Haemophilus influenzae

Encapsulated strain of Haemophilus influenzae on chocolate agar

plate. Encapsulated strains may produce larger colonies with a glistening mucoid
quality. Haemophilus influenzae requires two accessory growth factors: factor
X (haemin or other porphyrins) and V factor (NAD). The X and V factor
requirement is usually demonstrated by the absence of growth on porphyrin and
NAD deficient but otherwise nutritionally adequate media except near paper disc
impregnated with X and V factors. H.influenzae grown on agar plates usually shows
a highly regular shape under microscope. The rounded ends of short bacilli make
many appear round, hence the term coccobacilli.

Haemophilus influenzae, formerly called Pfeiffer's bacillus, is a non-motile Gramnegative pleomorphic rod-shaped bacterium first described in 1892 by Richard
Pfeiffer during an influenza pandemic. It is facultatively anaerobic. H.
influenzae was mistakenly considered to be the cause of influenza until 1933, when
the viral etiology of the flu became apparent. Still, H. influenzae is responsible for
a wide range of clinical diseases.
Diseases
Most strains of H. influenzae are opportunistic pathogens - that is, they usually
live in their host without causing disease, but cause problems only when other
factors (such as a viral infection or reduced immune function) create an
opportunity. Naturally-acquired disease caused by H. influenzae seems to occur in
humans only. In infants and young children, H. influenzae type b (Hib) causes
bacteremia, pneumonia, and acute bacterial meningitis. Occasionally, it
causes cellulitis, osteomyelitis, epiglottitis, and infectious arthritis. Due to routine
use of the Hib conjugate vaccine in the U.S. since 1990, the incidence of invasive
Hib disease has decreased to 1.3/100,000 in children. However, Hib remains a
major cause of lower respiratory tract infections in infants and children in

developing countries where vaccine is not widely used. Unencapsulated H.

influenzae causes ear (otitis media) and eye(conjunctivitis) infections and sinusitis
in children, and is associated with pneumonia.

Culture
Bacterial culture of H. influenzae is performed on Chocolate agar at 37C in an
enriched CO2 incubator. Blood agar growth is only achieved as a satellite
phenomenon around other bacteria. H. influenzae will grow in the hemolytic zone
of Staphylococcus aureuson blood agar plates. The hemolysis of cells by S.
aureus releases nutrients vital to the growth of H. influenzae. H. influenzae will not
grow outside the hemolytic zone of S. aureus due to the lack of nutrients in these
areas.
Colonies of H. influenzae appear as raised or convex, smooth, pale, grey
or transparent. Gram-stained and microscopic observation of a specimen of H.
influenzae will show Gram-negative coccobacilli with no specific arrangement.
Further serological is necessary to distinguish the capsular polysaccharide and
differentiate between H. influenzae b and non-encapsulated species. H.
influenzae is a finicky bacterium to culture, and any modification of culture
procedures can greatly reduce isolation rates. Poor quality of laboratories in
developing countries has resulted in poor isolation rates of H. influenzae.

Haemophilus influenzae basic characteristics

GRAM-NEGATIVE RODS OR COCCOBACILLI
NONMOTILE
NON-SPORE-FORMING
CATALASE: POSITIVE
OXIDASE: POSITIVE
FACULTATIVELY ANAEROBIC
Identification of Haemophilus influenzae
requirement of X+V factor in cultivation media; X = hematin, V =
nicotinamide adenine dinucleotide(NAD)
growth on chocolate agar but not on blood agar (when cultivated in pure
culture)
satelite phenomenon with Staphylococcus aureus (on blood agar)
Antibiotic treatment of Haemophilus influenzae infections
Should be always guided by in vitro susceptibility tests!!
Selection of appropriate antibiotics depends on diagnosis!!

