental factors..............................328
7.2.1. Physical factors..........................................................................328
7.2.2. Chemical factors........................................................................ 335
7.2.3. Chemotherapeutics..................................................................... 345
7.3. Antibiotics and their mechanisms of action..........................................348
7.3.1. Antibiotics inhibiting cell wall synthesis................................... 348
7.3.2. Antibiotics disrupting the plasma membrane.............................353
7.3.3. Antibiotics disturbing the metabolism of nucleic acids............. 354
7.3.4. Antibiotics inhibiting protein synthesis..................................... 355
7.3.5. Antibiotics affecting phosphorylation process...........................358
RECOMMENDED REFERENCES.......................................................................................361
INDEX ...................................................................................................................................363
viii
ACKNOWLEDGEMENTS
Scientific reviewers:
Prof. PharmDr. Josef Jampílek, Ph.D., Professor of Medicinal Chemistry, Department of Chemical Biology,
Faculty of Science, Palacky University Olomouc, Slechtitelu 27, 783 71 Olomouc, Czech Republic;
Department of Analytical Chemistry, Faculty of Natural Sciences, Comenius University in Bratislava,
Ilkovicova 6, 842 15 Bratislava, Slovakia
Prof. Lorenzo Drago, Ph.D., Professor of Clinical Microbiology, Department of Biomedical Sciences for
Health, University of Milan, 20133 Milan, Italy
Prof. Aidan Coffey, Ph.D., Professor of Microbiology, Department of Biological Sciences, Cork Institute
of Technology, Bishopstown, Cork, Ireland
This educational book contains the modern knowledge of bacterial physiology and biochemistry.
The book includes seven chapters that describe the subject of bacterial physiology, chemical composition,
functional cell structures, nutrition, growth, and the processes of cell differentiation. Special attention
is paid to the bacterial metabolism, including the processes of catabolism and anabolism. The energy
of biochemical reactions, carriers of hydrogen, the role of ATP in the bacterial cells, and types of
phosphorylation are also discussed. The physiological role of bacteria in ecosystems, intercellular and
internal population interactions, quorum-sensing regulation of gene expression, and luminescent bacteria
and bioluminescence are described. The effect of environmental factors on bacteria, including physical
and chemical factors, and their mechanisms are presented. The description of chemotherapeutics and
antibiotics and mechanisms of their action is also provided. This book can be helpful for bachelor and
master students who study General Microbiology, Medical Microbiology, Veterinary Microbiology, and
Molecular Biology, for microbiologists, biochemists, biologists, experts in the cell and molecular biology,
and for readers interested in the study of bacterial physiology and biochemistry.
Acknowledgments: The author sincerely thanks Igor Starunko from Ivan Franko National Univer-
sity of Lviv (Ukraine) for his help and the technical preparation of the illustrative material and layout of
the book. The author also expresses special thanks to the scientific editor, Prof. Josef Jampílek, as well as
the reviewers, Prof. Lorenzo Drago and Prof. Aidan Coffey, for their time and help, critical comments,
and recommendations during preparation of this book.
Ivan Kushkevych
ix
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chapter 1. introduction into bacterial physiology and biochemistry
ABBREVIATIONS
ATP Adenosine triphosphate

Abe Abequose
ADP Adenosine diphosphate
AMP Adenosine monophosphate
APS Adenosine-5′-phosphosulfate
BChl Bacteriochlorophylls
BLAST Basic local alignment search tool
CAP Catabolite activator protein
Chl Chlorophylls
CTP Cytidine triphosphate
Cyt Cytochrome
Da Daltons
DAP Diaminopimelic acid
dATP Deoxyadenosine triphosphate
dCTP Deoxycytidine triphosphate
dGTP Deoxyguanosine triphosphate
DHB 2,3-Dihydroxybenzoate
DNA Deoxyribonucleic acid
dTTP Deoxythymidine triphosphate
dXDP Deoxyribonucleotide diphosphate
EDP Entner–Doudoroff pathway
EDTA Ethylenediaminetetraacetic acid
EMBL European Molecular Biology Laboratory
EMP Embden–Meyerhof–Parnas pathway
FA Fatty acids
FACS Fluorescence activated cell sorting
FAD Flavin adenine dinucleotide
Fd Ferredoxin
Feb Ferrienterochelin-binding protein
FMN Flavin mononucleotide
Gal Galactose
GDP Guanosine diphosphate
Glu Glucose
GluN Glucosamine
GSB Green sulfur bacteria
GSH Reduced glutathione
GSSG Oxidized glutathione
xi
abbreviations
GTP Guanosine triphosphate
GTP Guanine triphosphate
Hase Hydrogenase
HDHD 3-Hydroxydecanoyl-3-hydroxydecanic acid
Hep Heptose
HM Hydroxy muramic acid
HMP Hexose-monophosphate pathway
HPLC High-performance liquid chromatography
KDO 2-Keto-3-deoxy-octonoic acid
KEGG Kyoto Encyclopedia of Genes and Genomes
LPS Lipopolysaccharide
LTPP Lipothiamine pyrophosphate
Man Mannose
MR-test Methyl red test
NAD Nicotinamide adenine dinucleotide
NAG N-Acetylglucosamine
NAM N-Acetylmuramic acid
NRPS Nonribosomal peptide synthetases
OAc O-acetyl
ORF Open reading frame
PAPS 3′-Phosphoadenosine-5′-phosphosulfate
PAS 4-Aminosalicylic acid
PCR Polymerase chain reaction
PEP Phosphoenolpyruvate
QS Quorum-sensing
Rha Rhamnose
RNA Ribonucleic acid
SDS Sodium dodecyl sulfate
TCA Tricarboxylic acid
TDP-rha Thymidine diphosphate rhamnose
THF Tetrahydrofolic acid
TPP Thiamine pyrophosphate
UMP Uridine monophosphate
UTP Uridine triphosphate
UV Ultraviolet
VP-test Voges–Proskauer test
XDP Ribonucleotide diphosphate
xii
FOREWORD
All things are hidden, obscure and debatable if the cause

of the phenomena is unknown, but everything is clear if its cause be known.
Louis Pasteur
B acterial Physiology and Biochemistry is an educational book designed for

