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2.2 Microbiology > Denise - Bacillus > Flashcards

Denise - Bacillus Flashcards

1
Q

Are bacillus endospore formers or not

A

They are endospore formers

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2
Q

What type of bacteria are bacilli

A

Gram positive bacilli
Aerobic
Endospore formers

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3
Q

What are the three bacillus species of interest

A

Bacillus cereus
Bacillus thurgenesis
Bacillus anthracis

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4
Q

How many Bacillus species are there

A

over 200 species identified

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5
Q

What infections does B. cereus cause

A

Extensively found in food and food processing environments

Opportunistic pathogen in other sites

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6
Q

Where is B. thuringenesis found

A

Insecticide

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7
Q

Where is B. anthracis found

A

Its a virulent pathogen of mammals

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8
Q

Where are all three Bacillus species found

A

Isolated from soil species

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9
Q

What is the B. cereus group

A

B. cereus sensu lato

Genetically highly homogenous

A subdivision of the genus Bacillus

Six recognised species

Closely related by genome sequence

Same phylogenetic cluster

Differentiated by phenotypic characteristics and pathological properties

Many distinguishing features between the species are encoded by genes located on plasmids

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10
Q

How are the B cereus group species similar to each other
(5)

A

B. anthracis, B. cereus, B. thuringiensis

Closely related by genome sequence

Same phylogenetic cluster

Differentiated by phenotypic characteristics and pathological properties

Many distinguishing features between the species are encoded by genes located on plasmids

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11
Q

Write about the clinical significance of Bacillus
(4)

A

Most species have little pathogenic potential

Opportunistic pathogens

Two species are clinically important (B. anthracis, B. cereus)

Major problem with contamination in the Lab

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12
Q

Write about B. cereus
(7)

A

Gram positive
Facultative anaerobe
Endospore former
Distinct colonial morphology
Large rods
Spores do not swell the sporangium
Most strains are motile and B-haemolytic

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13
Q

Write about the endospore forming aspect of B. cereus

A

Form endospores within a few days on commonly used agar media

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14
Q

What is the distinct colonial morphology of B. cereus

A

Ground glass appearance
Cells are large rods

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15
Q

Write about B. cereus reservoirs
(5)

A

Ubiquitous
Soils, sediments, dust and plants
Spores can be passively spread and can be found outside natural habitats
Germinates, grows and sporulates in soil, thus demonstrating a saprophytic life cycle
Has been reported to be present in stools of healthy humans at varying levels

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16
Q

Write about soil to food transmission of B. cerueus

A

Isolated from a range of different foods
Cross-contamination can distribute spores or cells to other foods, such as meat products
At harvest cells or spores may accompany plant material into food production areas and establish on food-processing equipment

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17
Q

What food is B. cereus found in
(5)

A

Rice
Dairy products
Spices
Dried foods
Vegetables

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18
Q

Write about B.cereus endospores
(7)

A

Heat treatment
Food processing
Sanitisers
Spore can adhere to surfaces in the food processing lines
Not removed by regular cleaning of surfaces
Role in biofilm persistence in food industry equipment such as dairy pipelines
The biofilm protects spores and vegetative cells against inactivation by sanitisers

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19
Q

Write about B. cereus epidemiology
(5)

A

Notifiable to HPSC
About 1 case a year
Under estimated
No outbreaks in Ireland
Outbreaks reported in neonatal units in UK and France

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20
Q

Write about B. cereus foodborne disease
(5)

A

Two distinct foodborne disease types, emetic and diarrhoeal
Mild and self-limiting
B. cereus was established as an organism of foodborne disease in the 1950s
Outbreaks of the diarrhoeal type of disease in hospitals in Norway in 1947
The emetic syndrome was first identified after several outbreaks caused by eating cooked rice in the UK in early 1970s

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21
Q

Write about B. cereus food association
(6)

A

Wide environmental distribution of spores allows B. cereus access to a variety of foods

Proteinaceous foods associated with the diarrhoeal disease

The emetic syndrome is associated with starchy foods

No legislation requires screening of food items for this pathogen contamination

Dried infant formulae have an established maximum limit of 50 CFU/g

Outbreaks on neonatal units

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22
Q

What is the B. cereus diarrhoeal disease associated with

A

Proteinaceous foods:
- meat
- stews
- sauces
- milk
- vegetables
- fish

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23
Q

What foods are associated with the B. cereus emetic syndrome

A

Starchy foods:
- rice products
- potato
- pasta
- noodles
- pastry products

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24
Q

Write about the B. cereus diarrhoeal syndrome
(7)