IF SUSCEPTIBLE:
ampicillin
amoxicillin
ampicillin/sulbactam
amoxicillin/clavulanate
cefuroxime
ceftriaxone

cefotaxime
ALTERNATIVES:
Trimethoprim-Sulfomethoxazole(TMP-SMX)
azithromycin
clarithromycin
doxycycline

Staphylococcus aureus

Staphylococcus aureus on Columbia agar with 5% defibrinated sheep blood (BioRad). Individual colonies on agar are round,convex, and 1-4 mm in diameter with a
sharp border. On blood agar plates, colonies of Staphylococcus aureus are
frequently surrounded by zones of clear beta-hemolysis. The golden
appearance of colonies of some strains is the etymological root of the bacteria's
name; aureus meaning "golden" in Latin.
Methicillin-resistant strains of Staphylococcus aureus (i.e. MRSA) often have
only weak or no beta-hemolysis and special cultivation media with oxacillin, mannitol
and NaCl for their isolation are used. MRSA is able to grow on this media and
produce colonies of certain color, depending on used pH indicator (in this picture
pink).
Staphylococcus aureus may occur as a commensal on human skin; it also occurs in
the nose frequently (in about a third of the population) and throat less
commonly. The occurrence of S. aureus under these circumstances does not always
indicate infection and therefore does not always require treatment (indeed,
treatment may be ineffective and re-colonisation may occur). It can survive on
domesticated animals such as dogs, cats and horses. S. aureus can infect other
tissues when barriers have been breached (e.g., skin or mucosal lining). This leads
to furuncles (boils) and carbuncles (a collection of furuncles). In infants S.
aureusinfection can cause a severe disease Staphylococcal scalded skin syndrome
(SSSS). Deeply penetrating S. aureus infections can be severe. Prosthetic joints
put a person at particular risk for septic arthritis, and staphylococcal
endocarditis (infection of the heart valves) and pneumonia, which may be rapidly
spread.

Staphylococcus aureus basic characteristics

GRAM-POSITIVE COCCI IN CLUSTERS
NONMOTILE
NON-SPORE-FORMING
CATALASE: POSITIVE
OXIDASE: NEGATIVE
FACULTATIVELY ANAEROBIC

Identification of Staphylococcus aureus

production of coagulase (free coagulase)
presence of clumping factor (cell-bound coagulase)
sodium chloride tolerance (7.5%)
mannitol fermentation
hyaluronidase production (positive decapsulation test with S.equi)
Antibiotic treatment of Staphylococcus aureus infections
Should be always guided by in vitro susceptibility tests!!
Selection of appropriate antibiotics depends on diagnosis!!

IF SUSCEPTIBLE:
ampicillin/sulbactam
amoxicillin/clavulanate
oxacillin
nafcillin
cefazolin
ceftriaxone
Macrolides
Clindamycin

ALTERNATIVES:
Trimethoprim-Sulfomethoxazole(TMP-SMX)
vancomycin

MRSA
vancomycin
teicoplanin
linezolid
quinupristin/dalfopristin
TMP-SMX
rifampicin

Streptococcus pneumoniae

Streptococcus pneumoniae in clinical material occurs in two distinctive forms.

Encapsulated, virulent strains isolated e.g., from sputum in patients with acute
pneumonia, often forming highly mucoid, glistening colonies (production of capsular
polysaccharide) surrounded by a zone of alfa-hemolysis. After prolonged
cultivation (48 hours in an aerobic atmosphere enriched with 5-10% carbon dioxide)
they are often able to form colonies about 5 mm in diameter.
In throat swabs Streptococcus pneumoniae can occur in its avirulent
form (oropharyngeal carriage of pneumococci is common and they are considered to
be part of normal flora). The colonies are only 0.5-2 mm in diameter, surrounded by
zone of alfa-hemolysis and due to autolysis, often develop a dimpled rather
a craterlike appearance. These colonies are someties morphologically
indistinguishable from those of viridans streptococci but unlike viridas
streptococci are sensitive to optochin and soluble in sodium desoxycholate (bile
salts).
Streptococcus pneumoniae is a significant human pathogenic
bacterium. S.pneumoniae was recognized as a major cause of pneumonia in the late
19th century and is the subject of many humoral immunity studies. Despite the
name, the organism causes many types of pneumococcal infection other than
pneumonia, including acute sinusitis, otitis media, meningitis, bacteremia, sepsis,
osteomyelitis, septic arthritis, endocarditis, peritonitis, pericarditis, cellulitis, and
brain abscess. S.pneumoniae is the most common cause of bacterial meningitis in
adults and children, and is one of the top two isolates found in ear infection, otitis
media. Pneumococcal pneumonia is more common in the very young and the very
old. S.pneumoniae can be differentiated from viridans streptococci, some of which
are also alpha hemolytic, using an optochin test, as S.pneumoniae is optochin
sensitive. S.pneumoniaecan also be distinguished based on its sensitivity to lysis
by bile. The encapsulated, gram-positive coccoid bacteria have adistinctive

