MEDICAL MICROBIOLOGY I

Lesson 11
Enterobacteriaceae Part I

Enterobacteriaceae
The family Enterobacteriaceae is the largest
most heterogeneous collection of medically
important Gram negative bacilli
A total of 32 genera and more than 130
species have been described
These genera have been classified based on
biochemical properties, antigenic structure,
and nucleic acid hybridisation and sequencing

Enterobacteriaceae
Enterobacteriaceae are ubiquitous organisms,
being found worldwide in soil, water and
vegetation, and are part of the normal intestinal
flora of most animals, including human
Some organisms (e.g. Salmonella typhi, Shigella
sp., Yersinia pestis) are always associated with
disease
Others (e.g. E. coli, Klebsiella pneumoniae,
Proteus mirabilis) are members of the normal
commensal flora that can cause opportunistic
infections

Enterobacteriaceae
A third group of Enterobacteriaceae exists those normally commensal organisms that
become pathogenic when they acquire
virulence factor genes on plasmids,
bacteriophages, or pathogenicity islands (e.g.
E. coli associated with gastroenteritis)
Infections with the Enterobacteriaceae can
originate from an animal reservoir, or through
the endogenous organisms in a susceptible
patient and can involve virtually all body sites

Enterobacteriaceae

Physiology and Structure

Moderately sized (0.3 - 1.0 x 1.0 - 6.0 m)
Gram negative bacilli
They are either non-motile or motile with
peritrichous flagella and do not form spores
All members can grow rapidly aerobically and
anaerobically (facultative anaerobes) on a
variety of non-selective (e.g. blood agar) and
selective (e.g. MacConkey) media

Physiology and Structure

Simple nutritional requirements - ferment
glucose, reduce nitrites, and are catalasepositive and oxidase-negative
The absence of cytochrome oxidase activity is
an important characteristic, because it can be
measured rapidly with a simple test and is
used to distinguish the Enterobacteriaceae
from many other fermentative and nonfermentative Gram negative bacilli

Physiology and Structure

Characteristics of the organisms colonies on
different media have been used to identify
common members of the family
Enterobacteriaceae
Ability to ferment lactose: differentiate lactosefermenting strains (e.g. Escherichia, Klebsiella,
Enterobacter, Citrobacter and Serratia sp.)
from non-lactose fermenting strains (e.g.
Proteus, Salmonella and Yersinia sp.)

Physiology and Structure

Resistance to bile salt: separate enteric
pathogen (e.g. Shigella, Salmonella) from
commensal organisms inhibited by bile salt

Enterobacteriaceae have prominent capsules,

whereas other strains are surrounded by a
loose-fitting, diffusible slime layer

Physiology and Structure

The heat-stable lipopolysaccharide (LPS) is the
major cell wall antigen and consists of 3
components:
The somatic O polysaccharide
A core polysaccharide
Lipid A

Physiology and Structure

The serology classification of the
Enterobacteriaceae is based on 3 antigens:
somatic O polysaccharide
Capsular K antigens
Flagella H proteins

Specific O antigens are present in each genus,

although cross-reactions between closely
related genera are common (e.g. Salmonella
with Citrobacter, Escherichia with Shigella)

Physiology and Structure

Different genera both within and outside the
family Enterobacteriaceae possess K antigens
e.g. E. coli K1 cross-reacts with Neisseria
meningitidis and Haemophilus pneumoniae,
and Klebsiella pneumoniae cross-reacts with
Streptococcus pneumoniae
H antigens are heat-labile, flagellar proteins
They may be absent from a cell, or they may
undergo antigenic variation and be present in
2 phases

Pathogenesis and Immunity

Virulence factors:
1.
2.
3.
4.
5.
6.

