7.2 Bacteriology

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OUR LADY OF FATIMA UNIVERSITY

COLLEGE OF PHARMACY Overview
Gram-positive cocci Aerobic Gram-positive bacilli
Staphylococcus aureus Bacillus anthracis
Streptococcus pyogenes Bacillus cereus
Streptococcus agalactiae Anaerobic Gram-positive bacilli
BACTERIOLOGY Streptococcus pneumoniae
Clostridium perfringens
Enterococcus faecalis
Clostridium difficile
PHARMACEUTICAL MICROBIOLOGY Gram-negative cocci Clostridium botulinum
AND PARASITOLOGY Neisseria gonorrheae Clostridium tetani
Neisseria meningitidis
PHMP 211
Aerobic Acid-fast bacilli
Mycobacterium tuberculosis
Mycobacterium leprae
OUR LADY OF FATIMA UNIVERSITY – COLLEGE OF PHARMACY – PHMP211
Staphylococcus aureus Staphylococcus aureus

• perfectly spherical cells about 1 • Staphylococcus aureus
micrometer in diameter. forms a fairly large yellow
• grow in clusters colony on rich medium.
• catalase-positive and oxidase-negative • S. aureus is often
• can grow at a temperature range of 15 hemolytic on blood agar.
to 45 degrees and at NaCl • Staphylococci are
concentrations as high as 15 percent. facultative anaerobes
• Nearly all strains of S. aureus produce
the enzyme coagulase.
– Only S. aureus is coagulase-positive
among the Staphylococcus.
OUR LADY OF FATIMA UNIVERSITY – COLLEGE OF PHARMACY – PHMP211 OUR LADY OF FATIMA UNIVERSITY – COLLEGE OF PHARMACY – PHMP211

expresses many potential virulence factors: expresses many potential virulence factors:
1. Fibrin/fibrinogen binding protein promotes attachment 7. Toxic Shock Syndrome Toxin-1 (TSST-1) is expressed
to blood clots and traumatized tissue adhesin.
systemically and is the cause of toxic shock syndrome
2. Coagulase converts fibrinogen to fibrin and coagulates
plasma. (TSS).
3. Staphylokinase lyses fibrin Signs and Symptoms:
4. Protein A is a surface protein of S. aureus, which binds IgG – Body temperature > 38.9 °C (102.02 °F)
molecules by their Fc region.
5. Leukocidin is a toxin that specifically acts on – Systolic blood pressure < 90 mmHg
polymorphonuclear leukocytes. – Diffuse rash, intense erythroderma, blanching with
6. Enterotoxins cause diarrhea and vomiting when ingested. subsequent desquamation, especially of the palms and
soles
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expresses many potential virulence factors: Laboratory Diagnosis
8. Exfoliatin toxin (ET) causes the scalded skin syndrome in • Culture
neonates, which results in widespread blistering and loss Treatment:
of the epidermis. • Beta-lactam antibiotics or cephalosporins.
Staphylococcus Staphylococcus
saprophyticus Staphylococcus
epidermidis
• Novobiocin sensitive • Non-hemolytic Organism Colony Catalase Coagulase Novobiocin Hemolysis
• Has the ability to adhere to • Coagulase negative appearance production production sensitivity on BAP
artificial materials in the • Novobiocin resistant S. aureus Golden + + Sensitive +
body (e.g. catheters and S. epidermidis White-gray + - Sensitive -
prosthetic heart valves) • Causes “honeymoon
cystitis” S. saprophyticus White-gray + - resistant -
• Frequently isolated:
infected indwelling
catheters, prostetic heart
valve endocarditis
Rebecca Lancefield Classification Lancefield’s Classification