morphology on gram stain, the so-called, "lancet shape". It has a polysaccharide

capsule that acts as a virulence factor for the organism; more than 90 different
serotypes are known, and these types differ in virulence.

Streptococcus pneumoniae basic characteristics

GRAM-POSITIVE COCCI
NONMOTILE
NON-SPORE-FORMING
CATALASE: NEGATIVE
OXIDASE: NEGATIVE
FACULTATIVELY ANAEROBIC

Identification of Streptococcus pneumoniae

optochin test (sensitive)
bile solubility test (positive)
capsular swelling reaction

Antibiotic treatment of Streptococcus pneumoniae infections

Should be always guided by in vitro susceptibility tests!!
Selection of appropriate antibiotics depends on diagnosis!!

IF SUSCEPTIBLE:
penicillin
ampicillin
amoxicillin
cephalosporins I, II
macrolides

PENICILLIN RESISTANT:
cephalosporins III (e.g., cefotaxime, ceftriaxone)

ALTERNATIVES:
vancomycin
chloramphenicol

Streptococcus agalactiae

Streptococcus agalactiae on Columbia agar with 5% sheep blood (Bio-Rad).

Colonies of group B streptococci tend to be larger andoften have less pronounced
zones of beta-hemolysis than do other beta-hemolytic streptococci; some group B
strains are nonhemolytic. The majority of group B streptococci produce a
diffusable extracellular protein (CAMP factor) that acts synergistically with
staphylococcal beta lysin to lyse erythrocytes.
Streptococcus agalactiae (also known as Group B streptococcus or GBS) is a betahemolytic gram-positive streptococcus. S. agalactiae is a species of the normal
flora of the female urogenital tract and rectum. Its chief clinical importance is
that it can be transferred to a neonate passing through the birth canal and can
cause serious group B streptococcal infection. In the western world, S.
agalactiae is one of the major causes of bacterial septicemia of the newborn,
which can lead to death or long-term sequelae. S. agalactiae can also cause neonatal
meningitis, which does not present with the hallmark sign of adult meningitis, a
stiff neck; rather, it presents with nonspecific symptoms such as fever, vomiting
and irritability and can consequently go undiagnosed until it is too late. Hearing loss
can be a long-term sequelae of GBS-meningitis. Somewhat more rarely, S.
agalactiae can also cause invasive group B streptococcal disease of the adult in the
pregnant, elderly, or immunosuppressed. S. agalactiae is present in up to onethird of women of childbearing age, and one in every thousand live births will be
affected by group B streptococcal infection. In the elderly or persons with
compromised immune systems septicemia or other serious infections are seen. This
occurs also rarely during pregnancy or maternity.
.

Streptococcus agalactiae basic characteristics

GRAM-POSITIVE COCCI

NONMOTILE
NON-SPORE-FORMING
CATALASE: NEGATIVE
OXIDASE: NEGATIVE
FACULTATIVELY ANAEROBIC
Identification of Streptococcus agalactiae
catalase test (negative)
CAMP test (positive)
PYR test ( negative unlike S.pyogenes )
Lancefield's group B antigen presence(latex agglutination)
Antibiotic treatment of Streptococcus pneumoniae infections
Should be always guided by in vitro susceptibility tests!!
Selection of appropriate antibiotics depends on diagnosis!!

