CLINICAL BACTERIOLOGY – RMT 2024

MOD 3 (LAB) – STRUCTURES & COMPOSITIONS OF MICROORGANISMS

Mr. Ken Cortes, RMT
02/11/22

STRUCTURES & COMPOSITION

OF MICROORGANISMS
Very relevant as they are associated with their VIRULENCE

- Ability of the microorganism to cause disease to a particular

like humans

STRUCTURES & COMPOSITION

Things or substances present within the organism which they utilize

for them to cause infection to human host

I. GRANULAR INCLUSION BODIES

Figure 1. Corynebacterium diphtheriae

• These circular structures you seen within the body of the

organism are actually the granules or the granular inclusion
bodies
• We can see that the organism has irregular shaped circles
within the body
• Granular inclusion bodies are actually accumulations of
METAPHOSPHATES and POLYPHOSPHATES. In fact, the
accumulation of metaphosphates and polyphosphates are the
composition of the granular inclusion bodies

Note:
If you are asked in the examination or the practical examination What is
the composition of the pointed bacterial structure? If the pointed bacterial
structure is a granular inclusion body, then you will answer
ACCUMULATION OF METAPHOSPHATES AND POLYPHOSPHATES.

Note:
This is not actually the non-sporing snapping diplobacilli. The VIRULENCE MECHANISM
representative organism of non-sporing snapping diplobacilli is
Mycobacterium tuberculosis, but this isn’t Mycobacterium tuberculosis.
This is another representative organism with granular inclusion bodies VIRULENCE

• Ability of the microorganism to cause infection or disease. It also means

what is the exact way to participate in the ability of the organism to
Granular inclusion bodies are:
cause infection or disease in any human host.
→ irregularly shaped circles or spheres present within the
1. IT ACTS AS ENERGY RESERVE FOR THE ORGANISM
body of the organism.
Note:
Note: You may answer plainly or directly in the practical examination for that
When we say irregularly shaped circles or spheres, it actually refers to matter – ENERGY RESERVE FOR THE ORGANISM
the granular term of the bacterial structure.

→ GRANULAR – irregular shaped circles In what particular way or mechanism does energy reserve mechanism of
→ INCLUSION – inside the body granular inclusion bodies will enable the bacterial structure to cause the
organism to produce infection or any human host?

→ The immune system of human beings are actually

composed of WBC’s and competent substances that fight
off infection whenever infection arises. WBC’s have
actually 5 types.

Clinical Bacteriology, CDU – BSMT2A

 1. NEUTROPHILS Note:
2. EOSINOPHILS Granular inclusion bodies participate as “energy reserves” for the
microorganism. They are also called “metachromatic granules” because
3. BASOPHILS it exhibits a special property called “metachromasia”.
4. MONOCYTES
5. LYMPHOCYTES
Metachromasia
→ From these 5, 3 types of these are actually phagocytes → A special property of granular inclusion bodies/granules where
namely NEUTROPHILS, EOSINOPHILS, & MONOCYTES it appears different colors from that of the dye used to stain it.
→ Phagocytes are actually WBC’s that engulfs or eats any
→ The main dye used to stain granular inclusion bodies is
foreign material that enters the body.
“methylene blue” (blue dye). When methylene blue is used to
→ Phagocytes are provoked by antigens so that these
dye the granular bodies, the granular inclusion bodies under
phagocytes will immediately chase the antigen and engulf the microscope will appear purple/violet. Thus, this property is
it or eat it just before that antigen or any foreign material called “metachromasia”.
to cause infection in the immune system to that particular
host or individual GRANULAR INCLUSION BODIES
→ Since microorganisms are not normally seen, when an ALSO KNOWN AS REP. ORGANISM MAIN DYE USED
“Cytoplasmic Corynebacterium Methylene blue (blue
organism gains entry, these microorganisms are granules” or “polar diphtheriae which is dye) which will
immediately detected as an antigen. bodies” the causative agent appear purple/ violet
for diphtheria. “metachromasia”
Remember:
And
• When an antigen enters the body, like microorganisms or leprae
bacteria, what happens is, these microorganisms will stimulate
the WBC’s or phagocytes, thus the phagocytes will try attack or
combat it by eating or engulfing it just before that
microorganism or bacteria can cause infection to the human
Corynebacterium diphtheriae
host.