Endotoxin
Capsule
Antigenic phase variation
Sequestration of growth factors
Resistance to serum killing
Antimicrobial resistance

Virulence Factor - Endotoxin

A virulence factor shared among all aerobic
and some anaerobic Gram negative bacteria
The activity of this toxin depends on the lipid
A component of lipopolysaccharide, which is
released at cell lysis

Virulence Factor - Endotoxin

Many of the systemic manifestations of Gram
negative bacterial infections are initiated by
endotoxin, including:
Activation of complement release of cytokines,
leukocytosis, thrombocytopenia, disseminated
intravascular coagulation, fever, decreased
peripheral circulation, shock, death

Virulence Factor - Capsule

Encapsulated Enterobacteriaceae are
protected from phagocytosis by hydrophilic
capsular antigens, which repel the
hydrophobic phagocytic cell surface
These antigens interfere with the binding of
antibodies to the bacteria and are poor
immunogens or activators of complement
The protective role of capsule is diminished,
however, if the patient develops specific anticapsular antibodies

Virulence Factor - Antigenic Phase

Variation
The expression of capsular K and flagellar H
antigens is under the genetic control of the
organism
Each of these antigens can be alternately
expressed or not expressed (phase variation),
a feature that protects the bacteria from
antibody-mediated cell death

Virulence Factor - Type III Secretion

Systems
A variety of distinct bacteria (e.g. Yersinia,
Salmonella, Shigella, Escherichia, Pseudomonas,
Chlamydia) have a common effector system for
delivering their virulence genes into targeted
eukaryotic cells
This system consists of approximately 20
proteins that facilitate secretion of bacterial
virulence factor into host cells
In the absence of the type III secretion system
the bacteria lose their virulence

Virulence factor - Sequestration of

Growth Factors
Nutrients are provided to the organisms in
enriched culture media, but the bacteria must
become nutritional scavengers when growing
in vivo
Iron is an important growth factor required by
bacteria, but it is bound in haeme proteins
(e.g. haemoglobin, myoglobin) or in ironchelating proteins (e.g. transferrin, lactoferrin)

Virulence factor - Sequestration of

Growth Factors
The bacteria counteract the binding by
producing their own competitive ironchelating compounds (e.g. siderophores
enterobactin and aerobactin)

Virulence Factor - Resistance to Serum

Killing
Virulent organisms capable of producing
systemic infections are frequently resistant to
serum killing
Although the bacterial capsule can protect the
organism from serum killing, other factors
prevent the binding of complement
components to the bacteria and subsequent
complement-mediated clearance

Virulence Factor - Antimicrobial

Resistance
As rapidly as new antibiotics are introduced,
organisms can develop resistance to them
The resistance can be encoded on transferable
plasmid and exchanged among species,
genera, and even families of bacteria

Escherichia coli
The genus Escherichia consists of 5 species, of
which E. coli is the most common and
clinically most important
This organism is associated with a variety of
disease, including sepsis, UTIs, meningitis, and
gastroenteritis
Many O, H, and K antigens have been
described, and they are used to classify the
isolates for epidemiologic purposes

Escherichia coli

Escherichia coli

Pathogenesis and Immunity

Specialised virulence factors:
Adhesins

Colonisation factor antigens CFA/I, CFA/II and CFA/III

Aggregative adherence fimbriae AFF/I and AFF/II
Bundle-forming protein (Bfp)
Intimin
P pili
Ipa protein
Dr fimbriae

Exotoxins

Heat-stable toxins Sta and STb

Shiga toxins Stx-1 and Stx-2
Haemolysin HlyA
Heat-labile toxins LT-I and LT-II

Epidemiology
Large numbers of E. coli are present in the GI
tract, and the bacteria are common causes of
sepsis, neonatal meningitis, infections of the
urinary tract, and gastroenteritis
The most common Gram negative bacilli
isolated from patients with sepsis
Responsible for causing more than 80% of all
community-acquired UTIs as well as hospitalacquired infections

Epidemiology
A prominent cause of gastroenteritis in
developing countries
Most infections (with the exception of
neonatal meningitis and gastroenteritis) are
endogenous; that is, E. coli that are part of the
patients normal flora are able to establish
infection when the patients defenses are
compromised

Clinical Diseases - Septicaemia

Originates from infections in the urinary or GI
tract (e.g. an intra-abdominal infection with
sepsis following intestinal perforation)
The mortality associated with E. coli
septicaemia is high for patients in whom
immunity is compromised or the primary
infection is in the abdomen or central nervous
system

Clinical Diseases - Urinary Tract Infection

(UTI)
Originate in the colon, contaminate the urethra,
ascend into the bladder, and may migrate to the
kidney or prostate
Disease is more common with certain specific
serogroups
These bacteria are particularly virulent because of
their ability to produce adhesins (primarily P pili,
AAF/I, AAF/II, and Dr.), which bind to cell lining the
bladder and upper UT, and haemolysin HlyA

Clinical Diseases - Neonatal Meningitis

E. coli and group B streptococci cause the
majority of central nervous systemic infections
in infants younger than 1 month
Approximately 75% of the E. coli strains
possess the K1 capsular antigen, which is
commonly present in the GI tract of pregnant
women and newborn infants

Clinical Diseases - Gastroenteritis

Divided into 6 groups:
1.
2.
3.
4.
5.
6.