IF SUSCEPTIBLE:
penicillin
ampicillin
amoxicillin
cephalosporins I, II, III
macrolides
clindamycin

ALTERNATIVES:
vancomycin

Bordetella pertussis

Bordetella pertussis is a slow-growing organism that requires specialized conditions

for growth. It is the most fastidious species within the genus. One of the media
used for its cultivation is charcoal agar supplemented with 10% horse (or sheep)
blood and cephalexin. Plates are incubated in air without elevated CO 2 at 35C for a
minimum of 7 days before being reported as negative (most isolates are detected
in 2 to 4 days). Colonies are small, shiny and round. With age they become whitish
grey. Repeated subculture of B.pertussis leads to loss of fastidiousness and
laboratory adaptation to a variety of media.
Bordetella pertussis is a Gram-negative, aerobic coccobacillus of the
genusBordetella, and the causative agent of pertussis or whooping cough. Unlike B.
bronchiseptica, B. pertussis is non-motile. There does not appear to be a zoonotic
reservoir for B. pertussishumans are its only host. The bacterium is spread by
coughing and by nasal dripping. The incubation period is 714 days.
Diseases
Pertussis (or whooping cough) is an infection of the respiratory system and
characterized by a whooping sound when the person breathes in. In the US it
killed 5,000 to 10,000 people per year before a vaccine was available. Worldwide in
2000, according to the WHO, around 39 million people were infected annually and
about 297,000 died. Bordetella pertussis infects its host by colonizing lung
epithelial cells. The bacterium contains a surface protein, filamentous
hemagglutinin, which binds to sulfatides that are found on cilia of epithelial cells.
Once anchored, the bacterium produces tracheal cytotoxin, which stops the cilia
from beating. This prevents the cilia from clearing debris from the lungs, so the
body responds by sending the host into a coughing fit. These coughs expel some
bacteria into the air, which are free to infect other hosts. Bordetella pertussis has

the ability to inhibit the function of the host's immune system. Two toxins, known
as the pertussis toxin (or PTx) andadenylate cyclase (CyaA), are responsible for
this inhibition. CyaA converts ATP to cyclic AMP, and PTx inhibits an intracellular
protein that regulates this process. The end result is that phagocytes convert too
much ATP to cyclic AMP, which can cause disturbances in cellular signaling
mechanisms, and prevent phagocytes from correctly responding to an infection.
The infection occurs most with children under the age of one when they are
unimmunized or children with faded immunity, normally around the age 11 through
18. The signs and symptoms are similar to a common cold: runny nose, sneezing, mild
cough, and low-grade fever. The patient becomes most contagious during the
catarrhal stage of infection, normally 2 weeks after the coughing begins. It may
become airborne when the person coughs, sneezes, or laughs. Pertussis vaccine is
part of the DTaP (diphtheria, tetanus, acellular pertussis) immunization. The
paroxysmal cough precedes a crowing inspiratory sound characteristic of pertussis.
After a spell, the patient might make a whooping sound when breathing in, or
vomit. Adults have milder symptoms, like prolonged coughing without the whoop.
Infants less than 6 months may not have the typical whoop. A coughing spell may
last a minute or more, producing cyanosis, apnoea and seizures. However, when not
in a coughing fit, the patient does not experience trouble breathing. This is
becauseBordetella pertussis inhibits the immune response and therefore very little
mucus is generated in the lungs. A prolonged cough may be irritating and sometimes
a disabling cough may go undiagnosed in adults for many months.
General Description
GRAM-NEGATIVE RODS
NONMOTILE
NONSPOREFORMING
CATALASE: POSITIVE
OXIDASE: POSITIVE
AEROBES
BASIC TESTS FOR IDENTIFICATION
Slow growth on selective media for pathogenic bordetellae

Growth on blood agar

Glass agglutination in polyclonal antiserum specific for B.pertussis

Urease production

Nitrate reduction

ANTIBIOTIC TREATMENT
MACROLODES
Azithromycin
Clarithromycin
Erythromycin
Alternative
Trimethoprim-sulfamethoxazole (Co-trimoxazole)

Salmonella spp.

