Microbial Diversity

Part 1: Acellular and Prokaryotic Microbes Viruses are classified by the following characteristics:
a) type of genetic material (either DNA or RNA),
Microbiology is the study of microbes, most of which b) shape of the capsid,
are too small to be seen by the naked eye. c) number of capsomeres,
d) size of the capsid,
2 Categories of microbes: Cellular and Accellular e) presence or absence of an envelope,
 Cellular - bacteria, archaea, algae, protozoa, fungi f) type of host that it infects,
o Prokaryotic – bacteria and archaea g) type of disease it produces,
o Eukaryotic – algae, protozoa, and fungi h) target cell, and
 Accellular - viruses, viroids, and prions i) immunologic or antigenic properties
o Not considered as living organisms
o more correctly referred to as acellular microbes  its genome is mostly either double-stranded DNA or
or infectious particles single-stranded RNA,
 few viruses possess single-stranded DNA or double-
ACELLULAR MICROBES stranded RNA
Viruses  viral genomes are usually circular molecules, but
 complete virus particles, called virions, are very some are linear (having two ends)
small and simple in structure  may be polyhedral (many sided – 20 sides,
 extremely small, observed using electron microscope icosahedron), helical (coiled tubes), bullet shaped,
 10 to 300 nm in diameter, although some—like spherical, or a complex combination of these shapes
Ebola virus—can be up to 1 micrometer in length  Each facet (side) consists of several capsomeres;
 smallest virus is about the size of the large thus, the size of the virus is determined by the size of
hemoglobin molecule of a red blood cell each facet and the number of capsomeres in each
 oncogenic viruses or oncoviruses—cause specific  the envelope around the capsid makes the virus
types of cancer appear spherical or irregular in shape in electron
 viruses are not alive – to replicate, viruses must micrographs
 The envelope is acquired by certain animal viruses
invade live host cells because there are no ribosomes
as they escape from the nucleus or cytoplasm of the
for protein synthesis or sites of energy production
host cell by budding
 lack most of the basic features of cells; thus, they  Budding is a form of viral shedding where viruses
consider viruses to be non-living entities. acquire their external envelope from either the host
 How to distinguish virus from living cells cell’s nuclear membrane or cell membrane
o The vast majority of viruses possess either DNA  viruses are then able to alter these membranes by
or RNA, unlike living cells, which possess both. adding protein fibers, spikes, and knobs that enable
o They are unable to replicate (multiply) on their the virus to recognize the next host cell to be
own; their replication is directed by the viral invaded
nucleic acid once it has been introduced into a
host cell Viruses have to infect a host cell to access the machinery
o Unlike cells, they do not divide by binary that it uses to replicate it DNA, and then hijack that
fission, mitosis, or meiosis. machinery in order to reproduce. The host cell is forced
o They lack the genes and enzymes necessary for to manufacture new viruses, which then leave and look
energy production. for new hosts to infect.
o They depend on the ribosomes, enzymes, and
metabolites (“building blocks”) of the host cell Origin of Viruses
for protein and nucleic acid production  existed for as long as bacteria/archaea have existed
 Typical virion consists of a genome of either DNA  Coevolution Theory
or RNA, surrounded by a capsid (protein coat), o Viruses originated in the primordial soup and
which is composed of many small protein units coevolved with bacteria and archaea.
called capsomeres (or capsomers). o Primordial soup generic term that describes the
 nucleic acid + capsid = nucleocapsid aqueous solution of organic compounds that
 Some viruses (enveloped viruses) have an outer accumulated in primitive water bodies
envelope composed of lipids and polysaccharides o This hypothesis has few supporters
 Bacterial viruses may have a tail, sheath, tail fibers.
  Phage attaches to a protein or
polysaccharide molecule (receptor) on the
surface of bacterial cell
 Retrograde Evolution Theory  The phage can only attach to bacterial cells
o viruses evolved from free-living prokaryotes that that possess the appropriate receptor—a
invaded other living organisms, and gradually protein or polysaccharide molecule on the
lost functions which were provided by host cell surface of the cell that is recognized by a
o Viruses arose from cellular organisms, most molecule on the surface of the phage
likely a bacterium. the cellular organisms  Most bacteriophages are species- and strain-
formed viruses by inquiring the ability to specific – they only infect a particular
gradually discard their genes until they become species or strain of bacteria.