use in advanced bachelor’s and master’s courses in biology, including micro-
biology, biochemistry, and molecular biology. This book contains curriculum taught to
biology students specializing in microbiology. The knowledge of this subject is required
within the above-mentioned specialization in partial and state exams. The content of the
text and the way of organization are based on the relevant microbiology curriculum, cor-
responding to the requirements of the new educational system at the faculties of science.
The necessary knowledge of biochemistry and organic chemistry and mastering the
basics of general microbiology are assumed. In a clear form defined by the relevant cur-
riculum, the text can also be used to prepare students of professional and teacher biology
for the partial exam in the physiological part of general microbiology. This book pro-
vides the most current, authoritative, and relevant presentation of bacterial physiology
and biochemistry and includes seven chapters about the subject of the book in general,
bacterial chemical composition and functional cell structure, nutrition and growth, pro-
cesses of cell differentiation, metabolism, and the influence of environmental factors.
The first chapter describes the subject of bacterial physiology and biochemistry,
bacteria in the phylogeny of living organisms and their shape diversity and evolution.
This chapter also presents methods for studying bacterial properties. Detailed informa-
tion on the chemical composition and functional cell structure of bacteria is described in
the second chapter. It includes the element and compound composition, and characteri-
zes the bacterial nucleus, cytoplasm, plasma membrane, cell wall, flagella, pilus and
fimbriae, bacterial capsule, endospores, and pigments.
An integral part of bacterial physiology is microbial nutrition and growth as well
as basic sources of nutrition, including sources of carbon and nitrogen. Therefore, the
third chapter of this book presents mineral nutrition, growth factors, energy sources,
and transport of substances across the plasma membrane. Furthermore, the growth and
multiplication of bacteria, their growth under static culture conditions, growth constants,
and deviations from the normal growth curve are described. This important chapter con-
cludes with the multiplication of bacterial populations under conditions of continuous
(dynamic) cultivation, synchronous multiplication, and the bacterial cell cycle.
The text of the fourth chapter presents the process of cell differentiation, characteris
tics of differentiation processes, polar differentiation, differentiation of photosynthetic
membranes in facultative phototrophic bacteria, and the formation of heterocysts in
cyanobacteria under bound nitrogen deficiency.
xiii
foreword
Special attention is paid to bacterial metabolism, energy of biochemical reactions,
hydrogen carriers, the role of ATP in cells, and types of phosphorylation. The chapter
describing metabolic processes is divided according to two metabolic pathways into
catabolism and anabolism sections. The processes of catabolism of carbon compounds
and fermentation of ethanol, lactic acid, pentose sugars, propionic acid, butyric acid
with solvent formation, mixed acids, and sugars and the fermentation pathway of poly-
saccharides are described in detail. Then anaerobic respiration is shown, including
nitrate reduction and denitrification, sulfate reduction, and reduction of carbon dioxide
to methane. The process of aerobic respiration in chemolithotrophic bacteria, including
oxidation of ammonia, reduced sulfur and iron compounds, hydrogen, and methane, is
represented in sections of this chapter. Aerobic respiration in chemoorganotrophic bac-
teria is also described, especially incomplete oxidation of alcohols and glucose as well
as complete oxidation of substrates. Nitrogen catabolism, protein, and amino acid dis-
similation are characterized. Anaerobic degradation and aerobic metabolism of amino
acids and heterocyclic compounds as well as fermentation and oxidation of heterocyclic
compounds complete the section about bacterial catabolism. Special attention is paid
to the processes of anabolism, including carbohydrate and lipid biosynthesis, carbon
dioxide consumption by heterotrophs, and molecular nitrogen fixation. The biosynthesis
of amino acids, nucleotides, nucleic acids, and proteins is shown. In addition, the regula-
tion of the metabolic process and the metabolism of phototrophic bacteria are summa-
rized in this chapter, which is one of the largest chapters of the book.
Physiological and biochemical processes depend on the environment. Therefore,
the next chapter characterizes microbial growth in nature, bacterial populations as part
of the ecosystem, and their physiological role in ecosystems. In addition, intercellu-
lar and internal population interactions, quorum-sensing regulation of gene expression,
luminescent bacteria, and bioluminescence are presented. The last chapter of this book
describes the influence of environmental factors on bacteria, including physical and
chemical factors, and chemotherapeutics. Antibiotics and their mechanisms of action are
also demonstrated. This book concludes with the description of antibiotics that inhibit
cell wall synthesis, disrupt the plasma membrane, and affect nucleic acid metabolism,
protein synthesis, and the phosphorylation process.
Prof. Lorenzo Drago, Ph.D.,

Professor of Clinical Microbiology,
Department of Biomedical Sciences for Health,
Faculty of Medicine and Surgery at University of Milan, Italy
xiv
CHAPTER 1
INTRODUCTION INTO BACTERIAL

I n this chapter, the studies of bacterial physiology and biochemistry and their
main goals are characterized. Bacteria in the phylogeny of living organisms and
diversity of cell shapes are described. Special attention is paid to the comparison of
cell structures and their metabolic properties in different organisms, including bacteria,
archaea, and eukaryotes. Moreover, main features of bacterial evolution and methods
of modern bacterial physiology and biochemistry are presented.
1.1. Subject of study

The subject of bacterial physiology is the study of the functions of bacterial cells,
that is, the study of all physical, chemical, and biological processes occurring in the cell
as well as physical, chemical, and biological transformations caused by bacteria in the
environment during their development. Such studies are impossible without familia
rizing with the organization of morphology and functional structures of the microbial
cell. For performing biological functions, bacterial cells are differentiated into architec-
tural and functional structures.
An important part of bacterial physiology is the study of the chemical composi-
tion of bacterial cells, which, albeit similar to the chemical composition of the cells of
higher organisms, has certain peculiarities. These features are related to the ability of
bacteria to adapt to the environment. Due to such adaptation, depending on the environ-
ment, different variants of the same microorganism by morphological and physiological
characteristics can be observed. The process of adaptation is explained not only by the
existence of various types of bacterial metabolism, but also by the transformation of
the saprophytic state into parasitic and the appearance of pathogenic microorganisms.
The following two types of living organisms are distinguished by the cell structure:
prokaryotic cells (bacterial and archaeal cells that do not have a formed
nucleus);
eukaryotic (cells with the nucleus).
Bacterial Physiology and Biochemistry 1
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Copyright © 2023 Elsevier Inc. All rights reserved.
bacterial physiology and biochemistry
Prokaryotic and eukaryotic cells differ in structural organization. Prokaryotes,

unlike eukaryotes, do not have membrane-bound compartments called organelles that
perform specialized functions. As mentioned above, prokaryotic cells do not have
a formed nucleus, and hereditary information (DNA) in prokaryotes is not separated
from other components of the cell. This feature is the most important difference
between prokaryotic and eukaryotic cells. A comparison of the cellular structure of
eukaryotic and prokaryotic cells is presented in Table 1.1.
Table 1.1. Comparison of bacteria, archaea, and eukaryotes
Properties Bacteria Archaea Eukaryotes
Nucleus with a nuclear membrane

Absent Absent Present
around the DNA
Complex of internal membrane

Absent Absent Present
organelles
Have peptidoglycan, Do not have Do not have

Cell wall
muramic acid muramic acid muramic acid
Esterified, Esterified, branched, Esterified,

Membrane lipids
not branched aliphatic chains not branched
Gas vesicles Present Present Absent
Contains thymine, Does not contain Contains thymine,

RNA transfer
N-formylmethionine thymine N-formylmethionine
Polycistronic mRNA Present Present Absent
Introns of the mRNA Absent Absent Present
Splicing of the mRNA Absent Absent Present
Ribosomes:
Size 70S 70S 80S
Sensitivity to chloramphenicol and Positive Negative Negative
kanamycin
DNA-dependent RNA polymerase

Number of enzymes One Few Three
Structure Simple subunit Complicated Complicated
(4 subunits) Subordinate structure Subordinate structure
(8–12 subunits) (12–14 subunits)
Sensitivity to rifampicin Positive Negative Negative
Polymerase II promoters Absent Present Present
Metabolism:
Similar ATPase Absent Present Present
Methanogenesis Absent Present Absent
Fixation of nitrogen Present Present Absent
Photosynthesis with participation
of chlorophyll Present Absent Present
Chemolithotrophs Present Present Absent
2
The comparison of similar properties of organisms, regardless of the presence of

the nucleus, is presented in Fig. 1.1.
Contain molecules that transmit Capable of grown and