Enterotoxigenic E. coli (ETEC)

Enteropathogenic E. coli (EPEC)
Enteroinvasive E. coli (EIEC)
Enterohaemorrhagic E. coli (EHEC)
Enteroaggregative E. coli (EAEC)
Diffusely adherent E. coli (DAEC)

Clinical Diseases - Gastroenteritis

1. ETEC
Most commonly in developing countries, most
common cause of Travellers diarrhoea
The inoculum for disease is high, so infections are
primarily acquired through consumption of faecally
contaminated food or water
Produce 2 classes of enterotoxins: heat-labile toxins
(LT-I, LT-II) and heat-stable toxins (STa and STb
LT-I is structurally similar to cholera toxin
This toxin consists of one A subunit and 5 identical B
subunits

Clinical Diseases - Gastroenteritis

2. EPEC
Major cause of infant diarrhoea in
impoverished countries, rare in older children
and adults, presumably because they have
developed protective immunity
Disease is characterised by bacterial
attachment to epithelial cells of the small
intestine with subsequent effacement
(destruction) of the microvilli, which result in
diarrhoea

Clinical Diseases - Gastroenteritis

Initially a loose attachment mediated by
bundle-forming pili (Bfp) occurs, followed by
active secretion of proteins by the bacterial
type III secretion system into host epithelial
cell
Translocated intimin receptor (Tir) is inserted
into the epithelial cell membrane and
functions as a receptor for an outer membrane
bacterial adhesin, intimin

Clinical Diseases - Gastroenteritis

3. EIEC
Closely related by phenotypic and pathogenic
properties to Shigella
Produce disease similar to shigellosis
The bacteria are able to invade and destroy the
colonic epithelium, producing a disease
characterised initially by watery diarrhoea
A series of bacterial genes carried on a plasmid
mediate invasion (plnv genes) into the colonic
epithelium

Clinical Diseases - Gastroenteritis

The bacteria then lyse the phagocytic vacuole
and replicate in the cell cytoplasm
Movement within the cytoplasm and into
adjacent epithelial cells is regulated by
formation of actin tails
This process of epithelial cell destruction with
inflammatory infiltration can progress to
colonic ulceration

Clinical Diseases - Gastroenteritis

4. EHEC
The most common strains producing disease in
developed countries
The ingestion of fewer than 100 bacilli can
produce disease
Symptoms: uncomplicated diarrhoea to
haemorrhagic colitis with severe abdominal
pain, bloody diarrhoea, and little or no fever

Clinical Diseases - Gastroenteritis

Haemolytic uremic syndrome (HUS)
Characterised by acute renal failure,
thrombocytopenia, and microangiopathic
haemolytic anaemia
Most cases attributed to the consumption of
undercooked ground beef or other meat
products, water, unpasteurised milk or fruit
juices, uncooked vegetables, and fruits

Clinical Diseases - Gastroenteritis

Express a Shiga-like toxin, induce lesions on
epithelial cells, and possess a 60 MDa plasmid
that carries genes for other virulence factors
The Stx toxins also stimulate expression of
inflammatory cytokines (e.g. TNF-, IL-6)
Death can occur in 3 - 5% of patients with HUS,
and severe sequelae (e.g. renal impairment,
hypertension, CNS manifestation) can occur in
as many as 30% of patients

Clinical Diseases - Gastroenteritis

5. EAEC
Implicated as a cause of persistent, watery
diarrhoea with dehydration in infants in
developing countries
The bacteria is characterised by their autoagglutination in a stacked brick arrangement mediated by bundle - forming fimbriae
Stimulate secretion of mucus, shortening of
the microvilli, mononuclear infiltration, and
haemorrhage are observed

Clinical Diseases - Gastroenteritis

6. DAEC
Adherence characteristic to cultured cells
Stimulate elongation of the microvilli with the
bacteria embedded in the cell membrane
The resulting disease is watery diarrhoea
found primarily in infants between 1 - 5 years
of age