Four different serotypes of Salmonella enterica ssp. enterica on Endo agar with
biochemical slope (see here). Glucose degradation is accompanied with formation of
acid compounds (red slope) and gas production (serotype Typhi without gas). All
strains are lactose negative and conspicuous is strongly positive reaction around
mannitol tablet and H 2S production with formation of black precipitate under glass
and in area of loop punctures. Serotype Typhi and Typhimurium isolated from
hemocultures. Highly mucoid strain of serotype Enteritidis isolated from a patient
with urinary infection.

Salmonella enterica is a rod shaped, flagellated, aerobic, Gram-negative bacterium,

and a member of the genus Salmonella. S. enterica has an extraordinarily large
number of serovars or strainsover 2000 have been described. The biomedically
most relevant subspecies is called S. enterica ssp. enterica, whose following
Serovars
have
special
clinical
significance
in
human
disease:
Salmonella enterica Serovar Typhi (historically elevated to species status as S.
Typhi) is the disease agent in typhoid fever. Salmonella enterica Serovar
Typhimurium (also known as S. Typhimurium) can lead to a form of human
gastroenteritis
sometimes
referred
to
as
salmonellosis.
Salmonella enterica Serovar Paratyphi A is associated with paratyphoid fever. It is
sometimes
known
as
Salmonella
Paratyphi.
Most cases of salmonellosis are caused by food infected with S. enterica, which
often infects cattle and poultry, though also other animals such as domestic cats
and hamsters have also been shown to be sources of infection to humans. However,
investigations of vacuum cleaner bags have shown that households can act as a
reservoir of the bacterium; this is more likely if the household has contact with an
infection source, for example members working with cattle or in a veterinary clinic.

Raw chicken and goose eggs can harbor S. enterica, initially in the egg whites,
although most eggs are not infected. As the egg ages at room temperature, the
yolk membrane begins to break down and S. enterica can spread into the yolk.
Refrigeration and freezing do not kill all the bacteria, but substantially slow or
halt their growth. Pasteurizing and food irradiation are used to kill Salmonella for
commercially-produced foodstuffs containing raw eggs such as ice cream. Foods
prepared in the home from raw eggs such as mayonnaise, cakes and cookies can
spread salmonella if not properly cooked before consumption.
General Description
GRAM-NEGATIVE RODS
MOTILE
NONSPOREFORMING
CATALASE: POSITIVE
OXIDASE: NEGATIVE
FACULTATIVELY ANAEROBIC
BASIC TESTS
FOR IDENTIFICATION
MacConkey growth

Indole production

Methyl red

Voges-Proskauer

Citrate(Simmons)
(depends on serotype!!
e.g., serotype Typhi = "-")

Hydrogen sulfide(TSI)
(depends on serotype!!)

Urea hydrolysis

Lysine decarboxylase
(serot. Paratyphi A
negative)

Arginine dihydrolase

Ornithine decarb.
(depends on serotype!!;
e.g., serotype Typhi = "-")

Motility (36 C)
(depends on serotype!!)

D-glucose/gas
(depends on serotype!!
e.g., serotype Typhi "+/-")

+/+

D-mannitol fermentation

Sucrose fermentation

Lactose fermentation

D-sorbitol fermentation
(depends on serotype!!)

Cellobiose

Esculin hydrolisis

Acetate utilization

ONPG test

+ positive ( > 90% of strains are positive)

D most positive (51 - 89%)
d most negative (11 - 50%)
- negative (0 - 10%)
ANTIBIOTIC
TREATMENT
Should be always guided by
in vitro susceptibility
tests!!
Gastroenteritis
Usually no ATB treatment necessary for uncomplicated diarrheal illness.
If treated:
ampicillin
amoxicillin
trimethoprim-sulfamethoxazole
Alternative
ciprofloxacin
chloramphenicol
ceftriaxone
Typhoid fever
chloramphenicol
ampicillin
amoxicillin
trimethoprim-sulfamethoxazole
Alternative
fluoroquinolones
(e.g., ciprofloxacin)
cefotaxime
ceftriaxone

Neisseria meningitidis

For cultivation of pathogenic Neisseria are used special media for cultivation and
isolation of nutritionally fastidious microorganisms. If this media contain intact
erythrocytes, Neisseria meningitidis grows on them without
hemolysis.
As Neisseria meningitidis is resistant to vancomycin and colistin, these antibiotics
are often added directly in the medium to inhibit other gram-positive and gramnegative bacteria. Meningococcus require an aerobic atmosphere for its growth.
Carbon dioxide enhances growth, but is not required. N.meningitidis is oxidase
positive.

