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Module: BIOM – 1009

Lecturer: Dr Ioannou

Date: 10/10/16

Medically Important Gram Negative Bacteria
o

As previously mentioned, Gram negative bacteria have a lipopolysaccharide (LPS) structure on its outer
surface, its complex and has some unique features
 Proteins called porins allow passive diffusion into the periplasmic space
 The presence of a small lipoprotein (Braun’s Lipoprotein) is covalently bonded to the
peptidoglycan and is embedded on the outer membrane by hydrophobic lipids
 The LPS consists of 3 parts:
 Lipid A
 This is an endotoxin molecule
o This can cause different effects to the body depending on dosage:
 Small doses can give mild fevers and bacterial death
 Large doses can cause low BP, increased heart rate and
liver/kidney failure, among other things
 It contains the hydrophobic membrane-anchoring region of the LPS
o This consists of phosphorylated N-acetylglucosamine (NAG) with 6/7
fatty acids bonded to it
 Core polysaccharide
 This contains some unusual sugars such as 2-keto-3-deoxyoctonate
 It is attached to the 6 position of one NAG
 Side chain of repeating sugars (O-Antigen)
 This is much longer than the core polysaccharide and it maintains the hydrophilic
domain of the LPS
 The composition of the O chain differs greatly between species and strains of
bacteria
 Some genera of Gram negative bacteria do not have the O-specific side chain, such as Neisseria
and Haemophilus

o

Gram negative bacteria and their toxins can cause bacterial sepsis during blood transfusions
...
coli and Enterobacter aerogenes can cause this
 Symptoms of sepsis in the first 4 hours after a transfusion include:
 Fever, rigors and high/low BP
 Stop the transfusion if the patient’s temperature increases by 2o and there is nausea and vomiting

o

Even after sterilization, the Gram negative bacteria may be dead, but the endotoxins can survive due to
their heat stability and insensitivity to pH

o

Some medically important Gram negative bacteria are Salmonella, E
...
coli O104, Haemophilus

influenzae, Legionella


The next 4 pages contain information on each

Salmonella
o

Salmonella is a genus of bacilli (rod shaped) Gram negative bacteria of the Enterobacteriaceae family
 The species found in warm blooded animals is Salmonella enterica
 This can then be divided in to 6 subspecies with over 2500 serotypes

o

The bacteria are usually found in the gastrointestinal tract of animals, birds, amphibians and sometimes
humans
 Poultry are especially susceptible
 Humans may carry Salmonella typhi (Typhoid Mary) in their GI tracts

o

There are many diseases associated with the salmonella bacteria, these include:
 Salmonellosis
 Typhoid Fever
 Gastroenteritis
 Enteric Fever

o

Most salmonella are motile with one exception, Salmonella gallinarum
 Salmonella move by means of flagella attached to them

o

Salmonella culture differently on different agar media, the left image is of a blood agar medium, and the
right is on DeoxyCholate Agar (DCA)

o

Key characteristics of Salmonella include:
 They are oxidase negative and indole negative
 They produce H2S (Hydrogen Sulphide)
 They grow at 37o in aerobic conditions, for an optimal time of 24 hours
 They are grown on selective agar such as:
 DCA, Bismuth sulphate, Hektoen agar and Selenite broth
 For further analysis, the sample may be cultured on Cysteine Lactose Electrolyte
Deficient (CLED) agar
o Salmonella are non-lactose fermenting

o

They are classified using the following techniques:
 The O antigen provides the serogroup
 The serotype is provided from one of the 46 serogroups
 The H antigen on the flagella allows the identification of different strains
 There are currently 2,523 known salmonella serotypes

o

Samples can be detected using:
 Faecal samples
 Selenite broth (this inhibits growth of Gram positive bacteria)
 Slide agglutinations (on slides suspected to be pure salmonella)

E coli O157 (separate to E
...
coli and is part of the Enterobacteriaceae family
 This particular strain is associated with virulence

o

This particular strain is rarely found, approximately 15% of cattle carry E
...
coli are found in most animal’s GI tracts and are usually harmless and part of the
normal bowel flora
 E
...
coli O157 is ~5% but can be higher in
susceptible individuals:
 The very young (They can contract Haemolytic Uremic Syndrome)
o This causes the destruction of red blood cells and so causes acute kidney
failure and low platelet count among other things
 The elderly

o

Humans can become infected by eating contaminated foods such as inadequately cooked beef burgers
and unpasteurised milk
 It can also be found in cooked meats, raw vegetables, yoghurt due to poor hygiene practice

o

E
...
coli have to be cultured behind a microbiological safe cabinet
 Only a low inoculum size is required to cause disease

o

Use of selective agar such as Sorbitol MacConkey (SMAC) or chromogenic agar is used to differentiate E
...
coli
...
coli O157 on 3 different agar media:

Haemophilus influenzae
o

H
...
influenzae will grow on enriched blood agar, such as chocolate
agar in an enriched CO2 atmosphere (5-10%)
...
influenzae

o

Growth of H
...
On discs with one or the other, the bacteria will not
grow:

o

The H
...
When absent, it is
associated with less sever diseases

Legionella

o

Legionella is a slender, Gram negative bacillus of the family Legionellaceae
 Legionella pneumophila is associated with Legionnaires disease/Legionellosis


The bacterium may not show up under a Gram stain unless it flooded with safranin for at least 10
minutes

o

The bacteria are widely distributed among ponds and hot or cold water systems
...
The left image shows this
 However, it takes 7 days for growth to occur

o

Another technique is the Legionella direct immunofluorescence test
...
It is shown on the right

o

Legionellosis is more dangerous to males than females it would seem:
 It affects more men than women
 It mostly affects people over the age of 40
 It is associated with travel
 It has the greatest mortality in males



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