Clostridium species - Microbiology
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Transcript of Clostridium species - Microbiology
College Of Dentistry - Mosul University
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CLOSTRIDIUM SPECIES
Done By: Rami Bassam
3rd StageGroup – B-
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Clostridium
Gram-positive . obligate anaerobes capable of producing endospores which protect them in harmful environment . Individual cells are rod shaped.
The spores are usually wider than the rods, and are located terminally or sub terminally.
Most clostridia are motile by peritrichous flagella.while others have a capsule like
Clostridium.perfringens
Characteristics
Light Microscope Clostridium .tetani
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Most Common Species
Clostridium consists of around 100 species that include common free-living bacteria as well as important pathogens There are five main species
responsible for disease in humans.C. perfringens: gas gangrene; food poisoningC. tetani: tetanusC. botulinum: botulismC. difficile: pseudomembranous colitisC.Sordellii : can cause a fatal infection in exceptionally rare cases after medical abortions
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SHAPES OF CLOSTIDIA The shape an position of spores varies in different species and is useful the identification of Clostridia *Central in Cl.bifermentans *Sub terminal in Cl.perfringens *Oval or terminal in Cl.tertium*Spherical and terminal giving drum stick appearance in Cl.tetani
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C. perfringens C. perfringens is a
relatively large Gram-positive short fat bacilli with blunt ends. It is capsulate and non-motile. Anaerobic. It grows quickly on laboratory media on blood agar ( B – Haemolytic )
Clostridium . PerfringensIn L.M
Clostridium . PerfringensIn E.M
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Media used for Cultivation
Liquid medium for cultivation cooked meat broth
Thiglyclolate broth
CMB contain unsaturated fatty acids which take up oxygen
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Clostridium. perfringens Culture & identification
1 =Control, 2 and 3= "Stormy clot fermentation“
Clostridium perfringens will produce acid and gases
B – HaemolysisOn blood agar media
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Litmus Milk TestIt distinguish between
different species of bacteria. The lactose (milk sugar), litmus (pH indicator), and casein(milk protein) contained within the medium can all be metabolized by different types of bacteria.
Milk is the first substrate used to maintain bacteria, this test allows for accurate depiction of bacterial types. The addition of litmus, other than explaining the pH type, acts as an oxidation-reduction indicator. The test itself tells whether the bacterium can ferment lactose, reduce litmus, form clots, form gas ,
Cl.perfringensStormy clot reaction
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Nagler’s Reaction(Lecithinase test )
This test is done to detect the lecithinase activity The M.O is streaked on the medium containing
egg yolk (contains lecithin) The plate is incubated anaerobically at 37 C
for 24 h Colonies of Cl. perfringens are surrounded by
zones of turbidity due to lecithinase activity and the effect is specifically inhibited if Cl. perfringens antiserum containing antitoxin is present on the medium
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C. perfringens Nagler Reaction
NOTE: Lecithinase (α-toxin; phospholipase) hydrolyzes phospholipids in egg-yolk agar around streak on right.
Antibody against α-toxin inhibits activity around left streak.
Nagler Reaction
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Positive Nagler ReactionProcedure of Nagler Reaction
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Specific Requirements for Anaerobic Cultivation
Anaerobic Jar
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Cl.perfringens Produce diseases
The organisms associated with gas gangrene attack soft tissues by producing toxins and aggressins, and some strains of the bacteria produce enterotoxins and cause food poisoning
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Collecting a Infected Tissues
If there are pieces of necrotic tissue in the wound, small pieces should be transferred aseptically into a sterile screw-capped bottle and examined immediately by microscopy and culture. Specimens of exudate should be taken from the deeper areas of the wound where the infection seems to be most pronounced.
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Virulence Factors Virulence factors
toxins – alpha toxin – causes RBC rupture, edema and tissue destruction
Enterotoxin collagenaseHyaluronidaseDNase
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Clostridium tetani Anaerobic bacteria of the genus species Clostridium it
is gram positive, slender bacillus and it has spherical terminal spores giving drum stick appearance
It is non capsulated & motile with peritrichus flagella
It produces a potent biological toxin, tetanospasmin, and is the causative agent of tetanus a disease
characterized by painful muscular spasms that can lead to respiratory failure and, in up to 40% of cases,
death.
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What is Tetanus? An infectious disease caused by
contamination of wounds from the bacteria Clostridium tetani, or the spores they produce that live in the soil, and animal feces
Infection follows when spores become activated and develop into gram-positive bacteria that multiply and produce a very powerful toxin
(tetanospasmin) that affects the muscles.
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Causes
Tetanus spores are found throughout the environment, usually in soil, dust, and animal waste.
Tetanus is acquired through contact with the environment; it is not transmitted from person to person.
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Causes The usual locations for the bacteria to enter the
body:
Puncture wounds (such as those caused by rusty nails, splinters, or insect bites.)
Burns, any break in the skin, and IV drug access sites are also potential entryways for the bacteria.
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Mode of action of tetanospasmin
1 .It inhibits the release of acetylcholine thus it interferes with neuromuscular transmission .
2 .Inhibition of postsynaptic spinal
neurons by blocking the release of an inhibiting mediator
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Laboratory Diagnosis:
Gram +ve stains grow on blood agar media aerobically
Inoculation of culture in to 2 mice one is protected with anti-toxin and the other is unprotected (dies with typical tetanic spasms )
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Clostridium difficile
Clostridium difficile " C. diff", is a
species of Gram-positive bacteria of the
genus Clostridium that causes
diarrhea and other intestinal disease when competing
bacteria are wiped out by antibiotics.
