Professional Documents
Culture Documents
Dr.T.V.Rao MD
Dr.T.V.Rao MD
Genus Clostridium
In Anaerobic spore bearing Gram positive bacilli Spores are wider than the body giving spindle shape
meaning spindle
Dr.T.V.Rao MD 2
Anaerobic. Large gram-positive rods. The spores are usually wider than the rods, and are located terminally or sub terminally. Most clostridia are motile by peritrichous flagella.
Dr.T.V.Rao MD 3
Clostridia
Large Gram positive bacilli. Straight or slightly curved rods with slightly rounded ends Anaerobic bacilli Spore bearing Spore do not germinate and growth does not normally proceed unless a suitably low redox potential Eh exists
SHAPES OF CLOSTIDIA
The shape an position of spores varies in different species and is useful the identification of Clostridia Central and equatorial in Cl.bifermentans Sub terminal in C.perfringens Oval or terminal in Cl.tertium Spherical and terminal giving drum stick in C.tetani
Dr.T.V.Rao MD
Clostridia
Clostridia are biochemically active, frequently possessing both saccharolytic and proteolytic properties, although in varying degrees. Many species are highly toxigenic. The toxins produced by the organisms of tetanus and botulism attack nervous pathways and are referred to as neurotoxins.
Dr.T.V.Rao MD 7
Gas gangrene
Dr.T.V.Rao MD
Dr.T.V.Rao MD
Clostridium perfringens
Large Gram-positive bacilli with stubby ends Capsulated Non motile Anaerobic Grown quickly on selective media Can be identified by Nagler reaction
Dr.T.V.Rao MD 10
C. perfringens
produce enterotoxins.
Dr.T.V.Rao MD 11
C. perfringens
C. perfringens is a relatively large Grampositive bacillus (about 46 1 m) with blunt ends. It is capsulate and non-motile. It grows quickly on laboratory media, particularly at high temperatures (approximately 42C), when the doubling time can be as short as 8 min. I
Dr.T.V.Rao MD 12
Saccharolytic organisms Cl. perfringens, Cl. septicum Ferment carbohydrates Acid and gas are produced
Proteolytic organisms Cl. sporogenes Digest proteins with blackening bad smell production
Dr.T.V.Rao MD
13
The Agent
Clostridium perfringens Gram-positive bacteria
Anaerobic rod 3-8 x 0.4 1.2 mu
5 types (A-E)
Types B and D produce the epsilon toxin
Dr.T.V.Rao MD 14
Resistance
Vegetative bacteria is killed like other bacteria Cl.perfringens destroyed by boiling Cl, botulinum not killed even at 105 c 0 for less than 100 minutes All spores are killed at 121 oc in 20 minutes Halogens , Glutaraldehyde are effective on spores Metronidazole and Pencillin and Chloramphenicol are effective
Dr.T.V.Rao MD
16
Virulence Factors
Virulence factors toxins
alpha toxin causes RBC rupture, edema and tissue destruction
Toxins
The toxins of Cl. perfringens
toxin (phospholipase C, lecithinase) is the most important toxin
Lyses of RBCs, platelets, leucocytes and endothelial cells Increased vascular permeability with massive hemolysis and bleeding tissue destruction Hepatic toxicity and myocardial dysfunction
Epsilon Toxin
Produced as an inactive protoxin Activated by trypsin
Removes a 13-residue N-terminal peptide
Pathology
Not highly invasive; requires damaged and dead tissue and anaerobic conditions Conditions stimulate spore germination, vegetative growth and release of exotoxins, and other virulence factors. Fermentation of muscle carbohydrates results in the formation of gas and further destruction of tissue.
Dr.T.V.Rao MD
22
Clostridial Cellulitis
Dr.T.V.Rao MD
26
Dr.T.V.Rao MD
27
Laboratory Diagnosis
Specimen: Histological specimen or wound exudates
Histological specimen transferred aseptically into a sterile screw-capped bottle & used immediately for microscopical examination & culture Specimens of exudates should be taken from the deeper areas of the wound where the infection seems to be most pronounced
Dr.T.V.Rao MD
30
Dr.T.V.Rao MD
31
C. perfringens
Laboratory Diagnosis
Specimens: pus, necrotic tissue, feces, food, etc. Smears: large gram-positive rods with or without spores, usually in the absence of leukocytes.
Culture: anaerobic culture on blood plate. Identification: Storming fermentation-- clot torn by gas in 24 hrs. Lecithinase test-- precipitate formed around colonies on egg yolk media. Biochemical tests.
Dr.T.V.Rao MD 32
Naglers Reaction
This test is done to detect the lecithinase activity The M.O is inoculated on the medium containing human serum or 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
Dr.T.V.Rao MD 33
NOTE:
Lecithinase (-toxin; phospholipase) hydrolyzes phospholipids in egg-yolk agar around streak on right. Antibody against -toxin inhibits activity around left streak.34 Dr.T.V.Rao MD
Nagler Reaction
35
36
Biochemical Tests
Cl. perfringnes characterized by:
It ferments many carbohydrates with acid & gas It acidified litmus milk with stormy clot production
C. perfringens Treatment
Treatment for Suppurative myositis or myonecrosis: Prompt and extensive debridement. Antibiotics (penicillin) administration. Hyperbaric oxygen may "detoxify" patients rapidly. Efficacy of antitoxins is doubtful. C. perfringens food poisoning requires only symptomatic care.
Dr.T.V.Rao MD
43
Dr.T.V.Rao MD
44
Diagnosis depends upon the isolation of similar strains of C. perfringens from the faeces of patients and from others at risk who have eaten the suspected food, and from the food itself. Numbers usually exceed 106 organisms/g faeces. The isolates can be sent to a reference laboratory for special typing to prove their relatedness.
Dr.T.V.Rao MD 49
Programme Created by Dr.T.V.Rao MD for Medical and Paramedical Students in Developing World
Email
doctortvrao@gmail.com
Dr.T.V.Rao MD
51