→ The specific name of the metachromatic granules of

• In the case of microorganisms with granular inclusion bodies, Corynebacterium diphtheriae is
these particular microorganisms will actually survive that o “Volutin granules of Babes-Ernst” or
phagocytosis or the phagocytes’ combat mechanisms in a o “Babes-Ernst granules”
particular way.
→ Morphology of Corynebacterium diphtheriae: “Chinese letter
• In layman’s story, when someone wants to catch you because appearance”.
you are actually at fault for that particular someone. When that
Corynebacterium diptheriae
someone wants to catch you and hurt you because of the bad
SPECIFIC NAME MORPHOLOGY
thing you have done to someone, Of course, your initial Volutine granules of Chinese letter appearance
reaction is to run away from that someone. Babes-Ernst

Or
• So if you try to connect that, when the phagocytes are already
provoked and stimulated because the microorganism had Babes-Ernst granules

entered the body, the microorganisms initial response is to run

Note: When writing the representative microorganism in exams,
away from the phagocyte because the microorganism knows
make sure to follow the general rule of the nomenclature of
that the phagocyte will actually destroy the microorganism. So,
microorganisms.
when that microorganism from the phagocyte, eventually, the
first set of energy used up in the running away from the • The 1st letter of the genus name should be capitalized and
phagocyte will be depleted. But since this organism has the following letters are in lowercase form.
granular inclusion bodies which serves as ENERGY RESERVE, • The specie name is written all in small letters.
once the original energy is exhausted from the escape of the (e.g., Corynebacterium diphtheriae)
phagocytes’ combat action. What will happen is that the
organism will utilize the granular inclusion bodies and will *in Celo, if there is an identification question asking for a rep. organism, follow the
general rule of nomenclature (if ALL CAPS you would be wrong)
utilize the ENERGY RESERVE so that it can keep away from the
particular phagocyte. Thus, it can escape the phagocyte’s GRANULAR INCLUSION BODIES
combat action, stay in the immune system for a long time, and FUNCTION STAINS
Food reserve • Acid-fast stain
for that long time, eventually that particular microorganism will • Loefller’s methylene blue
cause infection or disease. (appearing as reddish blue)

Clinical Bacteriology, CDU – BSMT2A

 iv. So that if, the WBC’s cannot actually catch these microorganisms
II. FLAGELLA
because the flagella propels continuously so that the organism
can escape totally the combat, engulfing, destructive action of the
phagocytes, what will happen is
v. The antibodies, from the immune system, which serves as the 2nd
line of defense in fighting off infection, will now be produced
specific to the flagella of the organism because that flagellum is
the ANTIGEN of the organism.
vi. So, what the antibodies will do is, the antibodies will destroy the
antigen part, which is the flagella. So, once the flagella are
destroyed, this will yield the microorganism as NON-MOTILE.
Once it is non-motile, the phagocytes can successfully chase that
non-motile organism, eat, engulf, and eliminate it from the body
so that the patient will actually recover from the infection

