Week 6 Flashcards
GENERAL CHARACTERISTICS Bacillus
- Gram-positive or gram-variable, straight, round- or square-ended
rods, occur singly or in chains (range from few to many cells in
length) - motile via peritrichous flagella with exceptions of Bacillus anthracisand Bacillus mycoides
- produce endospores, 1 per mother cell (sporangium) in presence of oxygen
- depending on species endospores may be: −cylindrical, oval, round or kidney-shaped
−centrally, subterminally or terminally placed
−characteristically swelling or not swelling sporangium - aerobic or facultatively anaerobic
- diversity of metabolic types and nutritional requirements: − psychrophiles, mesophiles (majority), thermophiles −alkalophilic, neutrophilic, acidophilic
- some species are producers of antibiotics
ENDOSPORES Bacillus
- spore - dormant structure capable of surviving for prolonged
periods - endowed with capacity to re-establish vegetative stage of growth under appropriate environmental conditions
- at some point in vegetative cell cycle of spore-forming organisms growth arrested
- cell undergoes progressive changes that results in formation of
endospore - endospore formation occurs during stationary phase of growth after depletion of certain nutrients in culture medium or environment
- single spore produced within vegetative cell
- differs from parent cell in its:
− morphology
−composition
−increased resistance to adverse environments −absence of detectable metabolic activity - resistance to radiation, drying, toxic chemicals depends on cystinerich, keratin-like spore coat proteins
- thermal resistance attributed to very low water content of
protoplast, which renders proteins and nucleic acids more resistant to denaturation - process of spore germination consists of 3 sequential phases:
−activation stage - conditions spore to germinate in suitable
environment
− germination stage - characteristic properties of dormant
spore lost
− outgrowth stage - spore converted into new vegetative cell
SPORULATION Bacillus
- Spore septum begins to isolate newly replicated DNA and a small portion of cytoplasm
- Plasma membrane starts to surround DNA, cytoplasm, and membrane isolated in step 1
- Spore septum surrounds isolated portion, forming forespores
- peptidoglycan layer forms between membranes
- Spore coat forms
- endospore is freed from cell
NATURAL HABITAT Bacillus
- widely distributed in nature; saprophytes in soil, dust, water, on materials of plant and animal origins
- part of normal intestinal microbiota of humans and animals
- majority of Bacillus spp. have little or no pathogenic potential,
rarely associated with disease - some species (e.g., Bacillus anthracis, Bacillus cereus, Bacillus subtilis) may cause human and animal infections
- world-wide in distribution
VIRULENCE FACTORS - BACILLUS ANTHRACIS
- capsular polypeptide
− D-glutamic acid
−interferes with phagocytosis - anthrax toxin
−complex exotoxin consisting of 3 protein components: - protective antigen - PA
- lethal factor - LF
- oedema factor - EF
− none have any toxic activities by themselves, acting
synergistically produce systemic effects of anthrax
−inhibit phagocytosis and intracellular signal transduction
resulting in inactivation of transcriptional factors in cell
nucleus - production of both virulence factors enhanced by CO2
CLINICAL SIGNIFICANCE - BACILLUS ANTHRACIS
- causative agent of anthrax
- exploited and developed as agent of biological warfare by
several countries - biological agent associated with bioterrorism
- anthrax primarily disease of herbivorous animals
- humans accidentally encounter disease by direct contact with
certain animal products (hair, wool, bones)
Anthrax in animals
* severe, usually takes form of septicaemia
* infected animal remains free of symptoms until few hours before death
* mortality rate about 80%
Anthrax in humans
* humans can become infected via one of 3 possible routes:
