Gram Negative Bacteria: Miscellaneous but Important!
27 October 2020 22:31
Important Gram Negative Bacteria 1
Fastidious Gram-negative Rods:
- Legionella pneumophila: Legionnaire's Disease
- Bordetella pertussis: Whooping cough
- Haemophilus influenzae: meningitis and septicaemia in children < 6 months
Gram- negative obligate anaerobes (very diverse group):
○ Bacteroides
○ Fusobacterium
○ Prevotella
○ Porphyromonas
The method gram staining:
The method was developed by Hans Christian Gram and he was a Danish microbiologist. The staining
method differentiates two main gram negative and gram positive bacteria. The staining method
involves crystal violet and iodine which get taken up into the cell wall (peptidoglycan layer).
If there is a thick peptidoglycan layer, the crystal violet colour will bind to the gram positive and the
stain won't be removed by alcohol or any decolourising agent. However, if you use a decolourising
agent on gram negative, it will wash off and the counter stain safranin or fuchsine will bind the thin
peptidoglycan layer.
Safranin/Fuchsine counterstain is applied which stains all cells, allowing the identification of Gram-
negative bacteria as well.
Fastidious meaning that the gram negative bacteria are difficult to grow within lab environment due
to not sufficient/ appropriate environment, nutrient etc. Therefore it is difficult to identify.
Legionella Pneumophila:
It causes pneumonia and very often associated with outbreaks.
1st Outbreak:
Bellevue-Stratford Hotel, Broad St, Philadelphia
The 58th State Convention of the American Legion of Pennsylvania, July 1976, 4000 World War II
Legionnaires and their families and friends. Around 600 staying at the hotel at which the convention
is hosted. On day two of the convention Legionnaires began to fall ill… chest pains, breathlessness.
- Eventually 34 dies, 221 ill (..72 walking past lobby)
- Epidemiological evidence suggested exposure in lobby
- Centre for Disease Control (CDC) feared terrorist plot
- Rickettsiologist Dr Joseph McDade 1977. Inoculated guinea pigs with material from deceased
and noted these fastidious bacilli within macrophages.
Dr Joseph extracted the infectious material from the deceased individuals and inoculated in an
animal model (guinea pig) to study the organism and its pathogenesis. Within the tissue section of
the guinea pig, he noted fastidious gram negative bacterium within macrophage.
It is later believed that the bacteria was present within the air conditioning system in the hotel which
then transmitted onto people walking past lobbies.
Dr Carl Fliermans researched the bacteria species in:
• L.pneumophila lipids resembled those of the thermophilic bacteria in Yellowstone National
Park, there are thermal pools within the national park and the bacteria tended to live as
biofilm.
• Discovered the bacteria in thermal waters discharged from nuclear reactor at Savannah River
Gram Negative- Legionella^J Bordetella Page 1
27 October 2020 22:31
Important Gram Negative Bacteria 1
Fastidious Gram-negative Rods:
- Legionella pneumophila: Legionnaire's Disease
- Bordetella pertussis: Whooping cough
- Haemophilus influenzae: meningitis and septicaemia in children < 6 months
Gram- negative obligate anaerobes (very diverse group):
○ Bacteroides
○ Fusobacterium
○ Prevotella
○ Porphyromonas
The method gram staining:
The method was developed by Hans Christian Gram and he was a Danish microbiologist. The staining
method differentiates two main gram negative and gram positive bacteria. The staining method
involves crystal violet and iodine which get taken up into the cell wall (peptidoglycan layer).
If there is a thick peptidoglycan layer, the crystal violet colour will bind to the gram positive and the
stain won't be removed by alcohol or any decolourising agent. However, if you use a decolourising
agent on gram negative, it will wash off and the counter stain safranin or fuchsine will bind the thin
peptidoglycan layer.
Safranin/Fuchsine counterstain is applied which stains all cells, allowing the identification of Gram-
negative bacteria as well.
Fastidious meaning that the gram negative bacteria are difficult to grow within lab environment due
to not sufficient/ appropriate environment, nutrient etc. Therefore it is difficult to identify.
Legionella Pneumophila:
It causes pneumonia and very often associated with outbreaks.
1st Outbreak:
Bellevue-Stratford Hotel, Broad St, Philadelphia
The 58th State Convention of the American Legion of Pennsylvania, July 1976, 4000 World War II
Legionnaires and their families and friends. Around 600 staying at the hotel at which the convention
is hosted. On day two of the convention Legionnaires began to fall ill… chest pains, breathlessness.
- Eventually 34 dies, 221 ill (..72 walking past lobby)
- Epidemiological evidence suggested exposure in lobby
- Centre for Disease Control (CDC) feared terrorist plot
- Rickettsiologist Dr Joseph McDade 1977. Inoculated guinea pigs with material from deceased
and noted these fastidious bacilli within macrophages.
Dr Joseph extracted the infectious material from the deceased individuals and inoculated in an
animal model (guinea pig) to study the organism and its pathogenesis. Within the tissue section of
the guinea pig, he noted fastidious gram negative bacterium within macrophage.
It is later believed that the bacteria was present within the air conditioning system in the hotel which
then transmitted onto people walking past lobbies.
Dr Carl Fliermans researched the bacteria species in:
• L.pneumophila lipids resembled those of the thermophilic bacteria in Yellowstone National
Park, there are thermal pools within the national park and the bacteria tended to live as
biofilm.
• Discovered the bacteria in thermal waters discharged from nuclear reactor at Savannah River
Gram Negative- Legionella^J Bordetella Page 1
