Gram Negative Bacilli, bench to bedisde (word document)

Question 1

What is the outermost part of a bacteria called?

Answer 1

cell wall - composed of petidoglycans.

Question 2

What is just inside the cell wall? What are the functions?

Answer 2

plasma membrane. A selective barrier. Contains enzymes vital to live function of the cell.

Question 3

What are the two groups of bacteria?

Answer 3

Gram positive

Gram negative

Question 4

What is the structure of the outside of gram positive bacteria? gram negative?

Answer 4

Gram positive have an outer surface of cell membrane followed by the cell wall which is a thick peptidoglycan layer.
Gram negative have a thin cell wall (which is the thin peptidoglycan layer) as well as an additional outer membrane composed of phospholipids and lipopolysaccharides. The outer layers have channels - porins, non-specific, and specific channels.

Question 5

What is the genetic material form in bacteria?

Answer 5

In most bacteria the genetic material is contained in one circular chromosome.
This has genes composed of DNA. Bacteria have prokaryotes which means there is no cell nucleus. In addition the circular chromosome, many bacteria contain circular molecules of DNA called plasmids, which may cary genes for resistence to antibiotics and production of toxins for survival.

The bacteria contain ribosomes for the production of proteins.

Question 6

What happens in gram-staining of gram-negative bacteria?

Answer 6

The thin peptidoglycan layer of their cell wall is sandwiched between an inner cell membrane and a bacterial outer membrane. In Gram staining, the outer lipid-based membrane of gram-negative bacteria is removed by an alcohol solution. The alcohol also decolorizes the then exposed peptidoglycan layer by dissolving away the previously applied crystal violet. A counterstain (safranin or fuchsine) is then added which recolorizes the bacteria red or pink.

Gram-positive bacteria on the other hand have a thicker peptidoglycan layer in their cell wall outside the cell membrane, which retains the crystal violet stain during the alcohol wash, so long as it is timed correctly. The counter stain may also be absorbed by gram positive bacteria but the darker crystal violet stain predominates visually.

All gram-positive bacteria are bounded by only a single unit lipid membrane and they generally contain a thick layer (20–80 nm) of peptidoglycan responsible for retaining the Gram stain.

In contrast to gram-positive bacteria, all archetypical gram-negative bacteria are bounded by both a cytoplasmic membrane and an outer cell membrane, and they contain only a thin layer of peptidoglycan (2–3 nm) in between these two membranes. The presence of both inner and outer cell membranes defines a new compartment in these cells, the periplasmic space or the periplasmic compartment.

Question 7

what is the periplasmic space?

Answer 7

The periplasm is a space bordered by two selective permeable barriers, i.e., biological membranes, which are the inner membrane (i.e., cytoplasmic membrane) and the outer membrane in Gram-negative bacteria. Strictly speaking, there is no periplasmic space in Gram-positive bacteria because there is only one biological membrane, the cytoplasmic membrane. A region termed “inner wall zone” (IWZ) has been observed between the cytoplasmic membrane and the mature cell wall.[1][2]

Question 8

What are the 4 groups of facultative and aerobic gram negative bacilli? (excluding anaerobic GNB)

Answer 8

1. Fermentative (enterobacteriaceae)
2. Non-Fermentative (Environmental)
3. Fastidious/Miscellaneous
4. Curved and Spiral

Question 9

Where are enterobacteriaceae found?

Answer 9

Organisms in this family of gram-negative bacilli are commonly found in soil and the GI tract of humans and other species. They are also referred to as enteric bacilli or as coliforms.

Question 10

What type of infections commonly encounter the enterobacteriaceae?

Answer 10

urinary tract infections (E. coli, Klebsiella and others), gastrointestinal infections and food poisoning (Salmonella, Shigella, E. coli), bacteremia, sepsis and nosocomial pneumonia.

Question 11

What are important features of enterobacteriaceae?

Answer 11

• Large Gram negative rods
• Facultative anaerobes (capable of growth aerobically or anaerobically)
• Grow in the presence of bile salts (therefore suited to survival and growth in the GI tract)
• Grow on MacConkey media (see box below)
• Oxidase negative (see oxidase test – next page)
• Ferment carbohydrates (fermentation patterns are used in speciation)
• Lipopolysaccharide (endotoxin) in cell wall bears type-specific polysaccharide O chain

Question 12

What is an important test used in the preliminary classification of a GNB?