simpler organisms which are the viruses we see  Those that infect E. coli are called
today coliphages.
o This theory has little support  Some bacteriophages can attach to more
 Escaped Gene Theory than one species of bacterium
o viruses are pieces of host cell RNA or DNA that 2. Penetration
have escaped from living cells, and are no longer  the phage injects its DNA into the bacterial
under cellular control cell, acting much like a hypodermic needle,
o most widely accepted explanation for the origin the capsid remains on the outer surface of
cell
of viruses
 then the phage DNA “dictates” what occurs
within the bacterial cell
Bacteriophages (phages)
 sometimes described as the phage DNA
 viruses that infect bacteria
taking over the host cell’s “machinery”
 obligate intracellular pathogens, they must enter a 3. Biosynthesis
cell to replicate  phage genes are expressed, resulting in the
 3 categories, based on their shape production (biosynthesis) of viral pieces
o Icosahedron bacteriophages: almost spherical  host cell’s enzymes (e.g., DNA polymerase
shape, with 20 triangular facets; smallest and RNA polymerase), nucleotides, amino
icosahedron phages are 25 nm in diameter acids, and ribosomes are used to make viral
o Filamentous bacteriophages: long tubes DNA and viral proteins
formed by capsid proteins assembled into a 4. Assembly
helical structure; they can be up to about 900 nm  viral pieces are assembled to produce
long complete viral particles (virions)/ complete
o Complex bacteriophages: icosahedral heads phages
attached to helical tails; they may also possess  viral DNA is packaged up into capsids
base plates and tail fibers 5. Release
 There are single-stranded DNA phages, double-  host cell bursts open and all of the new
stranded DNA phages, single-stranded RNA phages, virions (about 50–1,000) escape from the
and double-stranded RNA phages cell
 Virulent Bacteriophages  the complete phages escape from the
o do not actually enter the bacterial cell—rather, bacterial cell by lysis of the cell
they inject their nucleic acid into the cell  lytic cycle ends with lysis (destruction) of
o always cause the lytic cycle, which ends with the the host cell
destruction (lysis) of the bacterial cell
o the whole process (from attachment to lysis)  Lysis is caused by an enzyme (referred to as an
takes less than 1 hour endolysin) that is coded for by a phage gene.
o once it enters a host cell, it always initiates the  after assembly, the appropriate viral gene is
lytic cycle, which results in the destruction of expressed, the enzyme is produced, and the bacterial
cell cell wall is destroyed
o Lytic Cycle Steps  With certain bacteriophages, a phage gene codes for
1. Attachment an enzyme that interferes with cell wall synthesis,
 Adsorption of the phage to the surface of the leading to weakness and, finally, collapse of the cell
bacterial cell. wall

 Temperate Phages (lysogenic phages)

 o do not immediately initiate the lytic cycle,  fitting the virus pieces together/assembled to
rather, their DNA remains integrated into the produce complete virions
bacterial cell chromosome, generation after 6. Release
generation  After the virus particles are assembled, they
 Bacteriophages are involved in two (lysogenic must escape from the cell by lysis/budding
conversion and transduction) of the four major  Some animal viruses escape by destroying
ways in which bacteria acquire new genetic the host cell, leading to cell destruction and
information some of the symptoms associated with
infection with that particular virus
 Other viruses escape the cell by a process
known as budding – become surrounded
with pieces of the cell membrane, thus
becoming enveloped viruses.
Animal Viruses
 Viruses that infect humans and animals
 Some are DNA viruses; others are RNA viruses
 may consist solely of nucleic acid surrounded by a  Inclusion Bodies
protein coat (capsid), or more complex o remnants or collections of viruses
 ex; they may be enveloped or they may contain o often seen in infected cells and are used as a
enzymes that play a role in viral multiplication diagnostic tool to identify certain viral disease
within host cells o found in the cytoplasm (cytoplasmic inclusion
bodies) or within the nucleus (intranuclear
 Steps in Multiplication of Animal Virus inclusion bodies), depending on the disease
1. Attachment o cytoplasmic: rabies, AIDS, Guarnieri bodies of
 Phage attaches to a protein or polysaccharide smallpox
molecule (receptor) on surface of bacterial cell o intranuclear: herpes, poliomyelitis
 Animal viruses can only attach to cells bearing
the appropriate protein or polysaccharide Latent Virus Infections
receptors on their surface  ex: herpes, cold sores (fever blisters), shingles
 Why certain viruses cause infections in dogs, but (painful nerve disease caused by a herpesvirus)
not in humans or vv? RECEPTORS. Viruses  viral infections in which the virus is able to hide
can only attach to and invade cells that bear a form host’s immune system by entering cells and
receptor that they can recognize remaining dormant
2. Penetration  limited by the defense systems of the human body—
 entire virion usually enters the host cell, phagocytes and antiviral proteins called interferons
sometimes because the cell phagocytizes that are produced by virus-infected cells
(engulf/ingest) the virus  once acquired, herpres virus infection never
3. Uncoating completely go away
 the viral nucleic acid escapes from the
capsid Antiviral Agents
 the viral nucleic acid “dictates” what occurs  Antibiotics function by inhibiting certain metabolic