Age highly organized
hereditary information reproduction
Cells of all organisms, regardless of the presence of the nucleus,

have the similar properties
Have a cellular metabolism, in which

Express hereditary information
energy is used and they convert
in transcription and translation processes
the compounds into cellular structures
Fig. 1.1. Comparison of similar properties of organisms.
Thus, prokaryotic and eukaryotic cells differ in structural organization, although

the cells of all organisms, regardless of the presence of the nucleus, have similar
properties. So, bacterial physiology studies the chemical composition of bacterial
cells, physical and biological processes occurring in the cells, and transformations of
different metabolic compounds.
1.2. Bacteria in the phylogeny of living organisms

and diversity of cell shapes
Microbiology is a branch of science that covers the study of viruses, archaea,
bacteria, fungi, algae, and protozoa. There are no significant differences in the intracel-
lular structure of bacteria and archaea. There are fundamental biochemical differences
between them, which reflects their evolutionary origin. About 10 years ago, most sci-
entists believed that evolution was in two ways: one of them led to the formation of
prokaryotic cells (bacteria), and the other to the emergence of eukaryotic cells. The
terms “bacteria” and “prokaryotes” were considered synonymous. These views have
undergone radical changes in the 80s of the last century, when Carl Richard Woese,
a molecular biologist, began analyzing information molecules that directly reflect
hereditary cell information.
The analysis of ribosomal RNA showed that there are three principal lines of evo-
lution that form three separate domains of cell evolution:
bacterial cells;
rchaeal cells;
eukaryotic cells (mushroom, algae, protozoa, plants, animals).
3
The conducted studies allowed determining:

•• organelles of eukaryotes;
•• cells involved in the formation of energy (mitochondria and chloroplasts) come
from prokaryotic cells that have lost their ability to live independently;
•• organisms evolved in three different ways from one common predecessor, which
led to the formation of a large variety of microorganisms, plants, and animals
that exist today.
Due to the received information, a new system of classification of living orga
nisms was created. It is based on the analysis of rRNA macromolecules.
It is assumed that there was a certain common ancestor “progenote,” which gave
rise to the three branches of the evolutionary tree (Fig. 1.2). How it was, it is unknown.
Bacteria Archaea Eukaryota

Green Slime molds
filamentous bacteria
Spirochetes Entamoebae Animals
Gram Fungi
Methanosarcina
positives Methanobacterium Halophiles
Plants
Proteobacteria Methanococcus
Thermococcus celer Ciliates
Cyanobacteria
Thermoproteus Flagellates
Planctomyces Pyrodictium
Trichomonads
Bacteroides
Cytophaga Microsporidia
Thermotoga Diplomonads
Aquifex
Fig. 1.2. Phylogenetic tree of life by Carl Woese et al. (1990).
Archaea are highly specialized prokaryotic organisms, and although they are simi-
lar to bacteria in the structural organization, they have a number of fundamental diffe
rences. The most important feature of the archaea is the specificity of their ribosomal
and transport RNA; their ribosomes differ in shape. Differences were also found in
other components of the protein synthesis system. Archaea do not have fatty acids and
polyhydric alcohols as part of membrane lipids and usually have from 20 to 40 carbon
atoms. The lipid layer of the membrane is formed by a monomolecular layer, which,
obviously, gives its strength. Externally, archaea often have surface layers formed in
a certain way by structured and regularly packed protein or glycoprotein molecules of
the correct and sometimes strange form. The structure of the cell wall of the archaea may
include peptides and polysaccharides. Some archaea are characterized by processes that
are not intrinsic to other organisms. For example, some representatives of this group
of prokaryotes form methane (methanogens) in their process of life. Most archaeas are
extremophiles, that is, they develop under extreme conditions, at high temperatures
(+90 °C) or in saturated saline solutions. Acidophilic archaea grow in the environment,
4
where the pH is as low as in the concentrated sulfuric acid. There are autotrophic forms
of archaea that do not require organic food but are satisfied with the energy obtained
through oxidation–reduction reactions, with the involvement of inorganic molecules.
Bacteria that differ in their morphological and physiological properties are an
extremely diverse group of prokaryotic microorganisms. They can be spherical, cylin-
drical, spiral, and pleomorphic (Fig. 1.3).
Spherical (cocci) Rods (bacillus)

Micrococcus
Plane of
division Single bacillus
Diplococci
Diplobacilli
Streptococci
Streptobacilli
Tetrad
Sarcina
Coccobacilli
Staphylococci
Spiral shapes
Vibrio Spirillum Spirochete
Fig. 1.3. Basic shapes of bacterial cells: spherical (coccus, cocci), rods (bacteria, bacillus), and spirals.
5
Size, shape, and arrangement of bacterial cells (Table 1.2):

•• the size, shape, and arrangement of bacterial cells depend on the species;
•• even in pure culture, individual cells of the population are not present in the
same phase of the cell cycle at the same time;
•• therefore, the size (and sometimes the shape) varies;
•• for bacterial cells, there is a considerable variation in size due to species affinity
and the physiological state of the cell and the external environment.
Table 1.2. The size of some bacterial cells
Organism Size (µm) Organism Size (µm) Organism Size (µm)
Chlamydium About 0.3 Streptococcus 1.0–1.5 Clostridium 0.5–0.8×3.0–8.0
Bdellovibrio 0.3–0.5×0.5–1.4 Sarcina 4.0–4.5 Shigella 0.4–0.6×1.0–3.0
Rickettsia 0.1–0.3×1–4 Azotobacter 2–3×3–6 Spirochaeta 0.2–7×5–500
Staphylococcus 0.8–1.0 Proteus 0.5–1×1–3 Spirillum 0.5×3
Escherichia coli 0.5–1×1–3 Micrococcus About 1.0 Corynebacterium 0.32–0.8×1.0–8
Bacillus subtilis 0.9–1.1×1.8–4.8 Pseudomonas 0.3–0.5×1.0–1.8 Treponema 0.2×4–14
Mycobacterium 0.3–0.5×1–4 Acetobacter 0.4–0.8×1.0–2.0 Corynebacterium 0.3–0.8×1.0–8.0
Chromatium 3–4×5–6 Lactobacillus 0.7–1.0×3.0–8.0 Brucella 0.3–0.5×0.5–2
Some bacteria are capable of photosynthesis and use the energy of sunlight and
CO2 to accumulate biomass. Among them, there are cyanobacteria, which were called
blue-green algae. Another group of bacteria obtains energy by metabolizing such
inorganic compounds as ammonium and sulfur. Bacteria can decompose different
organic compounds, from glucose to hydrocarbon oil. There are bacteria that can grow
only under specific conditions, for example, in human tissues, where they can cause
diseases. Purple nonsulfur bacteria can grow both as anaerobes (photoorganohetero-
trophs) and aerobes (chemoorganoheterotrophs). They are found in water with a high
content of organic matter and low sulfur content. Intracellular parasites (rickettsiae)
can use nutrient substances of the host, its ATP, and coenzymes. Representatives of
many genera of bacteria, in particular Rhizobium, are capable of fixing nitrogen, while
Agrobacterium causes tumor growth in plants.
Chemolithotrophic bacteria obtain energy by restoring inorganic compounds,
and nitrifying bacteria can oxidize ammonia and nitrites. Colorless sulfur bacte-
ria Thiobacillus can oxidize sulfur, hydrogen sulfide, and thiosulfates to sulfates.
Photolithotrophs (purple sulfur bacteria) can oxidize hydrogen sulfide to sulfur and
accumulate it in their cells. Methylotrophic bacteria use methane, methanol, and other
monocarbon compounds as the only source of carbon and energy. Representatives of
the Pseudomonas genus play an important role in the processes of mineralization (and
6
at the same time cause diseases). Various diseases are caused by enterobacteria, which
are widely used in experimental studies. The species of myxobacteria, which forms
fruiting bodies, moves along the surface in search of food, and excretes enzymes that
can cause lysis of bacteria and yeasts, has a complex life cycle.
Representatives of the Clostridium genus, which form endospores, cause food
poisoning (botulism), gangrene, and tetanus. Staphylococci are important pathogens.
At the same time, species of the Lactobacillus genus, capable of lactic fermentation,
are widely used in the food industry. The Рropionibacteria genus and streptomyces,
capable of degrading many organic compounds and producing antibiotics, are of
industrial significance.
Thus, despite the fact that prokaryotes and eukaryotes are very different in struc-
ture, the metabolisms of these two groups of organisms are similar. Photosynthetic
eukaryotes are algae (unicellular or multicellular), which can be organized into comp
lex structures. Based on the study of the structure and organization, blue-green algae
were considered as cyanobacteria, because they do not have a nucleus. Microbial
physiology covers the research of thousands of different microorganisms. On examp
les of the best-investigated bacteria, the basic principles of their life will be discussed.
1.3. Bacterial evolution