Neisseria meningitidis is a heterotrophic gram-negative diplococcal bacterium best

known for its role in meningitis and other forms of meningococcal disease such as
meningococcemia. N. meningitidis is a major cause of morbidity and mortality in
childhood in industrialized countries and is responsible for epidemics in Africa and
in Asia.Approximately 2500 to 3500 cases of N. meningitidis infection occur
annually in the United States, with a case rate of about 1 in 100,000. Children
younger than 5 years are at greatest risk, followed by teenagers of high school
age. Rates in sub-Saharan Africa can be as high as 1 in 1000 to 1 in 100.
Meningococci only infect humans and have never been isolated from animals
because the bacterium cannot get iron other than from human sources (transferrin
and lactoferrin). It exists as normal flora in the nasopharynx of up to 5-15% of
adults. It causes the only form of bacterial meningitis known to occur
epidemically.
Meningococcus is spread through the exchange of saliva and other respiratory
secretions during activities like coughing, kissing, and chewing on toys. Though it
initially produces general symptoms like fatigue, it can rapidly progress from
fever, headache and neck stiffness to coma and death. The symptoms are easily
confused with those of meningitis due to other organisms such as Hemophilus
influenzae andStreptococcus pneumoniae.Death occurs in approximately 10% of
cases. Those with impaired immunity may be at particular risk of meningococcus
(e.g. those with nephrotic syndrome or splenectomy; vaccines are given in cases of
removed
or
non-functioning
spleens).
Septicaemia caused by Neisseria meningitidis has received much less public
attention than meningococcal meningitis even though septicaemia has been linked
to infant deaths. Meningococcal septicaemia typically causes a purpuric rash that
does not lose its colour when pressed with a glass ("non-blanching") and does not
cause the classical symptoms of meningitis. This means the condition may be
ignored by those not aware of the significance of the rash. Septicaemia carries an
approximate 50% mortality rate over a few hours from initial onset. Note that not
all cases of a purpura-like rash are due to meningococcal septicaemia; however,
other possible causes need prompt investigation as well (e.g. ITP a platelet
disorder or Henoch-Schnlein purpura). Other severe complications include
Waterhouse-Friderichsen syndrome (a massive, usually bilateral, hemorrhage into
the adrenal glands caused by fulminant meningococcemia), adrenal insufficiency,
and
disseminated
intravascular
coagulation.
Diagnosis
The gold standard of diagnosis is isolation of N. meningitidis from sterile body
fluid. A CSF specimen is sent to the laboratory immediately for identification of
the organism. Diagnosis relies on culturing the organism on a chocolate agar plate
or a blood agar plate enriched with a growth supplement. Further testing to
differentiate the species includes testing for oxidase (all Neisseria show a
positive reaction) and the carbohydrates maltose, sucrose, and glucose test in
which N. meningitidis will oxidize (that is, utilize) the glucose and maltose.
Serology determines the group of the isolated organism. Clinical tests that are
used currently for the diagnosis of meningococcal disease take between 2 and 48
hours and often rely on the culturing of bacteria from either blood or
cerebrospinal fluid (CSF) samples. However, polymerase chain reaction tests can

be used to identify the organism even after antibiotics have begun to reduce the
infection. As the disease has a fatality risk approaching 15% within 12 hours of
infection, it is crucial to initiate testing as quickly as possible but not to wait for
the results before initiating antibiotic therapy.
General Description
DIPLOCOCCI
NONMOTILE
NONSPOREFORMING
CATALASE: POSITIVE
OXIDASE: POSITIVE
AEROBES
BASIC TESTS
FOR IDENTIFICATION
Growth on blood agar
(most strains)