− Cutaneous route
−Inhalation
−Ingestion
Cutaneous anthrax (malignant pustule)
* most common form (95% to 99% of all cases world-wide)
* organisms gain access through small abrasions or cuts, multiply locally
* incubation period 2 to 3 days
* at site of inoculation small, pruritic, non-painful papule, develops into haemorrhagic vesicle
* vesicle ruptures, leading to slow healing painless ulcer covered with black eschar surrounded by oedema
* may spread to lymphatics, may be regional adenopathy
* 20% mortality rate in untreated cases
Pulmonary anthrax (wool-sorter’s disease)
* acquired by inhalation of spores
* symptoms those of respiratory infection, fever, malaise, myalgia, unproductive cough
* rapidly progresses to very severe infection with marked respiratory distress, cyanosis, haemorrhagic necrosis of mediastinum* 80% to 90% mortality rate in untreated cases
Oropharyngeal anthrax and Gastrointestinal anthrax
* acquired by ingestion of organism
* infections associated with cervical and oral pain and oedema,
and nausea, vomiting and diarrhoea, respectively
* occasionally there is loss of blood (either through
haematemesis or in stools)
* like pulmonary form, there is a high mortality rate if untreated* in all forms, there may be invasion of bloodstream and
localisation in meninges
* anthrax infection in humans provides permanent immunity
CLINICAL SIGNIFICANCE - BACILLUS CEREUS
Emetic or vomiting syndrome
* short incubation period of 1 to 6 hours
* characterised by severe nausea, vomiting, abdominal cramping* frequently mistaken for staphylococcal food poisoning
* patients recover after 10 to 24 hours after onset
* associated with fried Oriental rice dishes, cream and milk products, pastas, reconstituted infant formula
Diarrhoeal syndrome
* longer incubation period of 8 to 16 hours
* characterised by nausea, abdominal cramping, profuse watery diarrhoea, tenesmus
* commonly confused with clostridial food poisoning
* patients recover after 12 to 36 hours after onset
* associated with meat, vegetable dishes, cakes, sauces, dairy products
- in patients with underlying diseases or immunosupressed patients causes:
− bacteraemia
− meningitis
−endocarditis
− necrotising pneumonia
− peritonitis
− osteomyelitis
COLLECTION, TRANSPORT, AND STORAGE OF SPECIMENS Bacillus
Anthrax
* specimens in cutaneous infections:
−swab samples of serous fluid of vesicles
− material beneath edge of black eschar
−3 sets of blood cultures
* specimens in pulmonary infections:
−sputum sample
−3 sets of blood cultures
* specimens in gastrointestinal infections:
− gastric aspirates
−faeces
−food
−3 sets of blood cultures
Bacillus cereus food poisoning
* isolation and identification of organism from stools insufficient
evidence as bacterium may be present in stools of healthy persons
* epidemiologically implicated food should be collected
CULTURE MEDIA Bacillus
Selective media
* Polymyxin-Lysozyme-EDTA-Thallous acetate (PLET) agar medium− best selective medium for Bacillus anthracis
* Phenylethyl alcohol (PEA) blood agar medium
Non-selective media
* 5% sheep blood agar medium
* Nutrient agar medium
INCUBATION CONDITIONS Bacillus
- 35oC to 37oC
- for 18 to 24 hours
- in ambient air
ISOLATION PROCEDURES Bacillus
Bacillus anthracis
* colonies non-haemolytic, white to grey, large (4mm to 5mm in
diameter), with ground-glass appearance
* usually rough, flat with many comma-shaped outgrowths of long filamentous chains of bacilli (“Medusa head” appearance)
* when lifted with inoculating needle edge of colony stands up
like egg white - “tenacity”
* poor filiform growth followed by outgrowth of delicate lateral
extensions
DIRECT EXAMINATION Bacillus
- Gram-stained smears
− of infected tissue or fluid - Fluorescent antibody stain
−for encapsulated strains of Bacillus anthracis - M’Fadyean-stained smears − of infected tissue, blood, fluid
− polychrome methylene blue stain used to reveal capsules - PCR assay