About 3.8 billion years ago, the Earth’s atmosphere contained hydrogen, methane,
ammonia, and water but did not contain CO2, oxygen, and organic compounds. In
the 50s of the last century, S. Miller and G. Urei have shown that under the influence
of electrical discharges, organic compounds, in particular, sugars, amino acids, and
nucleotides, can be formed in such an atmosphere.
In the early decades of the 20th century, Aleksandr Oparin (in 1924) and John
Haldane (in 1929, before Oparin’s first book was translated into English) independently
suggested that if the primitive atmosphere was reducing (as opposed to oxygen-rich),
and if there was an appropriate supply of energy, such as lightning or ultraviolet light,
then a wide range of organic compounds might be synthesized (Fig. 1.4). These com-
pounds could provide energy, growth, and reproduction necessary for life. An alterna-
tive is the hypothesis about the origin of micelles in oceanic thermal waters. Although
it is not known how the first organisms evolved, experiments show that in conditions
similar to the atmospheric conditions of the Earth at that time, micelles resembling
cells are formed in a mixture of organic compounds. The micelles are separated
from the environment, and within the micelles, there may be some reactions between
organic compounds, which could lead to the formation of cells. These first microor-
ganisms were supposed to be resistant to external factors: high temperature, acid pH,
and high salt concentration. They had to be anaerobic, because they had no oxygen;
they had to use organic compounds for energy, growth, and development. According
to these features, primitive organisms are reminiscent of microorganisms, which are
now classified as archaea. They are physiologically highly specialized and can grow
under adverse conditions (temperature, acidity). The gradual change in the conditions
7
on Earth led to the emergence of new organisms, called autotrophs (using CO2 as a car-
bon source, capable of synthesizing complex organic compounds from CO2). Primary
anaerobic microorganisms developed mechanisms for the capture and use of chemical
and solar energy for biosynthetic purposes. In the form in which these processes are
observed on Earth now, they received the name of chemosynthesis and photosynthesis.
It is believed that the first photosynthetic organisms on Earth were the ancestors of
cyanobacteria (blue-green algae). Photoautotrophs are capable of using light as an
energy source for the synthesis of organic compounds. The synthesized organic com-
pounds could be a carbon source for other microorganisms called heterotrophs. The
first photosynthetic bacteria were obligatory anaerobes. Later, there were new photo
synthetic microorganisms that could form oxygen from water. It is believed that it
was 2 billion years ago. The formation of oxygen in the process of photosynthesis had
a huge impact on evolutionary development. Due to aerobic breathing, the metabolism
of organic compounds became possible.
kJ kJ kJ T
11106 170 6 1 per 4 109 years
m2 h m2 h m2 h m2
Radiation Lightning bolts Volcanoes Meteorites
H 2 H 2O
CH4 CO2
Biosphere NH3
Primary 2.5 1018 g
atmosphere
kJ
Prokaryotes 120 2
4 ×10 9 years ago
m h
HCN
Ocean H2CO
Nucleotides 23 C2nH2n + 2
c ≤ 10 g
Polysaccharides Sedimentary CH3OH
Amino acids rocks C nH n
Nitrogenous bases
Carbohydrates
Carboxylic acids
Fig. 1.4. The scheme of life's origin based on Oparin–Haldane hypothesis.
Alberts and Watson et al. believe that the emergence of anaerobic prokaryotes in
the conditions of the depletion of nutrients from the environment and the ability to eat
one another (phagocytosis) was another important stage in the development of life
8
on Earth. According to these authors, while some organisms used phagocytosis, other
anaerobic prokaryotes could enter into symbiosis with their aerobic cells absorbed and
create a mutually beneficial association. Thus, phagocyted aerobic cells could not be
used as a power source but left inside the host cell for a more complete and effective
oxidation of organic matter.
This could give significant advantages to host cells in the struggle for existence.
During the evolutionary development of new microorganisms, which adapted to new
conditions, this process continued.
The idea of symbiosis, that is, the formation of a mutually beneficial associa-
tion of two or more organisms, arose in the second half of the 19th century. In 1867,
A. S. Famincin published that lichens are a mutually beneficial association of mush-
rooms and algae. The name “symbiosis” was proposed by De Bari. In 1907, based
on his research as well as on the work of A. F. Schimper (Germany), who showed the
ability of chloroplasts to self-replant in plant cells, Famincin suggested that chloro-
plasts could be single-celled algae and symbionts of plants. This idea was supported
and developed by K. S. Merezhkovsky and B. M. Kozopolyansky in the 20s of the
20th century. At the same time, it was suggested that not only chloroplasts but also
mitochondria are symbionts, since both are capable of self-replication in cells of higher
plants. However, this hypothesis was not mentioned as too extravagant for a long time.
Only in the 50s and 60s of the last century, biochemists received data on the content of
plants and mitochondria in chloroplasts. These organelles isolated from different orga
nisms. Mitochondrial DNA that differed from the DNA of nuclei was similar to the
DNA of prokaryotes (in particular, it had a circular structure). In addition, the presence
of prokaryotic ribosomes in chloroplasts of plants was established by N. M. Sisakyan
and other scientific groups. The process of protein biosynthesis in chloroplasts and
mitochondria on many grounds resembled the synthesis of protein in bacterial cells.
Chloroplasts and mitochondria appeared to be similar to bacterial cells by other
biochemical features, in particular, by the presence of a specifically constructed phos-
pholipid named cardiolipin in their membranes, which is characteristic only of bacterial
membranes and absent in mammalian eukaryotes (Fig. 1.5). However, there were many
arguments against this hypothesis. The main of them is the weak autonomy of life, first
of all, the process of biosynthesis of proteins, in chloroplasts and in mitochondria. It
turned out that in these organelles, a small amount of proteins is synthesized, first of all,
primarily necessary for the formation and functioning of these organelles.
The next step in the development of the hypothesis about the endosymbiotic origin
of eukaryotic organisms was the work of Lynn Margulis, an American researcher. It
discovered a very large immunological, that is, structural, proximity of the flagella
and some elements of the cytoskeleton of eukaryotic cells with spirochetes, which
are bacteria that have a spiral form of cells. This enabled L. Margulis to suggest
that the cytoskeleton and eukaryotic cell may have originated from spirochete-like
prokaryotes.
Recently, it was shown that ATPase (an enzyme that utilizes ATP), isolated from
yeast vacuoles, is structurally distinct from ATPases of other organelles of yeast and
9
practically corresponds to ATPase of some Archaea. On this basis, it was assumed that
the precursors of the vacuoles of lower eukaryotes, for example yeasts, are ancient
prokaryotes belonging to the archaea and entered into a symbiotic relationship with
the host cell. Recently, it has been shown that another enzyme, polyphosphatase
(utilizing inorganic polyphosphates), isolated from various organelles of yeasts (nuclei,
mitochondria, cellular membranes, cytosols, and vacuoles) has completely different
characteristics and properties. These data also indicate that many intracellular structures
of eukaryotic organisms have different origins.
Oxygen-breathing
The young earth bacterium
supported many
types of bacteria
Photosynthetic
bacterium
Most membrane-enclosed
organelles, including the nucleus,
ER, and Golgi, probably originated
from deep folds in the plasma
membrane
Mitochondria and chloroplasts