Answer 12

Lactose fermentation
Lactose fermentation can be easily detected by examining growth on MacConkey media. MacConkey media contains lactose as a source of energy. As lactose is metabolized, the pH of the media around the bacterial colony falls and a pH indicator in the media causes colonies to take on a pink colour.

Question 13

What are two important characteristics of a MacConkey media?

Answer 13

MacConkey media:
• Selective media – inhibits growth of gram positive bacilli
• Differential media – Lactose fermenters = PINK Non Lactose fermenters = WHITE

Question 14

where are the non-fermentative/environmental gram negative bacilli found?

Answer 14

The non-fermentative gram negative bacilli are environmental organisms. Many of these organisms are water-loving and contaminate aqueous environments in the hospital (nebulizer and ventilator equipment, irrigation solutions etc.)

Question 15

What are problems caused by the non-fermentative/environmental gram negative bacilli, who is most at danger of these?

Answer 15

They are important nosocomial / opportunistic pathogens – causing urinary tract infections, pneumonia, intravenous catheter associated bacteremias, and sepsis, mainly in hospitalized patients. Patients in Intensive Care Units and those receiving mechanical ventilation are particularly vulnerable to infection with these organisms.

Question 16

What is one common non-fermentative/environmental gram negative bacilli?

Answer 16

Pseudomonas spp.

Question 17

For pseudomonas spp. what is characteristic of them, what issues do they cause, what are the antibiotic choices for them?

Answer 17

Remember it is a non-fermentative/environmental gram negative bacilli.
• Some strains produce a slime layer that makes the organism very difficult for the immune system to eradicate. These slime producing isolates are common in patients with CF.
• Importance: lung infections in patients with Cystic Fibrosis, ventilator associated pneumonia, bacteremia in neutropenic patients, skin / graft infections in burn patients, urinary tract infections in patients with prolonged catheterization.
• Antiobiotic choices: aminoglycosides, ciprofloxacin, imipenem, piperacillin, ceftazidime

Question 18

What are some other pseudomonads (organisms closely related to pseudomonas)? what type of bacilli are they?

Answer 18

non-fermentative/environmental gram negative bacilli.

• Stenotrophomonas spp. — nosocomial and opportunistic pathogen, especially in patients treated with broad spectrum antibiotics. Highly antibiotic resistant.
• Burkholderia spp. — important pathogen in patients with advanced Cystic Fibrosis

Question 19

What are other important ones in the non-fermentative/environmental gram negative bacilli?

Answer 19

• Flavobacterium spp. — Sepsis and meningitis in sick neonates
• Acinetobacter spp.
• Alcaligenes spp.

Question 20

What is the oxidase test? Why is it important?

Answer 20

Oxidase test: a rapid test that is often used to differentiate various species of gram-negative bacilli and gram-negative cocci. A small amount of growth from a colony is rubbed on a strip of filter paper with the oxidase reagent on it. A purple colour will appear within a few seconds if the organism is oxidase positive

Question 21

What type of gram negative bacilli is oxidase negative? What are some important oxidase positive ones?

Answer 21

Enterobacteriaceae are OXIDASE NEGATIVE
Important OXIDASE POSITIVE organisms include:
•	Pseudomonas 
•	Pasturella
•	Campylobacter
•	Neisseria (GNC)

Question 22

What are the the group of organisms includes non-fermentative gram-negative bacilli with special growth requirements - either special growth factors or special atmospheric conditions.?

Answer 22

Fastidious Gram Negative Bacilli
They are important causes of disease in humans and may fail to be identified in the lab if their specific growth requirements are not considered and the correct conditions for growth not provided.

Question 23

What do fastidious gram negative bacilli look like on gram stain?

Answer 23

, some of these organisms are very small bacilli that are intermediate between cocci and bacilli - they are therefore often referred to as “cocco-bacilli”.

Question 24

What are three important fastidious gram negative bacilli?