FLAGELLA IS THIN STRUCTURE

→ Staining or demonstration of flagella needs a specific

→ Are thin-like appendages/structures extending from the body
of the organism.
→ Composed of a fibrous protein called “flagellin”.
VIRULENCE MECHANISMS:
1. MOTILITY OF THE ORGANISM (SIMPLY, MOTILITY)
2. ANTIGENIC PROPERTY
1. VIRULENCE MECHANISM: MOTILITY
In what exact mechanism does the flagella enable the
microorganism to cause infection in the human host?
i. Any bacteria with flagella are motile (can travel from one place to
another).
ii. If a certain microorganism enters the body, the immune system
immediately detects that as an antigen in which the WBC and
phagocytes will immediately engulf it before it causes an infection.
iii. However, if the microorganism has flagella, the flagella will propel
microorganism so that it can escape or run away from the
phagocytes’ combat action.
iv. Unlike with the granular bodies that act as energy reserves, the
microorganism with flagella can continue propelling away from
the phagocytes. In other words, the microorganism with flagella
can “totally” escape from the combat/destructive action of the
phagocytes.
v. This means that the microorganism can stay in the body for long
periods of time which causes an infection.
reagent called MORDANT
→ The primary stain in the solution will actually pile up. The
MORDANT will bridge the color of the primary stain in the
solution to pile up in the flagella in the microorganism.
So, from a thin structure, once the primary stain is already
piled up, it would become thicker. After the staining
process, the thicker flagella would now be visible under
the microscope
Note:
MORDANT = “Tannic acid”
you would only answer “Tannic acid” if you are asked for the
specific reagent necessary for the demonstration of the
flagella
but if you are asked what reagent is necessary for the demo of
flagella, you would answer MORDANT
FLAGELLA IS CLASSIFIED ACCORDING TO TWO DIFF. CRITERIA
1. NO OF FLAGELLA PRESENT IN ORGANISM
2. LOCATION OF THE FLAGELLA IN THE BODY OF THE
ORGANISM
The criteria followed by MESSEA’s classification classifying according
to different types is the two criteria mentioned abovehand

2. VIRULENCE MECHANISM: POSSESSES ANTIGENIC

PROPERTIES (equivalent to how the organism is destroyed in the body – not
connected to the virulence of the organism)

Note:
in the examination, answer antigenic properties ONLY
i. When an organism possessing flagella enters the body, the
immune system will immediately detect that microorganism as
antigen because it is foreign
ii. But, in the case of microorganisms or bacteria with flagella, as it
enters the body, the immune system will detect the flagella of
the microorganism as the antigen and not the entirety of the
organism’s body.
iii. In other words, what really provokes the WBC’s (phagocytes) of
the immune system as these types of microorganisms enters the
body is the FLAGELLA

Clinical Bacteriology, CDU – BSMT2A

Types of flagella according to its main name of it’s classification MESSEA Note:
Spores/ endospores are actually spores within the organism
MESSEA’S Classification: These Light-shaded color of the organism (light or violet colored)
Monotrichous bacteria having one flagella. as shown above

Lophotrichous several flagella are located on one side; Spores/ endospores are composed of CALCIUM DIPICOLINATE. It
tuft (bundle)
can also be spelled/ called as:
Amphitrichous bacteria having flagella on each side.
- CALCIUM DIPICHOLINATE
Peritrichous flagella extending at different point; - DIPICOLINIC ACID
Atrichous no flagella - DIPICHOLINIC ACID

Because of the calcium dipicolinate as its composition, the spores/

endospores will enable the microorganism to resist physical agents
Representative Organism of Flagella: of disinfection.

Both are peritrichous Both are monotrichous Note:

→ Salmonella typhi (causative
agent of typhoid fever)
→ Proteus mirabilis/ Proteus
vulgaris (C.A. of UTI)
→ Pseudomonas aeruginosa
→ Vibrio cholerae
The virulence mechanism of spores/ endospores is very long, thus for
exam purposes, let’s uniform the answer if you are tasked for the
virulence mechanism, you may answer = RESIST DISINFECTION
But the more complete answer is that IT ENABLES THE ORGANISM TO
RESIST THE PHYSICAL AGENTS OF DISINFECTION

BACTERIAL SPORES
FUNCTION STAINS
• Virulence Factor • Writz
• Resistance to infection • Gram
• Dorner
• Conklin
• Schieffer

SPORES CAN BE CLASSIFIED INTO:

1. LOCATION WITHIN THE BODY OF THE ORGANISM

Figure 2. Pseudomonas aeruginosa a. Central spore
i. Center of the organism
FLAGELLA
FUNCTION STAINS
• Motility • Liefson’s stain
• Specific antigenically • Gray’s stain

III. SPORES/ ENDOSPORES

Figure 3. Central spore in the center

b. Subterminal spore
i. Located in the near end

c. Terminal spore
i. Located at the end portion
ii. Morphology/ appearance
1. Lollipop shape
2. Tennis-racket
3. Drumstick appearance

Spores have the other term – Endospores is that spores are found
within the body of the organism