within the host cell activities within cellular pathogens, and viruses are
4. Biosynthesis not cells.
 many viral pieces (viral nucleic acid and  But antibiotics may be prescribed in an attempt to
viral proteins) are produced prevent secondary bacterial infections that might
 quite complicated, depending on what type follow the virus infection
(1 or 2 D/RNA strand) of virus infected the  Drugs used to treat viral infections are called
cell antiviral drugs
 some animal viruses do not initiate  It is important to understand that antibiotics are not
biosynthesis right away, but rather, remain effective against viral infections
latent within the host cell for variable  been developed to interfere to virus-specific
periods enzymes and virus production by either disrupting
 Latency is the phase in certain viruses' life critical phases in viral cycles/multiplications or
cycles in which, after initial infection, inhibiting the synthesis of viral DNA, RNA, or
increase of virus particles ends proteins
5. Assembly
  has the largest capsid diameter (440 nm) and largest
Oncogenic Viruses (oncoviruses) and most complex genome of all known viruses
 viruses that cause cancers  its genome is predicted to encode over 1,000
 cancer; virus different proteins; larger than that of some bacteria
 nasopharyngeal carcinoma, Burkitt lymphoma, B-
cell lymphoma; Epstein-Barr virus Plant Viruses
 Kaposi sarcoma; human herpesvirus 8  more than 1,000 different viruses cause plant
 Hepatocellular carcinoma; hepa B and C diseases
 usually transmitted via insects (e.g., aphids, leaf
Human Immunodeficiency Virus hoppers, and whiteflies); mites; nematodes (round
 Cause of AIDS (2 single strand RNA) worms); infected seeds, cuttings, and tubers; and
 enveloped, 2 single-stranded RNA virus contaminated tools (e.g., hoes, clippers, and saws)
 Genus: Lentiviruses, Family: Retroviridae  Viroids
 Can attach to and invade cells bearing receptors that o even smaller and less complex infectious agents
the virus recognizes o consist of short, naked fragments of single-
 Each of its 72 surface knobs contains a glycoprotein stranded RNA (about 300–400 nucleotides in
(gp120) that can bind to CD4 receptor of host cell length) that can interfere with the metabolism of
 CD4 is the most important receptors, CD4 cells: plant cells and prevent the growth of plants,
cells that possess CD4 receptors sometimes killing the plants in the process
 T cells: most important CD4 cells o infectious RNA molecules that cause a variety of
 HIV infections destroy these important cells of the plant diseases
immune system o transmitted between plants in the same manner
 Macrophage, a type of phagocyte, have CD4 as viruses
receptors, so can be invaded by HIV. o cause of potato spindle tuber, citrus exocortis, &
 able to invade certain cells that do not possess CD4 diseases in chrysanthemums, palms, tomatoes
receptors, but do possess other receptors that HIV is
able to recognize  Prions
o smaller and less complex infectious agents
Mimivirus o small infectious proteins that apparently cause
 large 2-stranded DNA virus recovered from amebas fatal neurological diseases in animals and
 named Mimivirus because it “mimics” bacteria humans
 very large – can be observed using a standard o scrapie in sheep and goats; bovine spongiform
compound light microscope encephalopathy (BSE; “mad cow disease”); and
 7-nm-thick capsid with a diameter of 750 nm kuru, Creutzfeldt–Jakob (C–J) disease,
 array of 80- to 125-nm-long closely packed fibers Gerstmann–Sträussler–Scheinker (GSS) disease,
project outward from the capsid surface and fatal familial insomnia in humans.