• Group A - Streptococcus pyogenes
• Group B - Streptococcus agalactiae • A serologic classification of streptococci based on their
• Group C - Streptococcus equisimilis, Streptococcus zooepidemicus antigenic characteristics.
• Group D - Enterococci, Streptococcus bovis • The bacteria are divided into 13 groups by the
• Group E - Streptococcus milleri and mutans identification of their pathologic action.
• Group F - Streptococcus anginosus
• Group G - Streptococcus canis • Group A contains most of the streptococci that cause
• Group H - Streptococcus sanguis infection in humans.
• Group L - Streptococcus dysgalactiae • Groups B to O are less pathogenic and are often present
• Group N - Lactococcus lactis without causing disease.
• Group R&S - Streptococcus suis • Most are hemolytic; of those, the beta subgroup is the
• other Streptococcus species are classified as non-Lancefield most likely to be the cause of infection.
Streptococci
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Streptococcus pyogenes Streptococcus pyogenes

Streptococcus pyogenes is Virulence Factors
the main human pathogen 1. Streptokinase (fibrinolysin) transforms the
associated with local or
plasminogen of human plasma to plasmin.
systemic invasion and
poststreptococcal 2. Hyaluronidase aids in spreading infecting
immunologic disorders. microorganisms.
Individual cocci are spherical 3. Pyrogenic exotoxins A-C cause the rash that
or ovoid and are arranged occurs in scarlet fever.
in chains. 4. Streptolysin O is an oxygen-labile protein
Classification responsible for the area of complete hemolysis.
Streptococcus pyogenes
belongs to Lancefield group A
Streptococcus agalactiae
• Normal flora in Laboratory Diagnosis:
nasopharynx, oral cavity, • Culture: Streptococcus
intestinal tract, and vagina agalactiae is -hemolytic
Microscopic appearance on blood agar.
• Gram-positive cocci in Treatment
chains. • Ampicillin is the drug of
Virulence Factor choice.
• Capsular material
interferes with phagocytic
activity and complement
cascade activation
Streptococcus agalactiae Streptococcus pneumoniae

• Streptococcus pneumoniae are gram-positive diplococci.
• Pneumococci are often lysed by surface active agents such
as bile salts.
• They are normal inhabitants of the upper respiratory tract
of humans.
Classification
• Αlpha-hemolytic streptococcus
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Streptococcus pneumoniae Streptococcus pneumoniae

Microscopic appearance
• distinctive in that the cocci Treatment:
are encapsulated ovoid or 1. Penicillin has been the drug of choice .
lancet –shaped and are 2. Third generation cephalosporins are gaining acceptance
often seen in pairs on as the initial treatment for severe pneumococcal
gram-stained samples. infections.
Virulence Factors 3. Vancomycin is the drug of choice for highly resistant
1. Polysaccharide capsule strains of S. pneumoniae.
has anti-phagocytic
property.
2. IgA protease inactivates
IgA antibodies.
Optochin Test Quellung Reaction

S. pneumoniae is optochin- • Pneumococci + anti-serum Enterococcus faecalis
sensitive and methylene blue =
Viridans Streptococci is swollen capsule • normal flora in the intestinal tract as well as in the oral
optochin-resistant • (Viridans = negative) cavity.
Classification
• Group D
Virulence Factors
• Adhesions, cytolysins allow these to proliferate as
nosocomial pathogens.
Enterococcus faecalis Enterococcus faecalis

• Many infections may show a synergistic response when
treated with penicillin and an aminoglycoside, such as
gentamicin or streptomycin.
• Other antibiotics useful in patients who are allergic to
penicillin are vancomycin and erythromycin.
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Neisseria Neisseria gonorrhoeae

General Properties Microscopic appearance
• Neisseria gonorrh(o)eae ferments only glucose • gram-negative, non-motile
diplococcus, approximately
and differ antigenically from the other Neisseriae. 0.8 um in diameter.
• Gonococci usually produce smaller colonies than • Individual cocci are kidney-
those of the other Neisseria. shaped; the organisms
occur in pairs, the flat or
concave sides are
adjacent.
Neisseria gonorrhoeae Neisseria meningitidis