originated as bacterial cells that
came to live inside larger cells
Fig. 1.5. Hypothesis of сhloroplast and mitochondria origin from photosynthetic and oxygen-breathing
bacteria.
The most significant evidence of the origin of chloroplasts and mitochondria of

eukaryotic cells from prokaryotes was obtained in recent years in the study of the struc-
ture of one of the most conservative components of living cells — ribosomal RNA. The
prominent American molecular biologist Carl Richard Woese and his collaborators
have shown that the structure of the 16S RNA from ribosomes of plant chloroplasts
is not similar to the structure of the corresponding 18S RNA from the cytoplasmic
ribosomes of plants and is almost identical to the structure of similar RNA from some
10
cyanobacteria. At the same time, this group of researchers has found that the ribosomal
16S RNA derived from mitochondria of various eukaryotic organisms, by their struc-
ture, are not similar to the ribosomal 16S RNA from the cytosol of the same biological
objects but are extremely similar to the 16S RNA of some bacteria, in particular, of
the Paracoccus genus. All these data clearly indicate in favor of the high probability
of the fact that eukaryotic cells have endosymbiotic origin. However, the problem of
establishing the nature of the host cell remains unsolved. According to L. Margulis,
host cells could be prokaryotes-precursors of modern bacteria, which belong to myco-
plasmas (primitive constructed prokaryotes, which practically do not have a cell wall).
In recent years, based on the study of the structure of the ribosomal 16S RNA of
many prokaryotes and eukaryotes, T. Oshima, a Japanese researcher, concluded that
the host cell was a representative of the kingdom of Archaea. Incidentally, in contrast
to bacteria, representatives of Archaea have many biochemical and molecular genetic
properties common with those of eukaryotes.
1.4. Methods of studies of bacterial properties

Different types of microscopy and cultivation of microorganisms are traditional
methods that made it possible to investigate their structural and physiological charac-
teristics, in particular, metabolism and growth under different conditions, and to detect
the extreme heterogeneity of microorganisms. The use of new types of microscopy
(confocal, transmissive, scanning), new methods for obtaining pure cultures of micro-
organisms, immunological methods, and methods of classical genetics provided much
better disclosure of the basic properties of microorganisms and possibilities of the
practical use of microbial technologies.
Currently, microbial physiology is undergoing a dramatic revolution. Huge pro-
gress in this area is due to the following technological advances:
personal computers;
Internet;
development of DNA sequencing techniques.
In the recent past, the study of the physiology of microorganisms was a long, dif-
ficult, and ungrateful process. Even 60 years ago, when the full analysis of Escherichia
coli was launched, there was still no complete integrated picture of its biochemistry
and genetics. Today, we know in detail the biochemistry and genetics of some micro-
organisms, which are of special interest to humans. Examples are Rickettsia prowazekii
and Mycobacterium leprae, which cannot exist outside the living organism. Currently,
we know what special biochemical ways they have, how they obtain energy, which
protein factors determine their virulence, and what their place in the evolutionary tree
of bacteria is. All this became known thanks to modern sequencing techniques that
allowed looking into the genome and decoding it in a relatively short time. An open
reading frame (ORF) was identified, which encodes proteins and promoter sequences,
and these data were compared with those already known. A comparison of amino acid
sequences ORF of unknown function with known sequences (homologous search)
11
made it possible to identify proteins with similar sequences and motifs. Such an analy-
sis provides the prediction of the possible functions of ORF in a cell without conducting
biochemical experiments.
Data on the sequence of genes are from EMBL (EU) and GenBank (US). These data
can be found on the web pages of the National Institute of Health (http://www.ncbi.
nlm.nih.gov) or the European Institute of Bioinformatics (http://www.ebi.ac.uk/Tools/
index.html). With programs such as BLAST (basic local alignment search tool) or
FASTA, it is possible to compare DNA or amino acid sequences with known ones.
Personal computers and Internet allow you to find this information on websites
around the world (Table 1.3).
Table 1.3. Genomics and proteomics websites
Website name Information
http://web.bham.ac.uk/bcm4ght6/genome.html Gene database of E. coli
http://cgsc.biology.yale.edu/ Genetic center of E. coli
http://www.ucalgary.ca/%7Ekesander/ Genetic center of Salmonella
http://genome.wustl.edu/gsc/bacterial/salmonella.shtml Genome of Salmonella
http://ecocyc.PangeaSystems.com:1555/server.html Different biochemical paths in E. coli

Biochemical pathways of many
http://www.genome.ad.jp/kegg/
sequenced organisms
http://bomi.ou.edu/faculty/tconway/global.html Functional genomics of E. coli
http://susi.bio.uni-giessen.de/ecdc/ecdc.html Database of E. coli
http://www2.ebi.ac.uk/fasta33/ Homologues search
http://www.motif.genome.ad.jp/ Search for motifs in proteins