A

Toxicoinfection
Vegetative cells, ingested as viable cells or spores producing protein enterotoxins in the small intestine
Heat-labile enterotoxin
Diarrhoeagenic necrotising enterotoxin
Onset of watery diarrhoea and cramps within 6-15 hours post consumption of contaminated food
Incubation time is over 6 hours with a normal range between 8-16 hours
Duration of symptoms is 12-24 hours

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25
What is the B. cereus diarrhoeal syndrome toxin
Diarrhoeagenic necrotising enterotoxin Its a heat-labile enterotoxin
26
What is the incubation time for B. cereus diarrhoeal syndrome
6 hours
27
How long does B. cereus diarrhoeal syndrome last
12-24 hours
28
What are the three toxins associated with B, cereus diarrhoea
HBL (Haemolysin) NHE (Non Haemolytic Entertoxin) CytK
29
Write about HBL (Haemolysin) (5)
Tripartite-3 component toxin (L2, L1 and B) B is for binding, L1 and L2 are lytic components Cytotoxic and haemolytic 55% of all B. cereus strains Tertiary structure resemblance with the pore-forming toxin cytolysin A (ClyA)
30
Write about the genetics behind Tripartite-3 component toxins (L2, L1 and B)
Encoded by the genes hblC, hblD and hblA Cotranscribed from one operon
31
What does HBL (Haemolysin) actually do
Its dermonecrotic Vascular permeability Fluid accumulation in rabbit ileal loops Cytotoxic and haemolytic
32
What percentage of B. cereus strains produce HBL (Haemolysin)
55%
33
What does HBL (Hameolysin) resemble
Forming toxin cytolysin A (ClyA)
34
What is NHE
Non Haemolytic Enterotoxin
35
Write about Non Haemolytic Entertoxin (3)
Large food poisoning outbreak in Norway in 1995 caused by the hbl-negative B. cereus strain Produced by 90% of all B. cereus strains NHE is a three-component toxin with cytotoxic activity towards epithelial cells
36
What percentage of B. cereus strains express Non Haemolytic Enterotoxin?
90% of all B. cereus strains
37
What does Non Haemolytic Entertoxin cause
Rapid disruption of the plasma membrane Formation of pores in the lipid bilayers
38
What are the two B. cereus Beta barrel pore-forming toxins
CytK and Hlyll
39
Write about the B. cereus Beta Barrel pore-forming toxins, CytK and Hlyll (4)
Both single component protein toxins This toxin family includes B-toxin of C. perfringens and alpha-haemolysin of S. aureus Transmembrane pore at target cell surface CytK is a single 34 kDa protein with dermonecrotic, cytotoxic and haemolytic activities
40
What does CytK do?
Dermonecrotic Cytotoxic Haemolytic
41
What is emetic syndrome
Vomiting
42
Write about emetic syndrome (3)
Outbreaks after eating cooked rice in the UK in the 1970s Intoxication - ingestion of a heat stable toxin (emetic toxin "cerulide") produced in food before ingestion Nausea and vomiting within 0.5-6 hours after consumption of food: Rapid onset of symptoms
43
What toxin is responsible for emetic syndrome?
Emetic toxin "cerulide"
44
How long does it take for onset of symptoms for emetic syndrome
0.5 - 6 hours after consumption of food
45
Write about emetic toxin (cerulide) (6)
Ring structure of 3 repeats of 4 amino acids Produced by a nonribosomal peptide synthetase Encoded by the 24-kb cereulide synthetase (ces) gene cluster Plasmid Mechanism unkown -> receptor mediated Resistant to autoclaving, pH and protease
46
What is emetic toxin resistant to? (3)
Autoclaving pH Protease
47
Write about B. cereus oportunistic infections (6)
Local infection Bacteraemia and septicaemia Central nervous system infections Respiratory infections Endocarditis (in drug abusers) and pericarditis Immunocompromised patients Neonates
48
What local infections does opportunistic B. cereus cause (2)
Infections particularly of burns, traumatic or post-surgical wounds and of eye Sever keratitis, endophthalmitis and panopthalmitis
49
What central nervous system infections does B. cereus cause
Meningitis Abscesses Shunt-associated infections
50
When might endocarditis by B. cereus be seen
In drug abusers
51
Write about B. anthrax epidemiology (6)
Zoonotic disease Exposure to animals (sheep, goats, horses) Exposure to animal products Difficult to eradicate from endemic areas due to endospores Cutaneous anthrax account for 95-99% of human cases worldwide <1-20% mortality rate; 1-2 cases per year USA
52
What is the mortality rate of B. anthrax
<1-20% But there is only 1-2 cases per year USA
53
Write about anthrax infections (5)
Incubation period of 2-5 days Small pimple or papule develops over 24 hours The lesion is surrounded by oedema May be necrosis Not purulent
54
Describe how anthrax infections develop (4)
