Medical Microbiology I - Lecture11

MEDICAL MICROBIOLOGY I

Lesson 11 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 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 Some organisms (e.g. Salmonella typhi, Shigellasp., 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. 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 sporesperitrichous 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


Simple nutritional requirements - ferment glucose, reduce nitrites, and are catalase-positive and oxidase-negative

The absence of cytochrome oxidase activity is 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 non-fermentative Gram negative bacilli


Characteristics of the organisms colonies on

different media have been used to identify

common members of the family

EnterobacteriaceaeEnterobacteriaceae

Ability to ferment lactose: differentiate lactose-

fermenting strains (e.g. Escherichia, Klebsiella,

Enterobacter, Citrobacter and Serratia sp.)

from non-lactose fermenting strains (e.g.

Proteus, Salmonella and Yersinia sp.)


Resistance to bile salt: separate enteric

pathogen (e.g. Shigella, Salmonella) from

commensal organisms inhibited by bile salt

Enterobacteriaceae have prominent capsules, Enterobacteriaceae have prominent capsules,

whereas other strains are surrounded by a

loose-fitting, diffusible slime layer


The heat-stable lipopolysaccharide (LPS) is the

major cell wall antigen and consists of 3

components:components:

The somatic O polysaccharide

A core polysaccharide

Lipid A


The serology classification of the

Enterobacteriaceae is based on 3 antigens:

somatic O polysaccharide

Capsular K antigens 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)


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, 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. Endotoxin

2. Capsule

3. Antigenic phase variation3. Antigenic phase variation

4. Sequestration of growth factors

5. Resistance to serum killing

6. 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 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: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 anti-capsular 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 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 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 iron-

chelating proteins (e.g. transferrin, lactoferrin)

Virulence factor - Sequestration of

Growth Factors

The bacteria counteract the binding by

producing their own competitive iron-

chelating compounds (e.g. siderophores chelating 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 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 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

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 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 hospital-acquired infections

Epidemiology

A prominent cause of gastroenteritis in

developing countries

Most infections (with the exception of

neonatal meningitis and gastroenteritis) are 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 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 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 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. Enterotoxigenic E. coli (ETEC)

2. Enteropathogenic E. coli (EPEC)

3. Enteroinvasive E. coli (EIEC)3. Enteroinvasive E. coli (EIEC)

4. Enterohaemorrhagic E. coli (EHEC)

5. Enteroaggregative E. coli (EAEC)

6. Diffusely adherent E. coli (DAEC)

Clinical Diseases - Gastroenteritis1. 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 waterprimarily 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


2. EPEC

Major cause of infant diarrhoea in impoverished countries, rare in older children and adults, presumably because they have developed protective immunitydeveloped 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


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 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


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 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


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 adjacent epithelial cells is regulated by

formation of actin tails

This process of epithelial cell destruction with

inflammatory infiltration can progress to

colonic ulceration


4. EHEC

The most common strains producing disease in

developed countries

The ingestion of fewer than 100 bacilli can 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


Haemolytic uremic syndrome (HUS)

Characterised by acute renal failure,

thrombocytopenia, and microangiopathic

haemolytic anaemiahaemolytic 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


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 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


5. EAEC

Implicated as a cause of persistent, watery diarrhoea with dehydration in infants in developing countries

The bacteria is characterised by their auto-agglutination in a stacked brick arrangement -mediated by bundle - forming fimbriae

Stimulate secretion of mucus, shortening of the microvilli, mononuclear infiltration, and haemorrhage are observed


6. DAEC

Adherence characteristic to cultured cells

Stimulate elongation of the microvilli with the

bacteria embedded in the cell membranebacteria embedded in the cell membrane

The resulting disease is watery diarrhoea

found primarily in infants between 1 - 5 years

of age

Medical Microbiology I - Lecture11

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