Structures of two-dimensional
http://www.expasy.ch/cgi-bin/map1
electrophoresis
http://www2.ebi.ac.uk/clustalw/ Nucleotide and amino acid sequences
http://www.ncbi.nlm.nih.gov/PubMed/ Search for literature in PubMed
http://bmbsgi11.leeds.ac.uk/bmbknd/DNA/ genomic.htm Genome databases
http://www.biology.ucsd.edu/~msaier/transport/ Information about the transport system
The first complete sequence of the bacterial genome, Haemophilus influenzae, was
published in 1995. In 2002, more than 50 genomes of microorganisms were screened.
Much more of them are in the process of sequencing.
Computer analysis enables to determine the properties of proteins encoded in
a certain DNA sequence, their isoelectric points, molecular weights, three-dimensional
structures, etc. as well as the potential regulatory regions; promoters can be deter-
mined in the DNA sequence.
12
Particularly complete information can be obtained from KEGG (Kyoto Ency

clopedia of Genes and Genomes) at the website: http://www.genome.ad.jp/kegg/. In
this encyclopedia, the biochemical systems of some microorganisms are graphically
illustrated and genomes of some of them are sequenced. The name of the microor-
ganism and the complete information about the presence of its metabolic pathway,
enzymes of this path, the DNA sequence, and proteins of the desired path can be found
on “Metabolic paths.” This example shows that due to the extensive development of
genomics and the Internet, microbial physiology has been completely changed. Howe
ver, the greatest misconception would be that the homology found guarantees the func-
tioning of this protein or the given regulatory site in the microorganism under investi-
gation. Gene functions must be confirmed by genetic and biochemical methods.
A classic approach to determining the biochemical pathway is to use a selective
phenotype or to get a mutant that damages a certain stage of the metabolic pathway
using a selective medium. If such a mutant is received, it is possible to map the gene,
identify, clone, and sequence. However, it is not always easy to get such a mutant; it is
necessary to analyze hundreds of thousands of colonies. If there is a positive selection
method (after mutagenization, only mutant clones grow on the medium), it is easier
to do so. If selection is negative (the mutant does not grow on the selective medium),
then it is necessary to make impressions on an environment that does not contain this
growth factor and to select one colony with the desired phenotype from one thousand
colonies.
In recent years, the selection of mutants has been greatly facilitated by the develop
ment of the method of fluorescence-activated cell sorting (FACS). For this purpose,
a gene encoding a green fluorescent protein fused to a gene that is not expressed under
certain conditions (e.g., at low pH 4.5) and cannot form a colony is used. After
mutagenization of the fusion strain, the mutants are cultivated at high and then
changed pH and passed through a device that selects fluorescing and nonfluorescing
cells. This approach has greatly improved the work on selecting mutants.
Cloning and DNA analysis. The process of molecular cloning (selective accumu-
lation) of DNA molecules involves several steps:
•• fragmentation of DNA by processing endonuclease restriction;
•• combining these fragments in vitro with a vector DNA molecule (capable of
autonomous replication);
•• introduction of the vector into the recipient organism, in which recombinant
DNA is accumulated.
Genetic engineering was born in 1973, when the first recombinant DNA was cre-
ated in vitro by P. Berg’s group in the United States.
Finding out how the cell coordinates biochemical reactions is an important step
toward understanding of the concept of life. Unfortunately, a universal method for
studying all biochemical, structural, and physiological features of the cell does not
exist yet. However, today there are techniques that are most suitable for this purpose.
These include gene arrays and two-dimensional division of cellular proteins. These
13
methods bring us closer to understanding how one system in a cell can determine the
synthesis of all other systems.
The technology of gene arrays makes it possible to consider the expression of
genes at the level of the complete genome. The expression of each gene can be analy
zed many times due to the use of microchips in DNA.
The basic idea of this method is as follows: DNA sequences of each gene are
placed on a solid surface to allow rapid hybridization with a fluorescently labeled DNA
pool (or cDNA). DNA sequences isolated from individual genes are placed on the chip
as separate grid spots (Fig. 1.6). The square of the area is 1–2 square inches (2.5 cm2).
Further, RNA is rapidly extracted from culture, transferred to complementary DNA
(cDNA) using inverse transcriptase (RNA-dependent DNA polymerase). Due to the
inclusion of fluorescently labeled nucleotides in the incubation mixture, each cDNA
molecule is tagged. It binds to specific loci on the chip, which contains a separate
isolated gene.
Laser scanning and fluorescence measurements allow the chip surface to be read
out, and computer analysis detects genes that are expressed in original samples taken.
By applying such an analysis, one can compare the level of expression of individual
genes isolated from cultures grown under different conditions. The cDNA drugs are
labeled with different fluorescence labels. If the gene is expressed only under certain
conditions, the corresponding spot will be fluoresced in one color; if under other con-
ditions, the spot will have another color. If the gene is expressed in both conditions, the
color becomes mixed. Such an analysis can be used to study the effects of conditions
of cultivation, stress factors, and mutations on gene expression.
Proteomic analysis. Unlike a genome that has a constant composition, proteins
can vary depending on the internal and external environment. Under the influence of
stress factors, the level of expression of some proteins is reduced and of other proteins
increases. Gene array technology makes it possible to analyze changes in the protein
composition caused by the transcription level, and proteomic analysis takes into account
post-transcription and post-translational changes.
For proteomic analysis, two-dimensional electrophoresis in polyacrylamide gel
is used (Fig. 1.7). The cell-free extract containing a mixture of 1000–2000 different
proteins is initially subjected to isoelectric focusing; it is a process that allows the
proteins to be separated by differences in their isoelectric points. Each protein con-
tains carboxyl groups and amino groups, which are differently protonated and have
charge depending on pH. For certain pH values, they are separated. However, identi-
cal isoelectric points can have multiple proteins, so electrophoresis should be used in
polyacrylamide gel. Proteins move depending on their molecular weight, faster with
a lower molecular weight.
Proteins can be visualized by a radioactive label or dye (e.g., Kumasi). Then,
computer analysis should be used to compare the proteins. A protein selected for
further analysis can be analyzed in detail. For this purpose, genomic and proteomic
analysis is used. If a gene of a microorganism is sequenced, the protein is cut and sub-
jected to mass spectrometry. To do this, it is cut by proteases (trypsin) into fragments,
14
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CHAPTER XXI.
cyril’s nurses.
While the church bells were ringing daily in the tower above his
head, and the old Bridford chimes, famous long ago, were heralding
the birth of every hour with a fine old psalm tune that pealed out over
the busy, money-making city, like an echo of the past, Cyril
Culverhouse was lying at the bottom of a dark gulf of pain and
confusion, and all the outer world, and all the life that he had lived,
were cancelled and forgotten.
Strange images danced before his eyes like motes in the
sunshine, yet seemed to him neither strange nor unexpected. He
had a history of his own in that period of delirium, a new identity, new
surroundings, a mad, wild world, peopled out of his own brain.
Bishops and archbishops came and sat beside his bed, and held
long arguments with him, figments of a mind distraught, the shadows
that haunt fever-dreams, but to him intensely real. The dead came
back to life to hold converse with him, and he was not surprised. No,
he had always thought there was something in the ideas of the old
necromancers. The elixir of life was not an impossibility. Here was
Luther with his square solid face, and sensual humorous mouth.
Here was Pascal, full of quaint sayings and far-reaching thoughts.
The sick man talked for all of them. His talk was wildest raving to the
ears that listened, but to his own fancy it was profoundest wisdom.
There is no egotism, no belief in self, equal to that of the lunatic. For
him the stars and moon have been made, for him God willingly
performs miracles which overthrow all the laws of the universe. He is
the axis of the world, and lets it go round.
How long those days and nights of fever were! That was the chief
agony of them. The eternity of hours, so thickly peopled with
distorted shapes that every quarter of an hour was an era. Of actual
physical pain the sufferer had no consciousness; but weariness,
almost too heavy to be borne weighed upon him in the long strange
nights, when the faces of his watchers changed, and the very walls
of his room seemed new and unknown to him. He fancied that his
nurses had removed him into new lodgings while he slept, though it
seemed to him that he had never slept.
Sometimes he fancied himself in one place, sometimes in another.
He was at Oxford, in those old rooms of his looking into the college
garden. He was at Little Yafford, at Culverhouse, anywhere but
where he really was.
And his nurses, who were they? He faintly remembered Mrs.
Podmore leaning over his bed, fat and scant of breath, with a
medicine-glass in her hand, coaxing him to drink. He remembered