o All these diseases are fatal spongiform
 consists of a double-strand DNA core, surrounded
by two 4-nm-thick lipid membranes and a protein encephalopathies, in which the brain becomes
capsid riddled with holes (sponge-like)
o Stanley B. Prusiner, scientist who coined the
 numerous fibrils extend outward from capsid surface
 genome is at least 10 times larger than that of the term prion and studied the role of these
proteinaceous infectious particles in disease
large viruses in the smallpox family and larger than
the genome of some of the smallest bacteria o believed to be the most resistant to disinfectants
 possess close to 1,000 genes, some of its genes
codes for functions The Domain Bacteria
Characteristics
 contains several genes for sugar, lipid, and amino
acid metabolism, and some RNA molecules  2 domains of prokaryotes: Bacteria and Archaea
 reports said that it might be cause of pneumonia  23 phyla, 32 classes, 5 subclasses, 77 orders, 14
suborders, 182 family, 871 genera, and 5,007 species
Megavirus (megavirus chilensis)  broadly divided into three phenotypic categories
 Larger than mimivirus, double-stranded DNA virus (i.e., categories based on their physical
characteristics):
 discovered in a water sample collected in 2010 off
(a) those that are Gram-negative and have a cell wall
the coast of Chile
(b) those that are Gram-positive and have a cell wall
(c) those that lack a cell wall
 its Gram reaction, basic cell shape, and  staining methods have been devised to enable
morphological arrangement of cells are very scientists to examine bacteria
important clues to bacterium’s identification  bacteria are smeared onto a glass microscope slide
(resulting in what is known as a “smear”), air-dried,
Cell Morphology and then “fixed” (securely place/fastened)
 using light microscope, size, shape, and morphologic  2 methods of fixation: heat and methanol
arrangement of various bacteria are easily observed o Heat fixation
 Bacteria vary greatly in size, usually ranging from  accomplished by passing the smear through
spheres measuring about 0.2 µm in diameter to 10.0- a Bunsen burner flame
µm-long spiral-shaped bacteria, to even longer  too much heat can distort the morphology of
filamentous bacteria the cells
 Average coccus is about 1 µm in diameter, and the o Methanol Fixation
average bacillus is about 1 µm wide × 3 µm long  accomplished by flooding the smear with
 3 basic shapes: round or spherical (cocci, sing: absolute methanol for 30 seconds
coccus); rectangular or rod-shaped (bacilli, sing:  more satisfactory fixation technique
bacillus); curved and spiral (spirilla)  purpose of fixation
o Cocci: pairs (diplococci), chains (streptococci), o It kills the organisms.
clusters (staphylococci), 4s (tetrads), 8s (octads) o It preserves their morphology (shape).
o Bacilli: short or long, thick or thin, and pointed o It anchors the smear to the slide.
or with curved or blunt ends.  Specific stains and staining techniques are used to
: pairs (diplobacilli); chains (streptobacilli); long observe bacterial cell morphology (e.g., size, shape,
filaments; branched morphologic arrangement, composition of cell wall,
 Cocobacilli: rods that are quite short, capsules, flagella, endospores).
resembling elongated cocci.  simple stain is sufficient to determine bacterial
o Curved and spiral: usually occur singly, but shape and morphologic arrangement (pairs, chains,
some species may form pairs. clusters)
: referred as spirochetes (spayrokits) - vary in  Structural Staining Procedures: procedures used
size, length, rigidity, and the number and to observe bacterial capsules, spores, and flagella
amplitude of their coils  Gram Stain or Gram Staining Procedure
 divides by binary fission—one cell splits in half to o Dr. Hans Christian Gram, 1883
become two daughter cells. The time it takes for one o become the most important staining procedure,
cell to split into two cells is referred to as that because it differentiates between “Grampositive”
organism’s generation time. and “Gram-negative” bacteria
 After binary fission, the daughter cells may separate o Gram-Positive bacteria:
completely from each other or may remain  blue stained
connected, forming various morphologic  was not decolorized during the
arrangements decolorization
 may lose their characteristic shape because adverse  The thick layer of peptidoglycan in the cell
growth conditions (e.g., the presence of certain walls of Gram-positive bacteria makes it
antibiotics) prevent the production of normal cell difficult to remove the crystal violet–iodine
walls (cell wall–deficient bacteria or L-forms) complex during the decolorization step
 CWD bacteria revert to their original shape when  Peptidoglycan is a polymer consisting of
placed in favorable growth conditions sugars and amino acids that forms a mesh-
 Bacterial species having cells of different shapes is like layer outside the plasma membrane of
pleomorphic (able to assume different forms) most bacteria, forming the cell wall. ..