IDENTIFICATION Bacillus
- Gram stain and Spore stain of colony
- Catalase production (positive)
- Motility Test (Motile)
- Lecithinase Test (positive)
- Colony morphology
- MALDI-TOF MS
- Growth on PEA
SEROLOGIC TESTS Bacillus anthracis
Bacillus anthracis
* Indirect Haemagglutination (IHA) Test
* Enzyme-Linked Immunosorbent Assay (ELISA)
ANTIBIOTIC SUSCEPTIBILITIES Bacillus
Bacillus anthracis
* drug of choice is penicillin
* alternative drugs for penicillin-sensitive patients are ciprofloxacin, gentamicin, erythromycin
Bacillus cereus
* because of β-lactamase production, it is resistant to penicillin,
ampicillin, cephalosporins
* drug of choice is clindamycin
IMMUNISATION - BACILLUS ANTHRACIS
- active immunisation only known method of preventing anthrax in
herbivorous animals −living spore vaccine − derived from non-encapsulated strain of Bacillus anthracis(Sterne strain) - for protection of humans in high-risk work situations
− non-living vaccine
−consisting of aluminum hydroxide-adsorbed supernatant material from fermentor cultures of toxigenic but non-encapsulated strain of Bacillus anthracis
GENERAL CHARACTERISTICS Listeria
- Gram-positive coccobacilli, occur singly, in short chains or in palisade arrangement; may be filamentous in older cultures* motile at room temperature by means of 1 to 5 peritrichous flagella, do not form spores
- facultatively anaerobic, facultative intracellular organisms
- oxidase negative, catalase and aesculin positive
- Voges-Proskauer and methyl red positive
- urease, indole, and H2S negative
NATURAL HABITAT Listeria
- widely distributed in environment, isolated from soil, decaying
vegetable matter, silage, sewage, water, animal feed, meats, milk* part of normal microbiota of many mammals and birds - 1% to 15% of healthy humans are asymptomatic intestinal carriers
VIRULENCE FACTORS - SOLUBLE PRODUCTS Listeria
- Actin-assembly-inducing protein (ActA)
- Phospholipases C (PLCs)
- Zinc metalloproteinase (Mpl)
- Listeriolysin O (LLO)
VIRULENCE FACTORS - SURFACE
COMPONENTS Listeria
- Internalin A and internalin B (InlA and InlB)
- Endotoxin-like material
CELL CYCLE OF LISTERIAMONOCYTOGENES
- Listeria monocytogenes - facultative intracellular organism* invades and grows in variety of mammalian cells: macrophages, epithelial cells, fibroblasts
- subsequent to internalisation, organism escapes from host vacuole* bacterium undergoes rapid division in cytoplasm of host cell
- organism becomes encapsulated by short, host actin filaments* these filaments reorganise into long tail extending from only one end of bacterium
- tail mediates movement of organism through cytoplasm to surface of host cell
- at periphery, formed protrusions penetrate neighbouring cells
CLINICAL SIGNIFICANCE Listeria
- Listeria monocytogenes causes: spontaneous abortions in animals, basilar meningitis - “circling disease” in sheep
- causative agent of human listeriosis
- occur as sporadic cases or epidemics
- contaminated foods primary vehicles of transmission (raw vegetables, dairy products, meats)
- organism survives and multiplies:
− at refrigerator temperature
− in wide range of pHs - human-to-human transmission - infection of foetus
transplacentally - exhibits striking tropism for: foetus, placenta, CNS
ADULT INFECTIONS Listeria
- meningitis
− most common form of listeriosis (55% of cases) - septicaemia
− affects patients under 50 years of age
− 25% of cases - encephalitis
− 6% of cases - focal infections
− occur after episode of bacteraemia
NEONATAL INFECTIONS Listeria
Early type - granulomatosis infantiseptica
* results from infection in utero
* appears within 2 days of birth
* symptoms of septicaemia followed by foetal distress, pneumonia, diarrhoea, seizures, maculopapular skin lesions
* very high mortality rate (54% to 90%)
Late type - meningitis
* infection acquired either during or after birth rather than in utero* appears after 5th day of life