Acid production from:

Glucose

Maltose

Fructose

Sucrose

-glutamylaminopeptidase

Tributyrin hydrolysis

Polysaccharide synthesis

ONPG

Reduction of NO3

Selective/ Selective Diagnostic Media

Thayer-Martin agar
Appearance
Composition
Modified Thayer-Martin agar (MTM agar)
Composition
Martin-Lewis agar (ML agar)
Composition
New York City medium (NYC medium)
Composition

ANTIBIOTIC
TREATMENT
IF SUSCEPTIBLE:
penicillin G
ampicillin
ALTERNATIVES:
ceftriaxone
cefotaxime
chloramphenicole

Enterobacter cloacae

Enterobacter cloacae on Endo agar with biochemical slope. Glucose fermentation

with gas production (fig.E), urea positive (about 70%) and H 2S negative, lactose
positive (fig.F). Sucrose, mannitol and cellobiose positive
General Characteristics
GRAM-NEGATIVE RODS
MOTILE
NONSPOREFORMING
CATALASE: POSITIVE
OXIDASE: NEGATIVE
FACULTATIVELY ANAEROBIC

BASIC TESTS FOR IDENTIFICATION

MacConkey growth

Indole production

Methyl red

Voges-Proskauer

Citrate(Simmons)

Hydrogen sulfide(TSI)
Urea hydrolysis

Lysine decarboxylase

Arginine dihydrolase

Ornithine decarboxylase

Motility (36 C)

D-glucose acid/gas

+/+

D-mannitol fermentation

Sucrose fermentation

Lactose fermentation

D-sorbitol fermentation

Cellobiose

Esculin hydrolisis

Acetate utilization

ONPG test

+ positive ( > 90% of strains are positive)

D most positive (51 - 89%)
d most negative (11 - 50%)
- negative (0 - 10%)

ANTIBIOTIC
TREATMENT
Should be always guided by
in vitro susceptibility
tests!!
Selection of appropriate antibiotics depends on diagnosis!!
IF SUSCEPTIBLE:
Imipenem + aminoglycoside(e.g., gentamicin or amikacin)
Piperacillin/tazobactam + aminoglycoside Fluoroquinolones(e.g.,
ciprofloxacin, norfloxacin)
Trimethoprim-sulfamethoxazole (Co-trimoxazole)
Nitrofurantoin
Ceftazidime
ALTERNATIVES:
Cephalosporins III, IV

Corynebacterium diphtheriae

Corynebacterium diphtheriae can grow on media with sheep blood with or without
beta-hemolysis. Tinsdale agar (TIN) is used for the primary isolation and
identification of Corynebacterium diphtheriae. The medium differentiates
betweenC. diphtheriae and diphtheroids found in the upper respiratory tract. This
differentiation was based on the ability of C. diphtheriae to produce black (or
brown) colonies, surrounded by a brown/black halo. The dark halo is due to the
production of H2S from cystine, interacting with the tellurite salt.
Corynebacteria are Gram-positive, catalase positive, non-spore-forming, nonmotile, rod-shaped bacteria that are straight or slightly curved. Metachromatic
granules are usually present representing stored phosphate regions. Their size falls
between 2-6 micrometers in length and 0.5 micrometers in diameter. The bacteria
group together in a characteristic way, which has been described as the form of a
"V", "palisades", or "Chinese letters". They may also appear elliptical. They are
aerobic or facultatively anaerobic, chemoorganotrophs, with a 5165% genomic
G:C content. They are pleomorphic through their life cycle: they come in various
lengths and frequently have thickenings at either end, depending on the
surrounding
conditions.
Diseases
The most notable human infection is diphtheria, caused by Corynebacterium
diphtheriae. It is an acute and contagious infection characterized by
pseudomembranes of dead epithelial cells, white blood cells, red blood cells, and
fibrin that form around the tonsils and back of the throat. It is an uncommon
illness that tends to occur in unvaccinated individuals, especially school-aged
children, those in developing countries, elderly, neutropenic or
immunocompromised patients. It can occasionally infect wounds, the vulva, the
conjunctiva, and the middle ear. The virulent and toxigenic strains are lysogenic,
and produce an exotoxin formed by two polypeptide chains, which is itself
produced when a bacterium is transformed by a gene from the prophage.