 Ex; Mycoplasmas - do not have cell walls; thus, o Gram-Negative bacteria
when examined microscopically, they appear in  red stained
various shapes. Because they have no cell walls,  crystal violet was removed from the cells
mycoplasmas are resistant to antibiotics that inhibit during the decolorization step, then cells
cell wall synthesis. were stained by the safranin (a red dye), they
will be pink to red
Staining Procedures  thin layer of peptidoglycan in the cell walls
 most bacteria are colorless,transparent, difficult to of Gram-negative bacteria makes it easier to
see remove the crystal violet–iodine complex
during decolorization
  the decolorizer dissolves the lipid in the cell  flagella stain can be used to demonstrate the
walls of Gram-negative bacteria; this presence, number, and location of flagella on
destroys the integrity of the cell wall and bacterial cells
makes it much easier to remove the crystal  can be demonstrated by stabbing the bacteria into a
violet–iodine complex tube of semisolid agar or by using the hangingdrop
o Gram-variable Bacteria technique
 Neither blue to purple nor pink to red after o Hangingdrop: a drop of culture (population of
Gram staining bacteria grown in the laboratory) is placed on a
 Ex are members of the genus coverslip that is encircled with petroleum jelly
Mycobacterium, such as M. tuberculosis and (or any sticky material). The coverslip and drop
M. leprae are then inverted over the well of a depression
 Mycobacterium species are more often slide. The drop hangs from the coverslip, and
identified using a staining procedure called petroleum jelly forms a seal that prevents
the acid-fast stain evaporation. When examined microscopically,
 Acid-Fast Stain motile bacteria within the “hanging drop” will
 Paul Ehrlich, 1882—a German chemist be seen moving fast around in every direction.
 carbol fuchsin <fyuksin> (a bright red dye) o Growth (multiplication) of bacteria in semisolid
is driven into the bacterial cell using heat agar produces turbidity (cloudiness). Nonmotile
(by flooding the smear with carbol fuchsin, organisms will grow only along the stab line (so
and then holding a Bunsen burner flame turbidity will be seen only along the stab line).
under the slide until steaming of the carbol Motile organisms will spread away from the stab
fuchsin occurs). line (thus, producing turbidity throughout the
 The heat is necessary because the cell walls medium)
of mycobacteria contain waxes, which
prevent the stain from penetrating the cells, Colony Morphology (appearance of colonies)
so heat softens the waxes, enabling the stain  single bacterial cell that lands on the surface of a
to penetrate. solid culture medium cannot be seen, but after it
 Then a colorizing agent (a mixture of acid divides over and over again, it produces a mound or
and alcohol) is then used in an attempt to pile of bacteria, known as a bacterial colony
remove the red color from the cells (contains millions of organisms)
 Because mycobacteria are not decolorized  colony varies from one species to another
by the acid–alcohol mixture (again owing to  includes the size, color, overall shape, elevation, and
the waxes in their cell walls), they are said the appearance of the edge or margin of the colony
to be “acid-fast.”