Virulence Factors: • Neisseria meningitidis resides in its natural habitat
1. Pili are involved in bacterial adherence to host cells. within the nasopharyngeal tract of humans
2. Outer Membrane Proteins are the primary virulence • 5-15% of the human population carries the bacteria in
factors of N. gonorrheae which mediate attachment to its nonpathogenic form.
mucosal cells. Microscopic appearance:
3. Capsules allow intracellular survival. • Neisseria meningitidis is gram-negative cocci, ranging
4. IgA protease may help gonococcus evade secretory IgA on from 0.6-1.0 um in diameter.
mucosal surfaces. • The cocci are kidney bean shaped often seen as
diplococci.
• It is also capsulated and piliated.
Neisseria meningitidis Neisseria meningitidis

Virulence Factors: Laboratory Diagnosis:
1. Pili serve as adhesion factors for the oropharyngeal • Culture
mucosa and probably mediate attachment to the • Rapid diagnostic tests is based on the agglutination of latex
meningeal tissues as well. particles coated with antibodies to N. meningitidis
2. Capsular polysaccharide has antiphagocytic properties. Treatment
3. IgA proteases cleave IgA and protect bacteria against the • Penicillin G is effective when large doses are administered
effects of secretory IgA. intravenously.
4. Lipopolysaccharide (LPS, endotoxin) is ten times more • Ceftriaxone and other 3rd generation cephalosporins
potent than most other endotoxins.
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Aerobic Acid Fast Bacilli:

Mycobacterium
General Characteristics: • Obligate aerobes growing most successfully in tissues with
• Organisms belonging to the genus Mycobacterium are very a high oxygen content, such as the lungs.
thin, rod-shaped, and non- motile. • Facultative intracellular pathogens usually infecting
• They have an unusual cell wall structure that contains mononuclear phagocytes (e.g. macrophages).
mycolic acid which make them difficult to stain using Gram • Hydrophobic with a high lipid content in the cell wall.
stain.
• These organisms resist decolorization by acidified alcohol
after prolonged application of a basic fuchsin dye following
its application, hence they are called acid-fast organisms.
Microscopic appearance:
• Mycobacterium tuberculosis are gram-positive (no outer cell
membrane), non-motile pleomorphic rods, related to the
Actinomyces.
Virulence Factors:
1. Cording factor is a glycolipid derivative of mycolic acid that is
present in the outer surface of M. tuberculosis.
2. Sulfatides, glycolipids located on the surface of mycobacteria,
inhibit phagolysosome formation.
OUR LADY OF FATIMA UNIVERSITY – COLLEGE OF PHARMACY – PHMP211
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Laboratory Diagnosis
a. Positive tuberculin skin test, an immune reaction to a small
quantity of tuberculosis antigens.
b. X-rays of the chest and microscopic examination of sputum
confirm the skin test.
c. Detection of significant numbers of acid-fast bacilli (using
the Ziehl-Neelsen or Kinyoun stain) in sputum or tissue
samples is considered a positive diagnosis.
Mycobacterium tuberculosis Mycobacterium leprae

Treatment: • Mycobacterium leprae is • Characteristic lesions are
Antitubercular drugs: the causative agent of the grown in laboratory
• Rifampicin, Isoniazid, Pyrazinamide, Ethambutol, disease, leprosy. animals.
Streptomycin • In size and shape, it closely • Incubation period is
resembles M. tuberculosis. months to years.
Regimen
• Not grown in non-living • Route of infection is
– The treatment of tuberculosis involves simultaneous
bacteriologic media. through nasal mucus
administration of at least two drugs to avoid resistant
secretion.
strains of M. tuberculosis.
Duration
• Treatment should be continued for several months,
depending on the severity of the disease.
Mycobacterium leprae Mycobacterium leprae

• Disease: Hansen’s disease
or leprosy.
• The lesion involves the
cooler parts of the body,
Eg. Ear lobes. Clinical
triads: Anaesthetic skin
patches
• Dx: Lepromin skin test
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Mycobacterium leprae Aerobic Gram Positive Bacilli: Bacillus

