MLS 13: BACTERIOLOGY (LEC) MODULE PROF: CAPATAYAN, YM.

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MORPHOLOGY
MODULE 11: Pseudomonas, Vibrio
Haemophilus
Slender, rod shape (bacillus) a bacterium
Shape
I. Introduction
II. Staphylococcus
III. Micrococci about 1.5–3 mm × 0.5 mm (micrometer)
size
IV. Streptococci

Arrangement singly or in pairs

Of Cells
PSEUDOMONAS

PSUEDOMONAS AERUGINOSA
Motility actively motile bacterium
• Gram-negative, aerobic bacilli
• Most common non-fermentative bacilli; most common clinical isolate
• Non-fermentative, non-oxidative flagellated bacterium with a polar flagellum,
• Most common cause of infections of burn injuries, eye (in contact Flagella arranged in an amphitrichous manner.
lenses) infections and “swimmer’s ear” which is otitis externa
• Can be isolated in disinfectants (Iodine), causing nosocomial infections
• In rare circumstances, cause community-acquired pneumonias, as well non – sporing bacterium
Spores
as ventilator-associated pneumonias.
• Member of the Pseudomonas fluorescent group along with P.
fluorescens and P. putida non – capsulated bacterium but some strains
Capsule posses a slime layer
Fluorescent group Non-fluorescent Burkholderia
of Pseudomonas group of group
Pseudomonas Gram
Gram -ve (Negative) bacterium
P. aeruginosa P. stutzeri B. cepacian Staining
P. fluorescens P. mendocina B. mallei Reaction
P. putida B. pseudomallei

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 CULTURE REQUIREMENTS Beta
hemolysis
• Optimum temperature- only ones capable of growth in 35-degree C. Hemolysis -- --- ---
(in some
and 42-degree C. strains)
• Oxygen requirements- obligate anaerobe
• Cetrimide Agar medium is a selective medium for Pseudomonas spp.
Consisting of Cetrimide which inhibits the growth of many bacteria
while allowing the growth of Pseudomonas aeruginosa.

Cultural Nutrient Cetrimide MacConkey Blood Agar

Characteristics Agar Agar Agar Medium
Medium medium medium
(NAM)
Shape Irregularly
Irregular Circular Circular edged BIOCHEMICAL TEST
circular
Size 2-4mm 1-3mm 2-3mm 2-4mm • Strict aerobic requirement; Oxidase (+)
Elevation Low • Motile via polar flagellum
Low convex Low convex Flat
convex • Non-lactose fermenter
Surface Smooth Smooth Smooth Smooth • Pigment production: only P. aeruginosa produces the blue pigment
(fresh (fresh (fresh (fresh “pyocyanin” which mixes with fluorescein (pyoverdin) producing the
isolation): isolation): isolation): isolation): distinguishable blue-green color
Mucoid Mucoid Mucoid Mucoid
(When slime (When (When (When slime
layer is slime layer slime layer layer is NAME OF THE PIGMENT COLOR OF THE PIGMENT
formed) is formed) is formed) formed Pyocyanin Bluish-Green
Color Greenish Greenish Fluorescein Greenish-yellow
blue (due to blue (due Greyish Pyorubicin Reddish-brown
Colorless Pyomelanin Brown to black
pigment to pigment white
production) production)
Structure Translucent- Translucent-
Opaque Transparent
Opaque Opaque

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 MLS 13: BACTERIOLOGY (LEC) MODULE PROF: CAPATAYAN, YM.
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VIBRIO VIBRIO CHOLERAE

• Rigid, curved rods or comma shaped • Causes “rice eatery stool”

• Highly motile-single polar flagellum, non sporulated & non- • Produces choleragen toxin; other virulent factors include pili and
encapsulated mucinase
• Present in marine environments and surface water with cosmopolitan • May be transmitted via ingestion of undercooked seafood or
in distribution contaminated drinking water
• All species are halophilic (salt-loving) except V. cholerae and V. • V. cholerae 01 has two biotypes: classical and El Tor
mimicus
• Thiosulfate citrate bile salt sucrose agar (TCBS) is a selective and Classical El Tor
differential medium RBC Hemolysis Negative Positive
SUCROSE POSITIVE SUCROSE VP Negative Positive
NEGATIVE Polymyxin B (50U) Susceptible Resistant
V. cholerae V. parahaemolyticus Agglutination of Negative Positive
V. alginolyticus V. vulnificus chicken RBCs
V. fluvialis V. mimicus
V. furnissil • V. cholerae 01 is subdivided into (3) serotypes: Inaba, Ogawa,
Hikojima

MORPHOLOGY
• Gram negative, Rigid, short curved rods
that are actively motile
• Comma shaped, sheathed, polar
flagellum having size of about 1.5 × 0.2-
0.4 μm in size.
• facultative anaerobe, non-sporulated,
nonencapsulated.
• Growth is stimulated by sodium chloride
• pH 6.4 – 9.6 (optimum 8.2).
• Acid labile
• Temperature 16 – 40°C (Optimum
37°C).
• “darting motility”/ “swarm of gnarts”

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CULTURE

• facultative anaerobic organism.