Most common cause of nosocomial diarrhea.
Rate and severity of C. difficile-associated diarrhea (CDAD) increasing.
Clostridium difficile is a bacterium that can cause symptoms ranging from diarrhea to life-threatening inflammation of the colon.
Illness from C. difficile most commonly affects older adults in hospitals or in long
term care facilities and typically occurs after use of antibiotic medication
characteristic features:
Clostridia are anaerobic, spore-
forming rods (bacilli) .
Clostridia
habitantC. difficile bacteria can be found
throughout the environment — in soil, air, water, and human and animal feces. A
small number of healthy people naturally carry the bacteria in their large intestine.
But C. difficile is most common in hospitals and other health care facilities,
where a much higher percentage of people carry the bacteria.
Reservoir
Infectious AgentC.difficile
Means of Transmission
Portal of entry
Susceptible Host
Chain of infection Bowel and
Contaminated environment
Contact transmission from contaminated
hands,equipment or the
environmentFaecal/Oral
>65 years History of antibiotic use
Recent received healthcare
Underlying conditions Abdominal surgery
Weakened immunity
PathogenesisDisruption of
normal colonic flora
Colonisation with C. difficile
Production of toxin A +/- B
Mucosal injury and inflammation
Toxin production is cause of Pathogenesis
Toxigenic strains produce 2 major
toxins:toxin A
(enterotoxin)toxin B
(cytotoxin)Neutralised by C.
sordellii antitoxin
Signs and symptomsWatery diarrhea three or more times a day for two
or more daysMild abdominal cramping and tendernessWatery diarrhea 10 to 15 times a dayAbdominal cramping and pain, which may be
severeFeverBlood or pus in the stoolNauseaDehydrationLoss of appetiteWeight loss
Complications of CDADPseudomembraneous colitis
Perforation of the colon
Sepsis
Death
The SpecimenFresh is best (test within 2 hours)Liquid or loose, not solidIf unable to test within 2 hours, refrigerate at 4°C for up to 3 daysFreeze at -70°C (not -20°C) if testing will be delayedSpecimen quality will influence test results
Diagnosis of CDADEndoscopy
(pseudomembranous colitis)
CultureCell culture
cytotoxin testEIA toxin testPCR toxin gene
detection
ENDOSCOPY PICTURE
preventation
Hand washing
Contact precautions
Thorough cleaning
Perform Hand Hygiene after removing gloves.
Note : alcohol does not kill C. difficile spores, use of soap and water is more
efficacious
Clostridium Botulinum
Characteristic
Gram positiveObligate anaerobic bacillusSpores
Resistant to heat, light, drying and radiationSpecific conditions for germination
Anaerobic conditionsWarmth (10-50oC)Mild alkalinity
Transmission
IngestionOrganismSporesNeurotoxin
Wound contaminationInhalationPerson-to-person not documented
HUMAN DISEASEThree formsFoodborneWoundInfant
All forms fatal and a medical emergencyIncubation period: 12-36 hours
Diagnosis
The symptoms of botulism are similar to those of Guillain-Barré syndrome,
stroke, and myasthenia gravis.As a result, botulism is probably
substantially under-diagnosed.Serum electrolytes, renal and liver
function tests, complete blood tests, urinalysis, and electrocardiograms will
all be normal unless secondary complications occur.
The incubation period varies according to the mode of transmission, rate of
absorption of the toxin, and the total amount and type of toxin.
Foodborne botulism usually takes 24-36 hours to manifest itself.
Wound botulism often takes 3 or more days to appear.
Inhalation botulism has occurred very rarely, but incubation times may range
from several hours to perhaps days, again depending upon the type and
amount of toxin inhaled.
All four types of botulism result in symmetric descending flaccid paralysis of motor and autonomic nerves always
beginning with the cranial nerves. These symptoms are preceded by
constipation in cases of infant botulism.
Symptoms include:Double or blurred visionDrooping eyelidsDry mouthDifficulty SwallowingMuscle weakness
If left untreated symptoms may expand to include paralysis of
respiratory muscles as well as the arms and legs.
Asphyxiation due to respiratory paralysis is the most common cause of
death in botulism cases.
Prevention
Proper food preparation is one of the most effective ways to limit the risk of
exposure to botulism toxin.Boiling food or water for ten minutes can
eliminate some strains of Clostridium botulinum as well as neutralize the toxin
as well. However, this will not assure 100% elimination.
Limiting growth of Clostridium botulinum and the production of botulism toxin is an
alternative to their outright destruction.
Medical uses of BoNT
Now manufactured under the name “Botox”
Experimentally used for treating migraine headaches, chronic low back
pain, stroke, cerebral palsy, and dystonias (neurologic diseases
involving abnormal muscle posture and tension)
Frequent injections allows an individual to develop antibodies
Studies carried out to determine feasibility of other strains of BoNT
BoNT B manufactured for treatment of cervical dystonia in 2000 as “Myobloc”
BoNT A (Botox)
Botox injection patient 13 weeks after injection
Sadick, N. and A.R. Herman (2003). “Comparison of Botulinum Toxins A and B in the Aesthetic Treatment of Facial Rhytides.” Dermatologic Surgery 29:340-347.
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