Clinical Bacteriology, CDU – BSMT2A

 Central Spore Subterminal Spore Terminal Spore Note:
Center of the Located in the near Located at the end slime layer and capsule similar to each other composed of
organism end portion polysaccharides differ only on the amount present or
- Oval spore at - Oval elongated - Lollipop shape surrounding the microorganism
the center spores near - Tennis-racket
the end - Drumstick
appearance
REPRESENTATIVE ORGANISM
SLIME LAYER
Bacillus subtilis and Bacillus subtilis Clostridium tetani
Clostridium (common contaminant in the (C.A. for tetanus) • Polysaccharides surrounding the microorganism/s
laboratory)
botulinum present in little or scanty amount

CAPSULE

• Polysaccharides (bigger and visible) surrounding the

microorganism/s present in abundant or copious
amount; form definite layer
• More apparent under the microscope
• Water soluble structures;
o Usually Seen as unstained halo surrounding the
microorganism’s body
o Because when you try to stain, initially will take up the
color of the stain, but once you submit the smear to
water, then the color will be washed out

SLIME LAYER OR CAPSULE

FUNCTION STAINS
• Virulence Factor • Anthony stain
• Anti-phagocytic effect • Hiss
On the 6-7 o’clock appearance, there is a terminal spore that looks • Maneval’s stain
like a cotton bud

VIRULENCE MECHANISM Note:

primary stained washed from the water making it seem
In what way spores or endospores enables the organism to cause infection? unstained.

1. RESIST DISINFECTION
TESTS DONE
No matter how the laboratory surfaces is disinfected the NEGROSIN detects capsule that produces semi-opaque
background that makes the capsule visible
microorganism in that surface can still survive. Therefore, if any
person comes in contact with that surface where the spores resisted
QUELLUNG capsule is mixed with an antiserum, after the ab-ag
the disinfection can harbor or possess the microorganisms. It then REACTION reaction, it will swell
enters the human system causing infection.

REPRESENTATIVE ORGANISMS
SLIME • Sarcina lutea
IV. SLIME LAYER & CAPSULE LAYER

CAPSULE • Klebsiella pneumoniae

• S. pneumoniae

1. SLIME LAYER VIRULENCE MECHANISM

1. ADHESION/ ADHERENCE MECHANISM/ ADHESION

MECHANISM/ ATTACHMENT/ ATTACHMENT
MECHANISM

• Composed of POLYSACCHARIDES surrounding the
microorganisms present in different amounts or
accumulate around the cell wall in varying amount.
o elaborate mucilaginous substances
1. Adherence Mechanism
When an organism enters the body, it is recognized by the immune
system as an antigen. Phagocytes (white blood cell) are provoked
and stimulated and attacks the antigen to destroy the
microorganism. In the case of microorganisms with slime layer, when
these microorganisms enters the body, since they have slime layer
they have the capability to attach/adhere to smooth surfaces of the
normal cells in the body. When the immune system stimulates and

Clinical Bacteriology, CDU – BSMT2A

gives signals to the phagocytes to attack the organism the Corynebacterium diptheriae
SPECIFIC NAME MORPHOLOGY
phagocyte can no longer recognize them because the
Volutine granules of Chinese letter appearance
microorganism with slime layer is already hidden or has already fit Babes-Ernst
in within the smooth surfaces of the normal cells in the body. Since
Or
these are surfaces of the normal cells of the body, it means that the
phagocytes cannot recognize them anymore since they will think Babes-Ernst granules
that they are already normal cells in the body. When the
microorganism survived from the combat action of the phagocyte
and thrives in the system of the patient causing infection VIRULENCE MECHANSIM
ENERGY RESERVE FOR THE ORGANISM
2. CAPSULE VIRULENCE MECHANISM
2. FLAGELLA
1. PREVENTS PHAGOCYTOSIS/ ANTI-PHAGOCYTOSIS/
FLAGELLA
ANTI-PHAGOCYTIC EFFECT FUNCTION STAINS
2. ANTI-COMPLEMENTARY • Motility • Liefson’s stain
• Specific antigenically • Gray’s stain