 colony features serve as important “clues” in the
 Other bacteria are decolorized by the acid–
identification of bacteria
alcohol treatment; they are non–acid-fast
 size of colonies is determined by the organism’s rate
o Gram reaction serves as an extremely important
of growth (generation time), and is an important
“clue” when attempting to learn the identity
characteristic of a particular bacterial species
(species) of a particular bacterium
 also includes the results of enzymatic activity on
o color of the bacteria at the end of the Gram
various types of culture media
staining procedure depends on the chemical
composition of their cell wall
Atmospheric Requirements
 Gram and acid-fast staining procedures are referred
 in microbio lab, it is useful to classify bacteria on the
to as differential staining procedures because they
basis of their relationship to O2 and CO2
enable microbiologists to differentiate one group of
 In a liquid medium such as thioglycollate broth
bacteria from another
(THIO), the region of the medium in which the
organism grows depends on the oxygen needs of that
Motility
particular species
 A bacterium is “motile” if it is able to swim
 as per oxygen, a bacterial isolate can be classified
 A bacterium is “nonmotile” if it is unable to swim
into one of five major groups:
 most often associated with the presence of flagella or a. obligate aerobes
axial filaments. Bacteria do not have cilia - require an atmosphere containing molecular
 Most spiral-shaped bacteria and 1/2 of the bacilli are oxygen in concentrations comparable to that
motile by means of flagella, but cocci are nonmotile found in room air (i.e., 20%–21% O2) to grow
and multiply
b. microaerophilic aerobes (microaerophiles)
 - requires reduced oxygen concentration (5%)  some bacteria are characterized by the production of
- ex: Neisseria gonorrhoeae (causes certain gases, such as carbon dioxide, hydrogen
gonorrhea) and Campylobacter spp. (causes sulfide, oxygen, or methane
diarrhea)  bacteria are inoculated (treated) into various
c. facultative anaerobes substrates (e.g., carbohydrates and amino acids) to
- capable of surviving in either the presence or determine whether they possess the enzymes
the absence of oxygen; anywhere from 0% necessary to break down those substrates
O2 to 20% to 21% O2.  when particular organism is able to break down a
- members of the family Enterobacteriaceae, certain substrate, it serves as a clue to the identity of
most streptococci, and most staphylococci that organism
d. aerotolerant anaerobes
- does not require oxygen, grows better in the Pathogenicity
absence of oxygen, but can survive in  ability of virus to cause disease
atmospheres containing molecular oxygen  Many pathogens are able to cause disease because
(such as air and a CO2 incubator) o they possess capsules, pili, or endotoxins
e. obligate anaerobe (biochemical components of the cell walls of
- will grow where there is 0% O2 Gramnegative bacteria)
o they secrete exotoxins and exoenzymes that
Anaerobes
damage cells and tissues
o organisms that do not require oxygen for life and
 tested by injecting the organism in mice or cell
reproduction
culture
o they vary in their sensitivity to oxygen
o e.g., certain Neisseria, Campylobacter, and Genetic Composition
Haemophilus species  Molecular Diagnostic Procedures - test procedure
 as per CO2, room air contains less than 1% CO2 that analyzes the organism’s DNA or RNA
 Capnophiles (capnophilic organism)  DNA of an organism is unique to each species
o grow better in the laboratory in the presence of  DNA probes
increased concentrations of CO2. o make it possible to identify an isolate without
o e.g., Bacteroides and Fusobacterium species relying on phenotypic characteristic
 In microbio labs, , CO2 incubators are routinely o single-stranded DNA sequence that can be used
calibrated to contain between 5% and 10% CO2 to identify an organism by hybridizing with a
unique complementary sequence on the DNA or
rRNA of that organism
 trough the use of 16S rRNA sequencing, the degree
of relatedness between 2 different bacteria can be
Nutritional Requirements determined
 bacteria need some form of the elements CO, H2O,
O2, sulfur, phosphorus, nitrogen, etc. for growth Unique Bacteria (rudimentary)
 Certain microbes have specific vitamin  do not possess all attributes of typical bacterial cells
requirements, and some need organic substances  they are so small and difficult to isolate, so they
secreted by other living microorganisms during their were formerly thought to be viruses
growth
 Rickettsias, chlamydias, and mycoplasmas
 Fastidious (fussy): organisms with especially
demanding nutritional requirement Rickettsias, Chlamydias, and Closely Related
 usually characteristic of a species of bacteria and Bacteria
serve as important clues when attempting to identify  bacteria with a Gram negative–type cell wall
the organism
 obligate intracellular pathogens (must live within a
host cell) that cause diseases in humans and other
Biochemical and Metabolic Activities
animals
 As bacteria grow, they produce many waste products
 to grow in laboratory, they must be inoculated
and secretions, some are enzymes that enable them
(treated) into embryonated chicken eggs, laboratory
to invade their host and cause disease
animals, or cell cultures
 pathogenic strains of many bacteria, such as
staphylococci and streptococci, can be tentatively Rickettsia
identified by the enzymes they secrete
 named after Howard T. Ricketts, a US pathologist