• grows within a temperature range of 16- 40°C (optimum 37°C).
• Growth is better in an alkaline medium
• the range of pH being 6.4 – 9.6 (optimum 8.2).
• It grows well on ordinary media.
• Selective Medium – TCBS in which V. cholerae grows well on
Thiosulphate citrate bile sucrose (TCBS) agar, on which it produces
yellow colonies due to sucrose fermentation that are readily visible
against the dark green background of the agar.

Vibrio Species Associated with Human Disease

Species Source of Clinical
Infection Disease
Seawater Wound infection,
BIOCHEMICAL REACTIONS V.alginolyticus
external otitis
• fermentative, producing acid, but no gas. V.cholerae Water, food Gastroenteritis
• ferment glucose, mannitol, maltose, mannose and sucrose BUT NOT V.cincinnatiensis Unknown Bacteremia,
inositol, arabinose or lactose. meningitis
• Indole is formed - nitrates are reduced V.fluvialis Seafood Gastroenteritis,
to nitrites. wound infection,
• These properties contribute to the bacteremia
“cholera red reaction‟. V.furnissil Seawater Gastroenteritis
• Catalase positive. V.harveyi Seawater Wound infection
• Oxidase positive. (shark bite)
• Methyl red negative V.etchinikovii Unknown Bacteremia
• Urease negative V.mimicus Fresh water Gastroenteritis,
• “string” test can be used to identify V.cholera colonies wound infection,
bacteremia
• A loopful of the growth is mixed with a drop of 0.5% sodium
desoxycholate on a slide V.parahaemolyticus Shellfish, Gastroenteritis,
seawater wound infection,
• If the mixture looses its turbidity, becomes mucoid and forms a string
bacteremia
when the loop is drawn away slowly, the test is positive.

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 MLS 13: BACTERIOLOGY (LEC) MODULE PROF: CAPATAYAN, YM.
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V.vulnificus Shellfish, Bacteremia, • The standard medium used for growth of H. influenzae is a chocolate
seawater wound infection, agar plate (CAP), which can be prepared with heat-lysed horse blood,
cellulitis a good source of both hemin and NAD, although sheep blood can also
be used.
Key Biochemical & Physiological Characteristics of Important
Vibrios
v. cholerae V. V. vulnificus CULTURE
parahaemolyticus • CAP-Large, round, smooth, convex, colorless to grey, opaque colonies
Oxidase + + + • Encapsulated strains appear more mucoidal than non-encapsulated
D- glucose - - - strains, which appear as smaller, compact grey colonies.
(Gas)
• H. influenzae produce a pungent indole smell
Lactose - - V
• BAP-transluscent, low, convex, or flat pinpoint colonies, shoes
Sucrose + - V satellitism with S. aureus.
Myo-inositol - - -
Lysine + + +
decarboxylase
Arginine - _ -
dihydrolase
Omithine + + V
decarboxylase
Growth in 0% + _ -
NaCl
Growth in 6% V _ V
NaCl
TCBS growth Good Good Good
Colony color Yellow Green Green
on TCBS

HAEMOPHILUS

• Mucous membrane of the URT Requirement Requirement B-hemolysis

• Small, pleomorphic, gram-negative bacilli or coccobacilli with random for hemin (X for NAD (V on horse
arrangements factor) factor) blood agar
• Fastidious organisms which gros best at 35-37 degree C with ~5% CO2 H. influenzae + + -
(or in a candle jar) and requires hemin (X factor) and nicotinamide- H. parainfluenzae - + -
adenine-dinucleotide (NAD, also known as V factor) for growth H. haemolyticus + + +

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 H. - + + BIOCHEMICAL CHARACTERISTICS
parahaemolyticus
H. aphrophilus + - - • Glucose +
H. paraphrophilus - + - • Xylose +
• Lactose +
• Sucrose –
PATHOGENICITY • Mannitol –
• Nitrites reduced
• Type b Haemophilus influenzae can cause meningitis, epiglottis,
• Indole differs on
bacteremia, and cellulitis.
• B type causes meningitis
• Non-typable H influenzae can cause otitis media, sinusitis,
tracheobronchitis, and pneumonia
References

• Ma’am Yvonne Video Discussion

• USA Bacteriology Module Packet

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BADOLES ● BESA ● BUENVENIDA ● BUNCALAN ● DELA PENA ● JANEA ● MERISCO | MLS 3D (AY 2021-2022) 7