Answer 24

Hemophilus, bordetella, legionella.

Question 25

What type of category of gram negative bacilli is hemophilus? What are its characteristics?

Answer 25

Hemophilus
• Require X (haemin) &/or V (NAD) factors (found in media containing heated blood: “Chocolate agar”)
• Present as part of normal respiratory flora of humans and many animal species
• Important cause of meningitis, sepsis, respiratory, and bone and joint infections in young children
• Important cause of respiratory infections in adult patients with COPD

Question 26

What type of fastidious gram negative bacilli is important cause of whoooping cough and other respiratory syndromes?

Answer 26

Bordetella

Question 27

What fastidious gram negative bacilli is an important cause of pneumonia in some areas, and lives in aquatic habitats such as air-conditioning cooling towers and hospital water systems?

Answer 27

legionella

Question 28

For fastidious gram negative bacilli, pasteurella multocida, brucella, and francisella tularensis, why are they specifically important? and what happens with these?

Answer 28

Three other genera are important causes of zoonotic infections.
Pasteurella multocida
• Infections due to animal bites
Brucella
• Infection results from ingestion, inoculation or inhalation of bacteria in animal products such as milk, meat, pelts, placental tissue etc.
Francisella tularensis
• Infection (“Tularemia”) acquired from tick bites, contact with rabbits & other species, usually hunters, campers
• Presentation is variable: fever, pneumonia, sepsis, skin ulceration with enlarged lymph nodes

Question 29

What are the HACEK organisms?

Answer 29

Other important species in this group are the HACEK organisms – a group of slow growing, fastidious gram-negative bacilli that are noted for their ability to cause insidious infection of heart valves (“subacute endocarditis”).
These organisms should be suspected in cases of “culture negative endocarditis” as they will not be recovered in culture unless specific techniques and prolonged incubation are used. These organisms may be found as commensals in the oropharynx. In most cases of endocarditis, patients have periodontal disease that is the source of the organism.

Haemophilus aphrophilus
Actinobacillus 
Cardiobacterium
Eikenella
Kingella

Question 30

What leads to most cases of endocarditis for the source of the HACEK organisms?

Answer 30

periodontal disease

Question 31

What are 4 types of curved gram negative bacilli?

Answer 31

Vibrios
Aeromonas
Campylobacter
Helicobacter

Question 32

What type of gram negative bacilli is vibrios and what characteristics does it have?

Answer 32

This group of organisms cause a variety of infections from cholera to skin and soft tissue infections. These organisms are found in aquatic environments contaminated by sewage. Infection occurs by ingestion of contaminated seafood or water, or through inoculation of wounds sustained in contaminated water.

Question 33

What type of gram negative bacilli is aeromonas and what characteristics does it have?

Answer 33

This fermentative gram-negative bacillus is related to the Vibrios (although not a curved bacillus). It is found in aquatic environments and can cause enteritis or severe soft tissue infections

Question 34

What curved gram negative bacilli includes a number of species responsible for human disease, most commonly enteritis (sometimes accompanied by bacteremia or deep tissue infections or followed by a reactive arthritis)? What is one syndrome that is a potential complication of infection with this bacilli?

Answer 34

Campylobacter – This genus includes a number of species responsible for human disease, most commonly enteritis. Occasionally, enteritis in the normal host is accompanied by bacteremia or deep tissue infections or followed by a reactive arthritis. In immunocompromised hosts, bacteremia or focal infections may occur without clinical enteritis. Guillain Barre Syndrome is a potential complication of infection with Campylobacter. Human infections are acquired through ingestion of contaminated food or water.

Question 35

What is the curved gram negative bacilli that is the leading cause of peptic ulcers and gastritis? It produces urease, which enables it to split urea to ammonia producing a local increase in the pH of the gastric mucosa, thus allowing the organism to survive the otherwise highly acidic environment of the stomach.

Answer 35

H. pylori is the leading cause of peptic ulcers and gastritis. H. pylori produces urease, which enables it to split urea to ammonia producing a local increase in the pH of the gastric mucosa, thus allowing the organism to survive the otherwise highly acidic environment of the stomach.