 have no connection to the disease called rickets,  can be cultured on artificial media in the laboratory,
which is the result of vitamin D deficiency where they produce tiny colonies (called “fried egg
 appear to have leaky cell membranes colonies”) that resemble sunny-side-up fried eggs
 most rickettsias must live inside another cell to  absence of a cell wall prevents mycoplasmas from
retain all necessary cellular substances staining with the Gram staining procedure
 diseases caused by Rickettsia species are arthropod-
borne, they are transmitted by arthropod carriers Especially Large and Especially Small Bacteria
 lice, fleas, and ticks transmit the rickettsias from one  size of a typical
host to another by their bites or waste products o coccus is 1 micrometre (µm) in diameter
 ex: typhus and typhus-like diseases (e.g., spotted o bacillus is 1.0 µm wide × 3.0 µm long, some are
fever rickettsiosis), involve production of a rash long thin filaments—up to about 12 µm in
 bacterias closely related to rickettsia length or even longer, but still 1 micrometre
o Coxiella burnetii - transmitted primarily by wide
aerosols, but can be transmitted to animals by  most bacteria are microscopic, requiring the use of a
ticks microscope to be seen
o Bartonella Quintana – associated with trench  Thiomargarita namibiensis
fever (a louse-borne disease), cat scratch disease, o Largest bacteria, can be seen with unaided eye
bacteremia, and endocarditis o colorless, marine, sulfide-oxidizing bacterium
o Ehrlichia spp. and Anaplasma spp. o its single spherical cells are 100 to 300 µm, but
- cause human tick-borne diseases such as may be as large as 750 µm (0.75 mm)
human monocytic ehrlichiosis (HME) and o do not form filaments
human granulocytic anaplasmosis.  Epulopiscium fishelsonii
- intraleukocytic pathogens, meaning that they o Also a large bacterium
live within certain types of white blood o isolated from the intestines of the reef
cells. surgeonfish
o a bacillus that is about 80 µm wide x 3600 µm
Chlamydia (energy parasites) (0.6 mm) long.
 refers to Chlamydia spp. and closely related o Its cells are about five times longer than
organisms (such as Chlamydophila spp.) eukaryotic Paramecium cell
 Although they can produce ATP molecules (major o Its volume is about a million times greater than
energy-storing molecules of cells), they prefer to use the volume of a typical bacterial cell
ATP molecules produced by host cells  Nanobacteria – Tiny bacteria
 obligate intracellular pathogens (must live within a o sizes are expressed in nanometers because these
host cell) that are transferred by inhalation of bacteria are less than 1 µm in diameter
aerosols or by direct contact between hosts—not by o have been found in soil, minerals, ocean water,
arthropods
human and animal blood, human dental calculus
 Chlamydia trachomatis: cause different diseases, (plaque), arterial plaque, and even rocks
including trachoma (leading cause of blindness) (meteorites) of extra-terrestrial origin
 Chlamydophila pneumonia: cause pneumonia o scientists believe that they were formed by
 Chlamydophila psittaci: causes a respiratory disease geological, rather than biological, processes
called psittacosis

Mycoplasmas (former pleuropneumonia-like organisms) Photosynthetic Bacteria

 smallest of the cellular microbes  include purple bacteria, green bacteria, and
 lack cell walls, so they assume many shapes, from cyanobacteria (blue-green algae)
coccoid to filamentous; thus, they appear  use light as an energy source, but they do not all
pleomorphic (assume many forms) when examined carry out photosynthesis in the same way
 may be free-living or parasitic and are pathogenic to  capable of converting light energy to chemical
many animals and some plant energy, ex: cyanobacteria
 In humans, pathogenic mycoplasmas cause primary  oxygenic photosynthesis
atypical pneumonia and genitourinary infections; o photosynthesis that produce oxygen
some species can grow intracellularly o cyanobacteria
 they have no cell wall, so they are resistant to
treatment with penicillin and other antibiotics that
 anoxygenic photosynthesis
work by inhibiting cell wall synthesis (formation)
 o photosynthesis that does not produce oxygen  Many, but not all, archaea are “extremophiles,”
o purple and green bacteria they live in extreme environments, such as
 In photosynthetic eukaryotes (algae and plants), extremely acidic, alkaline, hot, cold, or salty
photosynthesis takes place in plastids environments, or environments where there is
 Cyanobacteria extremely high pressure
o Exists 3.3 to 3.5 billion years ago  Other archaea, called methanogens, produce
o vary widely in shape; some are cocci, some are methane, which is a flammable gas
bacilli, and others form long filaments.  all archaea possess cell walls, but their cell walls
o In cyanobacteria, photosynthesis takes place on contain no peptidoglycan (all bacteria contains)
intracellular membranes known as thylakoids  16S rRNA sequence data suggest that archaea are
o Thylakoids are attached to the cell membrane at more closely related to eukaryotes than they are to
various points and are thought to represent bacteria
invaginations of the cell membrane
 the action or process of being turned inside GRAM STAINING PROCEDURE - SIMPLIFIED
out or folded back on itself to form a cavity  the gram stain is the most widely employed staining
or pouch in microbiology. it is a differential stain because it
o Attached to the thylakoids, in orderly rows, are divides bacteria into two classes - gram positive and
numerous phycobilisomes—complex protein gram negative.