Laboratory Diagnosis: General Characteristics:
• Diagnosis is based primarily on clinical signs and symptoms. • The members of the family Bacillaceae are gram-positive
• Scrapings or biopsies from lesions reveal characteristic rod-shaped bacteria, which form endospores.
histopathology and, in lepromatous disease, large numbers
of acid fast.
Culture and isolation:
• M. leprae cannot be cultured in bacteriologic media.
• It requires cell culture system.
Treatment
• very easily cured by using Dapsone + Rifampin +/-
Clofazimine in combination.
Bacillus anthracis Bacillus anthracis

• rod-shaped, endospore forming aerobic or facultatively Laboratory Diagnosis:
anaerobic, gram-positive bacteria. • Diagnosis is confirmed by observation of characteristic
• Only one endospore is formed per cell - needs oxygen to encapsulated bacilli in polychrome methylene blue-stained
sporulate smears of blood, exudate, lymph, cerebrospinal fluid, etc.,
Virulence Factors: and/or by culture.
1. Capsule composed of a high molecular weight polypeptide Treatment:
of D-glutamic acid develops in infected tissue. • Penicillin is the drug of choice.
2. Exotoxin produced by B. anthracis is composed of 3
different proteins.
Bacillus cereus
General Properties: Virulence Factors:
• Bacillus cereus is gram- • Enterotoxin may be
positive preformed in the food or
• Facultatively aerobic produced in the Intestine.
spore-former whose cells
are large rods and whose
spores do not swell the
sporangium.
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Bacillus cereus Bacillus cereus

Laboratory Diagnosis:
• Isolation of B. cereus from suspect foods and determining
their enterotoxigenicity by serological (diarrheal toxin) or
biological (diarrheal and emetic) tests
Treatment:
• Food poisoning is usually a self-limiting situation that
requires supportive treatment only.
Anaerobic Gram Positive Bacilli:

Clostridium
General Characteristics: • an anaerobic, gram- Virulence Factor:
• The Clostridia are anaerobic, spore-forming bacilli that positive spore-forming rod. 1. Lecithinase is the primary
usually stain gram-positive. • It is widely distributed in toxin - hydrolyzes lecithin
• Most species are anaerobes. the environment and and sphingomyelin.
• The pathogenic species produce soluble toxins, some of frequently occurs in the 2. Alpha-toxin – responsible
which are extremely potent. intestines of humans and for gas gangrene
many domestic and feral 3. Beta-toxin – responsible
• The clostridia are widely distributed in nature and are animals.
present in soil and in the intestinal tract of humans and for food poisoning and
animals enteritis necroticans
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Clostridium perfringens Clostridium difficile

Laboratory Diagnosis : • Organism diminution of normal gut flora by the activity of
• Toxin detection in the feces of patients is used in the various antimicrobial agents to become established in the
diagnosis of illness gut of hospitalized patients.
• Culture • Once established, elaboration of toxins results in diarrhea
Treatment: or potentially life-threatening inflammation of the colon.
• Penicillins are the drugs of choice for C. perfringens Virulence Factors:
infections, but they are usually used in the association with • Cytotoxins A and B are heat and acid-labile proteins
broad-spectrum antibiotics because therapy is initiated attached to the intestinal epithelial cells at the microvilli
before the precise nature of infection is known. level.
Clostridium difficile Clostridium difficile

Laboratory Diagnosis:
• Culture and isolation is followed by the identification of the
secreted exotoxins is probably the most definitive method,
but it takes several days.
• Detection of toxins in fecal material using Enzyme
Immunoassay have been developed and appear to be
adequately specific and sensitive.
Clostridium difficile Clostridium difficile