Small pimple or papule develops A ring of vesicles develops around the papule Ulcerates, dries and blackens into a characteristic eschar This enlarges and becomes thick and adherent to underlying tissue over the ensuing weeks
55
Write about anthrax infection severity (4)
Secondary sequelae to cutaneous anthrax include cellulitis and meningitis About 20% of untreated cases are fatal The mortality rate drops to zero if patients are treated
56
Write about B. anthrax cutaneous anthrax (6)
Endospores enter through a cut, they germinate and vegetative cells multiple Acute host inflammatory response - capsule prevents phagocytosis Exotoxin release Invasive damage Rapid development of a malignant pustule Associated with cases where humans handle infected animals and/or animal products
57
Write about intestinal anthrax
Cutaneous anthrax on intestinal mucosa It is sometimes accompanied by gastroenteritis Low level germination at one site leads to massive effusion, mucosal oedema and necrotic lessions
58
Write about pulmonary anthrax
Endospores are carried from the lungs by macrophages to the lymphatic system where they germinate and multiply to produce a fatal septicaemia
59
Write about Bacillus bioterrorism
2011 Sent through US Postal Service
60
Write about B. anthracis - Virulence: Capsule (6)
B. anthracis produces a number of virulence factors The poly D glutamic acid capsule - antiphagocytic - essential for virulence Plasmid pXO2 caosule
61
What are the two B. anthrax toxins associated with virulence
Oedema factor -> increases CAMP Lethal factor -> metalloprotease tissue necrosis
62
Write about the 2 plasmid encoded exotoxins, oedema factor and lethal factor
Both are AB toxins (activity Binding) The binding unit is shared by both toxins and is called a protective antigen Mediated cell entry of both toxins pXO1 (anthrax toxin)
63
What are some safety considerations for laboratory investigation behind Bacillus species
B. anthracis is a Class 3 pathogen Category 3 Lab facilities are required
64
What clinical samples is B. anthracis seen in? (3)
Cutaneous (95%) -> tissue Gastrointestinal (5%) -> faeces Inhalation (<1%) - Lung aspirates, pleural fluid, tissue samples, blood, CSF
65
How is B. anthraxis inoculated
Blood agar PLET agar Incubates at 37 degrees for 24-48 hours
66
What is PLET agar
Polymuxin-lysozyme-EDTA- thallous acetate agar
67
What samples might be seen for B. cereus
Numerous samples Environmental samples Food
68
What media is B. cereus isolated on
Isolation media Blood agar Selective agar - Bacillus cereus selective agar (PEMBA)
69
Write about B. cereus on PEMBA agar
PEMBA is selective and differential B. cereus does not ferment mannitol B. cereus produces lecithinase B. cereus is resistant to polymixin Colonies on PEMBA agar have peacock blue colonies Cannot differentiate between B. cereus and thuringenesis
70
What is the basic characterisation for Bacillus
Gram positive baciili (large box car ends Central spores) KoH negative Catalase positive Oxidase negative
71
What confirmatory tests are put up on day 2 and 3 for Bacillus species
Biochemical identification Maldi: Not Disc enough Toxin detection
72
Write about the detection of B. cereus emetic toxin (5)
Cell culture assays using Hep-2 cells These lack specificity HPLC-MS analysis can be done Maldi Tof MS Real time PCR ces genes
73
What is meant by cell culture assays using Hep-2 cells to detect B. cereus emetic toxin
Using Hep 2 cell lines with culture filtrates Cause vacuolization of this cell line Toxin induced cell damage
74
Why is HPLC-MS not really used for emetic toxin detection
Laborious and costly
75
Write about detection of B. cereus cytotoxin NHE
TECRA-BDE kit is used
76
What is TECRA -BDE kit
TECRA bacillus diarrhoeal enterotoxin visual immunoassay Detects the NheA component of the Nhe toxin by an ELISA sandwich testT The sensitivity reported by the manufacturer is >1 ng mL prepared sample The kit is intended for use on foods and environmental samples
77
Write about detection of B. cereus cytotoxin HBL detection
The Oxoid BCET-RPLA kit is used
78
What is the Oxoid BCET-RPLA kit
Semi quantitative Detects by reversed antibody agglutination The L2 component of Hbl in foods and in culutres of B. cereus The kit is intended for use on foods and environmental samples
79
What are the Duopath Cereus Enterotoxins: HBL and NHE
Immunochromatographic rapid test based on gold labelled antibodies
80
How are B. anthracis toxins detected
PCR ELISA to detect in serum or pleural fluid
81
Write about the treatment and prevention of anthrax
Vaccination for workers at high risk of exposure Ciprofloxacin for cutaneous Ciprofloxacin and rifampicin and vancomycin for inhalation

2.2 Microbiology (60 decks)

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