pigment aggregates where light harvesting
occurs  in the first step of the gram stain procedure, cells
o Believed by scientists believe as the first from a fresh culture are transferred to a clean slide
organisms capable of carrying out oxygenic and allow it to air dry. the cell should form a thin
photosynthesis and, thus, played a major part in
barely visible film that is achieved by smearing the
the oxygenation of the atmosphere
cells obtained from the culture. fresh cultures must
o If certain conditions exist, Cyanobacteria in
pond or lake water overgrow, then it creates a be used because as cells age they lose their ability to
water bloom—a “pond scum” that resembles a retain stain.
thick layer of bluish green (turquoise) oil paint.
o Conditions include  the cells are then fixed to the slide, so using the heat
 mild or no wind fixations, pass the smear slightly above the flame of
 balmy water temperature (15°–30°C) a bunsen burner but not too much because too much
 water pH of 6 to 9 heat can distort the morphology of the cells
 abundance of the nutrients nitrogen and
phosphorous in the water  the fixed cells are then stained with a crystal violet
o cyanobacteria are able to convert nitrogen gas dye for 30 to 40 seconds. Then rinse the slide using
(N2) from the air into ammonium ions (NH4 +) water to remove excess stain. at this point, all cell
in the soil or water (nitrogen fixation) appears purple under the microscope.
o Some cyanobacteria produce toxins
(cyanotoxins) that can cause disease and even
death in wildlife species and humans that  next, a solution of gram iodine is added and retained
consume contaminated water), such as: in the slide for about 1 minute. iodine combines
 neurotoxins (affect central nervous system) with crystal violet to form di-iodine complex, and
 hepatotoxins (liver) that decrease its solubility within the cell. at this
 cytotoxins (other types of cell) point, the cell still appear as purple.

The Domain Archaea  then, the cells are decolorized by washing with
 archaean prokaryotes were discovered in 1977 ethanol or acetone. this is the differential step.
 more closely related to eukaryotes than they are to gram positive bacteria retain the crystal violet. gram
bacteria; some possess genes otherwise found only negative bacteria do not. the ethanol or acetone
in eukaryotes should be added dropwise with the slide tilted at an
 contains 2 phyla, 8 classes, 12 orders, 21 families, angle until the drop coming off from the slide started
69 genera, and 217 species to become colorless. even gram-positive cells can
 vary widely in shape; some are cocci, bacilli, and lose the violet iodine complex during prolonged
others form long filament
excessive decolorization.
 excess ethanol is then washed off with water. when
viewed under the microscope, gram- positive cells
appear purple and gram-negative cells are colorless.

 finally, the rinsed cells are covered with the

counter stain safranin for 20 to 30 seconds. this
stains the gram negative bacteria red or pink.

 after rinsing with water, the slide is dried by a filter

paper. when viewed microscopically. the gram-
positive bacteria are purple to blue and the gram
negative bacteria are pink to red

Gram-Positive bacteria:
 was not decolorized during the decolorization
 The thick layer of peptidoglycan in the cell walls of
Gram-positive bacteria makes it difficult to remove
the crystal violet–iodine complex during the
decolorization step
 Peptidoglycan is a polymer consisting of sugars and
amino acids that forms a mesh-like layer outside the
plasma membrane of most bacteria, forming the cell
wall. ..

Gram-Negative
 thin layer of peptidoglycan in the cell walls of
Gram-negative bacteria makes it easier to remove
the crystal violet–iodine complex during
decolorization
 the decolorizer dissolves the lipid in the cell walls of
Gram-negative bacteria; this destroys the integrity of
the cell wall and makes it much easier to remove the
crystal violet–iodine complex