Sarah, making believe to step softly, in creaking shoes, whose every
movement was agony to him. But these things were lost in the
darkness of remote ages. His present nurses seemed to have been
tending him during a century.
There were two, one tall and slender, dignified of bearing, yet
gracious in every movement; the other short, small, and brisk. They
were dressed exactly alike, in the costume of some religious order,
as he supposed. They wore long black robes and white linen caps,
such as he remembered to have seen worn by the Sisters of Mercy
in Breton towns that he had visited years ago in one of his long
vacations. Admirable caps for ugly women, for the stiff linen borders
projected a quarter of a yard beyond the face, entirely concealed the
profile, and overshadowed the countenance at all times.
Cyril knew only that the taller of his two nurses had dark eyes and
a pale face, and that the little woman had black eyes of exceeding
sharpness, that flashed at him from the cavernous cap. They were
both admirable nurses, quiet, gentle, attentive, but in some phases
of his delirium he hated them, and accused them of all manner of evil
designs. They were poisoning him. Yes, the medicine they made him
take at stated intervals contained a slow poison—the Aqua Tofana of
the Middle Ages—that horrible stuff which the wicked witch Toffania
made by wholesale, and sent to all the cities of the earth as the
manna of St. Nicola of Bari; or it was the hemlock that Socrates
drank, or wolf’s bane, or deadly nightshade. He recognised the
flavour of the murderous herb. And then he stormed at his nurses,
and told them they had plotted his murder.
‘If you were honest women you would not hide your faces,’ he
cried. ‘You are murderesses, and have come here to kill me.’
One night, after an age of fever and hallucination, he sank into a
refreshing slumber. It was as if his spirit, newly escaped from a
burning hell, had slipped unawares into Paradise. Fair meadows and
flowing streams, an ineffable sense of coolness and relief, and then
deep rest and stillness.
When he awoke, the summer dawn filled the room. Through the
widely opened windows came the fresh breezes of the morning. A
soft cool hand was on his brow, the tall nurse’s dark figure stood
beside his bed.
All his delusions, all his hideous fancies, seemed to have run out
of his brain, like water out of a sieve, during that one sweet sleep.
Suddenly and completely as the leper at the Divine Healer’s bidding,
he was made sound and whole. Very weak still, with a sense of utter
helplessness and prostration, he yet felt himself cured. The fire that
had made life a torture had burnt itself out.
He looked up at his nurse. How purely white that quaint old head-
gear of hers looked in the morning sunshine. He remembered the
bright freshness of just such another morning in his holiday rambles
five years ago, and just such another black-robed figure and white
cap, a Sister of Mercy waiting for the starting of the diligence, in the
old market square at Vannes, the white dusty square, the scanty
trees, that seem to have been planted yesterday, the shabby old
cathedral looking down at him.
‘You are a Frenchwoman, are you not?’ he asked, the weakness
of his voice startling him a little.
‘Mais si,’ she answered, gently.
He tried to get her to talk, but she answered him only in
monosyllables. He tried to see her face, but the position in which she
held her head always prevented him.
‘Perhaps her cap is the prettiest thing about her, and she would
rather show that than her face,’ he thought.
Even that brief conversation exhausted him, and he fell asleep
again. Those weary hours of delirious wakefulness had left him long
arrears of sleep to make up. He slept on till dusk, and Dr. Saunders,
finding him locked in that deep slumber, pronounced him out of
danger.
‘Our medicines have never been able to touch him,’ he said
frankly. ‘It has been an unaided struggle between nature and
disease. I ought not to say unaided, though,’ he added,
apologetically, to the little nursing sister in the Breton cap. ‘Your care
has been a very powerful assistance.’
The little woman thanked him effusively in her broken English. The
taller nurse spoke only French, and as little of that as possible.
When Cyril awoke again, just before nightfall, the small nurse was
sitting by his bed.
‘Where is the other?’ he asked.
‘Gone.’
‘Gone?’
‘Yes. You are now much better—on the high road to recovery. You
no longer want two nurses. My companion has gone home.’
‘She is wanted for some other case, perhaps.’
‘No doubt she soon will be.’
‘To what order do you belong?’
‘To a community of nursing sisters.’
‘In Brittany?’
‘Yes.’
‘What part of Brittany?’
‘We never talk about ourselves. It is one of the rules of our order.
We come and go like the wind.’
‘But how was it that you came to me? Who sent for you?’
‘We were not sent for. We happened to hear of your illness—and
we knew you were a good man. It was our duty to come and nurse
you.’
‘What me?—a Protestant?’
‘We are not sectarian. We go wherever we are wanted.’
‘But how do you—Breton nuns—come to be in England?’
‘We are not nuns. We are a nursing sisterhood, bound by no vows.
We heard of the pest raging in this town, and came here to be
useful.’
‘You are very good people,’ said Cyril. ‘I am sorry the other sister
is gone. I should like to have talked to her, but this morning she
would answer me only in monosyllables.’
‘It is not good for you to talk, and it is one of our rules to talk as
little as possible.’
For three days the figure in the loose black gown was constantly at
Cyril’s bedside. He heard the little woman telling her beads in the
dead of night. If she were no nun she was at any rate a staunch
Roman Catholic; but she did not endeavour to convert him to her
own creed. She was a modest, unobtrusive little woman; but during
those three days she very often broke the rule of her order, and
talked to the patient a good deal. She talked of Brittany, which she
knew thoroughly, and sometimes of modern French literature, which
she knew better than she ought to have done as a member of a
religious sisterhood.
On the fourth day she was gone, and another figure, dressed in
black, with neat white cap and apron, was by Cyril’s bedside. The
face of this watcher was not hidden. He knew it well, a homely
English face that brought back the thought of his work in the courts
and back streets of Bridford.
‘Mrs. Joyce,’ he exclaimed. ‘Have you turned nurse?’
‘What more blessed privilege can I have, sir, than to take care of
you? I owe you what is a great deal more to me than my own life, the
life of my beloved son. Oh, sir, if he ever comes to be a Milton or a
Shakespeare, the world will bless you for your goodness, as I do
now.’
Cyril smiled at her enthusiasm. Perhaps every mother whose son
writes obscure verses in doubtful English believes with Mrs. Joyce
that she has produced a Milton.
‘I should have come before, sir, if the two ladies hadn’t been here.
But they were such good nurses I didn’t want to interfere with them.’
‘Do you know where they came from, or why they came?’
‘No, indeed, Mr. Culverhouse. They were foreigners, and I
suppose they came from foreign parts.’
‘Neither of my doctors sent for them, I believe.’
‘No, sir. Dr. Saunders told me they came and went like spirits, but
he was wishful there were more like them.’
‘And your son is really recovered?’
‘Yes, sir. It is a most wonderful cure. He rallied that night, and was
up and about at the end of the week. To both of us it seemed like a
miracle. I have read the gospel about the widow’s son every night
and morning after my prayers, and I have read it two or three times
to Emmanuel. Oh, sir, I hope and believe you have wrought a double
cure. I think my son’s heart is turned to holy things. He has read his
Bible very often lately. I have watched him, and I think he is
beginning to find out that there is truth and comfort to be found in it.’
‘He cannot read the gospel long without making that discovery.
Young men are too apt to form their judgment of the Bible from what
other people have written about it. When they go to the fountain
head they find their mistake.’
Cyril was not satisfied till he had questioned Dr. Saunders and Dr.
Bolling, the latter of whom had come to see him daily, without any
fee, about the two French nurses. But neither of these could tell him
more than he knew already.
‘I wish I did know more about them,’ said Mr. Saunders. ‘Whatever
institution they belong to, it’s an admirable one, and I’m sorry we
haven’t a few more institutions of that kind over here. I don’t think we
should have pulled you through if it hadn’t been for that excellent
nursing. No, upon my word I believe you owe those two women your
life.’
‘And I do not even know their names, or where they are to be
found,’ said Cyril, regretfully.
It worried him not a little to be under so deep an obligation, and to
have no mode of expressing his gratitude. At one time he thought of
putting an advertisement in the Times, thanking his unknown nurses
for their care. But on reflection this seemed idle. They were
doubtless what they represented themselves, sisters of some
religious order, who did good for the love of God. They had no need
of his thanks. Yet he puzzled himself not a little about the whole
business. Why should he have been selected, above all other
sufferers in the town of Bridford, as the recipient of this gratuitous
care?
As soon as he was able to leave his bed, Dr. Bolling insisted on
his going off to the sea-side to get strength before he went back to
his work. This vexed him sorely, but he could not disobey.
‘You’ve been as near the gates of death as a man can well go
without passing through them,’ said the doctor.
end of vol. ii.
J. AND W. RIDER, PRINTERS, LONDON.
Corrections
The first line indicates the original, the second the correction.
Contents to Vol. II
IX. ‘THOSE ARE THE KILLING GRIEFS WHICH DARE NOT SPEAK’ 138
IX. ‘THOSE ARE THE KILLING GRIEFS WHICH DARE NOT SPEAK’ 128
p. 137
Miss Scales eat her dinner