Four subspecies are recognized: C. diphtheriae mitis, C. diphtheriae intermedius,

C. diphtheriae gravis, and C. diphtheriae belfanti. The four subspecies differ
slightly in their colonial morphology and biochemical properties such as the ability
to metabolize certain nutrients, but all may be toxigenic (and therefore cause
diphtheria) or non-toxigenic. Unusually, the diphtheria toxin gene is actually
encoded by a bacteriophage which is found in toxigenic strains, not on the bacterial
chromosome itself.
General Description
GRAM-POSITIVE RODS
NONMOTILE
NONSPOREFORMING
CATALASE: POSITIVE
OXIDASE: NEGATIVE
FACULTATIVELY
ANAEROBIC
BASIC TESTS
FOR IDENTIFICATION
resistant to tellurite

cystinase activity

resistant to fosfomycin

growth on MacConkey agar

nitrate reductase

urea

esculin

glucose

maltose

sucrose

maninitol

xylose
sugar fermentation: positive (> 90% of strains)

ANTIBIOTIC
TREATMENT
Erythromycin
Penicillin
ALTERNATIVES:
Clindamycin

Listeria monocytogenes

Listeria monocytogenes is a non-fastidious bacterium growing well on commonly

used cultivation media. On blood agar it forms small colonies about 1-2 mm in
diameter after 24 hours of cultivation. Colonies are -hemolytic but many
strains ofL.monocytogenes produce only narrow zones of hemolysis that
frequently do not extend much beyond the edge of the colonies. In appearance,
they may resemble colonies of Enterococci or some Corynebacteria.
Listeria monocytogenes is a bacterium commonly found in soil, stream water,
sewage, plants, and food. Each bacterium is gram-positive and rod-shaped.
Listeria are known to be the bacteria responsible for listeriosis, a rare but
potentially lethal food-borne infection: the case fatality rate for those with a
severe form of infection may approach 25%. They are incredibly hardy and able
to grow in temperatures ranging from 4C (39F), the temperature of a
refrigerator, to 37C (99F), the body's internal temperature. Furthermore,
listerosis's deadliness can be partially attributed to the infection's ability to
spread to the nervous system and causemeningitis. Finally, Listeria has a
particularly high occurrence rate in newborns because of its ability to infect

the fetus by penetrating the endothelial layer of the placenta. Vegetables can
become contaminated from the soil, and animals can also be carriers. Listeria
has been found in uncooked meats, uncooked vegetables, unpasteurized milk,
foods made from unpasteurized milk, and processed foods. Listeria is killed by
pasteurization and cooking. There is a chance that contamination may occur in
ready-to-eat foods such as hot dogs and deli meats because contamination may
occur after cooking and before packaging.
General Description
GRAM-POSITIVE RODS
MOTILE
NONSPOREFORMING
CATALASE: POSITIVE
OXIDASE: NEGATIVE
FACULTATIVELY
ANAEROBIC
BASIC TESTS
FOR IDENTIFICATION
Growth at 4C

Motility at 25C

Motility at 37C

Tumbling motility in a wet mount

CAMP test reaction

(with S.aureus)

-hemolysis

+*

Innate resistance to CEPHALOSPORINS I,II,III

D-glucose

VP test

Methyl red

Esculin

Urea

Gelatin

Indole

H2S

D-xylose

After 24 hours of cultivation is -hemolysis often only weak, visible after

removal of colonies.
Selective/ Selective Diagnostic Media
PALCAM agar
LPM agar
Oxford agar

ANTIBIOTIC
TREATMENT
Ampicillin(or AMP + GEN)
Penicillin G (+GEN)
ALTERNATIVES
Trimethoprim-Sulfome-thoxazole(TMP-SMX)
POSSIBLE THERAPY (if susceptible)
Tetracycline
Chloamphenicol
Erythromycin

Streptococcus pyogenes
Group A streptococcus