COMPLEMENT highest form of defense against any invading

SYSTEM antigen. REPRESENTATIVE ORGANISM
Both are peritrichous Both are monotrichous
CELL-LYSIS end action of the complement system when → Salmonella typhi (causative → Pseudomonas aeruginosa
OR CELL the immune system is activated. agent of typhoid fever)
DEATH
→ Proteus mirabilis/ Proteus → Vibrio cholerae
vulgaris (C.A. of UTI)
1. ANTI-PHAGOCYTIC EFFECT

When the organism with capsule enters the body the immune
STAINING USED
system sends phagocyte to destroy the invading organism. Since MORDANT = “Tannic acid”
capsules are slippery structures or slimy structures when the
you would only answer “Tannic acid” if you are asked for the
phagocyte tries to open its mouth ready to eat the microorganism specific reagent necessary for the demonstration of the
it slips from the phagocyte. Because of the capsule’s slippery nature flagella
causing the microorganism to slip from the phagocytes’ mouth and
from there prevention of phagocytosis happens. but if you are asked what reagent is necessary for the demo of
flagella, you would answer MORDANT

2. ANTI-COMPLEMENTARY
VIRULENCE MECHANSIM
When the complement system is activated because the organism
MOTILITY
has provoked its action, the complement system will attack the ANTIGENIC PROPERTIES
organism and destroys the organism. Because of the capsule and
anti-complementary action when this type of organisms enters the
3. SPORES/ ENDOSPORES
body complementary system will not stimulate because the capsule
makes the complement system naive to its presence. BACTERIAL SPORES
FUNCTION STAINS
• Virulence Factor • Writz
• Resistance to infection • Gram
• Dorner
SUMMARY • Conklin
• Schieffer

1. GRANULAR INCLUSION BODIES

Central Spore Subterminal Spore Terminal Spore
GRANULAR INCLUSION BODIES Center of the Located in the near Located at the end
FUNCTION STAINS organism end portion
Food reserve • Acid-fast stain - Oval spore at - Oval elongated - Lollipop shape
• Loefller’s methylene blue the center spores near - Tennis-racket
(appearing as reddish blue) the end - Drumstick
appearance
REPRESENTATIVE ORGANISM
GRANULAR INCLUSION BODIES Bacillus subtilis and Bacillus subtilis Clostridium tetani
ALSO KNOWN AS REP. ORGANISM MAIN DYE USED Clostridium (common contaminant in the (C.A. for tetanus)
laboratory)
“Cytoplasmic Corynebacterium Methylene blue (blue botulinum
granules” or “polar diphtheriae which is dye) which will
bodies” the causative agent appear purple/ violet
for diphtheria. “metachromasia”

Clinical Bacteriology, CDU – BSMT2A

 Note:
The virulence mechanism of spores/ endospores is very long, thus for
exam purposes, let’s uniform the answer if you are tasked for the
virulence mechanism, you may answer = RESIST DISINFECTION

But the more complete answer is that IT ENABLES THE ORGANISM TO

RESIST THE PHYSICAL AGENTS OF DISINFECTION

4. SLIME LAYER

SLIME LYER OR CAPSULE

FUNCTION STAINS
• Virulence Factor • Anthony stain
• Anti-phagocytic effect • Hiss
• Maneval’s stain

Note:
primary stained washed from the water making it seem unstained.

TESTS DONE
NEGROSIN detects capsule that produces semi-opaque
background that makes the capsule visible

QUELLUNG capsule is mixed with an antiserum, after the ab-ag

REACTION reaction, it will swell

REPRESENTATIVE ORGANISMS
SLIME • Sarcina lutea
LAYER

CAPSULE • Klebsiella pneumoniae

• S. pneumoniae

SLIME LAYER VIRULENCE MECHANSIM

ADHESION/ ADHERENCE MECHANISM/ ADHESION MECHANISM
ATTACHMENT/ ATTACHMENT MECHANISM

CAPSULE LAYER VIRULENCE MECHANSIM

PREVENTS PHAGOCYTOSIS/ ANTI-PHAGOCYTOSIS/ ANTI-PHAGOCYTIC
EFFECT
ANTI-COMPLEMENTARY

Clinical Bacteriology, CDU – BSMT2A

Clinical Bacteriology, CDU – BSMT2A