Clinical Diseases: Treatment:
a. Antibiotic-associated • Discontinuing the offending antibiotic may be sufficient for
diarrhea is usually associated very mild cases.
imbalance of the normal • Oral metronidazole or vancomycin is use in mild to
intestinal flora and in most moderate cases.
cases will subside after
therapy is discontinued. • Intravenous metronidazole or oral vancomycin is
recommended.
b. Pseudomembranous
(necrotizing) colitis develops
as a consequence of the
release of cytotoxins by non-
invasive c. difficile in the
colon.
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Clostridium botulinum Clostridium botulinum

• Clostridium botulinum is an anaerobic, gram-positive
spore-forming rod that produces a potent neurotoxin.
• The spores are heat-resistant and can survive in foods that
are incorrectly or minimally processed.
• Seven types (A, B, C, D, E, F and G) of botulism are
recognized, based on the antigenic specificity of the toxin
produced by each strain.
– Types A, B, E and F cause human botulism. Types C and
D cause most cases of botulism in animals.
Clostridium botulinum
Virulence Factors:
• Toxins A to F are neurotoxins that interfere with
neurotransmission at the peripheral cholinergic synapses
by preventing the release of acetylcholine causing flaccid
paralysis.
Clinical Diseases:
a. Foodborne botulism is the name of the disease (actually a
foodborne intoxication) caused by the consumption of
foods containing the neurotoxin produced by C. botulinum.
b. Infant botulism - caused by the ingestion of C. botulinum
spores, which colonize and produce toxin in the intestinal
tract of infants (intestinal toxemia botulism).
Clostridium botulinum Clostridium tetani

Treatment: • obligate anaerobic bacillus
• Supportive measures are most important. with terminal spores that
• Ensuring proper pulmonary ventilation, using assisted stains gram positive in
respiration if necessary, is critical. fresh cultures, but may
stain gram negative in
older cultures.
Virulence Factor
• Tetanospasmin is a
neurotoxic that disrupts
nerve impulses to muscles.
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Clostridium tetani Clostridium tetani

Clinical Disease: Treatment:
• Tetanus is characterized by twitching of muscles around a • C tetani infection is treated with antimicrobial agents
wound, pain in neck and jaw muscles (trismus; lockjaw), (metronidazole or penicillin) and by local wound
and around the wound. debridement.
Laboratory Diagnosis: • Other measures include tetanus immunoglobulin and
• Diagnosis is primarily by the clinical symptoms. supportive therapy.
• The wound may not be obvious. • Tetanus toxoid
• C. tetani is recovered from only one-third of all implicated
wounds.
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7.2 Bacteriology

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7.2 Bacteriology

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24/10/2021

OUR LADY OF FATIMA UNIVERSITY

OUR LADY OF FATIMA UNIVERSITY – COLLEGE OF PHARMACY – PHMP211

Staphylococcus aureus Staphylococcus aureus

Staphylococcus aureus Staphylococcus aureus

Staphylococcus aureus Staphylococcus aureus

Rebecca Lancefield Classification Lancefield’s Classification

Streptococcus pyogenes Streptococcus pyogenes

Streptococcus agalactiae Streptococcus pneumoniae

Streptococcus pneumoniae Streptococcus pneumoniae

Optochin Test Quellung Reaction

Enterococcus faecalis Enterococcus faecalis

Neisseria Neisseria gonorrhoeae

Neisseria gonorrhoeae Neisseria meningitidis

Neisseria meningitidis Neisseria meningitidis

Aerobic Acid Fast Bacilli:

OUR LADY OF FATIMA UNIVERSITY – COLLEGE OF PHARMACY – PHMP211

Mycobacterium tuberculosis Mycobacterium leprae

Mycobacterium leprae Mycobacterium leprae

Mycobacterium leprae Aerobic Gram Positive Bacilli: Bacillus

Bacillus anthracis Bacillus anthracis

Bacillus cereus Bacillus cereus

Anaerobic Gram Positive Bacilli:

Clostridium perfringens Clostridium difficile

Clostridium difficile Clostridium difficile

Clostridium difficile Clostridium difficile

Clostridium botulinum Clostridium botulinum

Clostridium botulinum Clostridium tetani

Clostridium tetani Clostridium tetani

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