Miss Schales ate her dinner
p. 172
Yes, it’s regretable.

Yes, it’s regrettable.
p. 268
in His earthly pilgrimage did He exereise that ineffable

in His earthly pilgrimage did He exercise that ineffable
p. 305
Mrs. Piper no longer recived her morning visitors in it

Mrs. Piper no longer received her morning visitors in it
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Bacterial Physiology and Biochemistry Ivan Kushkevych Full Chapter

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Bacterial Physiology and Biochemistry

Copyright © 2023 Elsevier Inc. All rights reserved.

For information on all Academic Press publications

Publisher: Stacy Masucci

Cover input from Igor Starunko

CHAPTER 1. INTRODUCTION INTO BACTERIAL PHYSIOLOGY

CHAPTER 2. BACTERIAL CHEMICAL COMPOSITION AND FUNCTIONAL

CHAPTER 3. BACTERIAL NUTRITION AND GROWTH................................................. 91

CHAPTER 4. PROCESSES OF CELL DIFFERENTIATION.............................................. 151

CHAPTER 5. BACTERIAL METABOLISM....................................................................... 159

CHAPTER 6. GROWTH OF MICROORGANISMS IN NATURE ....................................311

CHAPTER 7. EFFECT OF ENVIRONMENTAL FACTORS

ATP Adenosine triphosphate

All things are hidden, obscure and debatable if the cause

B acterial Physiology and Biochemistry is an educational book designed for

Prof. Lorenzo Drago, Ph.D.,

INTRODUCTION INTO BACTERIAL

1.1. Subject of study

Prokaryotic and eukaryotic cells differ in structural organization. Prokaryotes,

Table 1.1. Comparison of bacteria, archaea, and eukaryotes

Properties Bacteria Archaea Eukaryotes

Nucleus with a nuclear membrane

Complex of internal membrane

Have peptidoglycan, Do not have Do not have

Esterified, Esterified, branched, Esterified,

Gas vesicles Present Present Absent

Contains thymine, Does not contain Contains thymine,

Polycistronic mRNA Present Present Absent

Introns of the mRNA Absent Absent Present

Splicing of the mRNA Absent Absent Present

DNA-dependent RNA polymerase

Polymerase II promoters Absent Present Present

The comparison of similar properties of organisms, regardless of the presence of

Contain molecules that transmit Capable of grown and

Cells of all organisms, regardless of the presence of the nucleus,

Have a cellular metabolism, in which

Fig. 1.1. Comparison of similar properties of organisms.

Thus, prokaryotic and eukaryotic cells differ in structural organization, although

1.2. Bacteria in the phylogeny of living organisms

The conducted studies allowed determining:

Bacteria Archaea Eukaryota

Fig. 1.2. Phylogenetic tree of life by Carl Woese et al. (1990).

Spherical (cocci) Rods (bacillus)

Vibrio Spirillum Spirochete

Size, shape, and arrangement of bacterial cells (Table 1.2):

Table 1.2. The size of some bacterial cells

Organism Size (µm) Organism Size (µm) Organism Size (µm)

Chlamydium About 0.3 Streptococcus 1.0–1.5 Clostridium 0.5–0.8×3.0–8.0

Bdellovibrio 0.3–0.5×0.5–1.4 Sarcina 4.0–4.5 Shigella 0.4–0.6×1.0–3.0

Rickettsia 0.1–0.3×1–4 Azotobacter 2–3×3–6 Spirochaeta 0.2–7×5–500

Staphylococcus 0.8–1.0 Proteus 0.5–1×1–3 Spirillum 0.5×3

Escherichia coli 0.5–1×1–3 Micrococcus About 1.0 Corynebacterium 0.32–0.8×1.0–8

Bacillus subtilis 0.9–1.1×1.8–4.8 Pseudomonas 0.3–0.5×1.0–1.8 Treponema 0.2×4–14

Mycobacterium 0.3–0.5×1–4 Acetobacter 0.4–0.8×1.0–2.0 Corynebacterium 0.3–0.8×1.0–8.0

Chromatium 3–4×5–6 Lactobacillus 0.7–1.0×3.0–8.0 Brucella 0.3–0.5×0.5–2

1.3. Bacterial evolution

Fig. 1.4. The scheme of life's origin based on Oparin–Haldane hypothesis.

Mitochondria and chloroplasts

The most significant evidence of the origin of chloroplasts and mitochondria of

1.4. Methods of studies of bacterial properties

Table 1.3. Genomics and proteomics websites

Website name Information

http://web.bham.ac.uk/bcm4ght6/genome.html Gene database of E. coli

Particularly complete information can be obtained from KEGG (Kyoto Ency