1. Blood Culture GROUP MEMBERS: Aimen Niaz Rafia Hafeez Adeena Shafique
Zujaja tul Misbah
Sammia Rehman
Syeda Fatma H. Bukhari
PRESENTED BY:
Adeena Shaique
Sammia Rehman
Syeda Fatma H. Bukhari

2. PURPOSE OF BLOOD CULTURE
WHAT IS BLOOD CULTURE
A blood culture is the laboratory test in which blood is injected into bottles with media to determine the presence of microorganism invaded in blood of patient
PURPOSE OF BLOOD CULTURE
DIAGNOSIS
PROGNOSIS
THERAPY

3. BACTERAEMIA VS. SEPTICAEMIA
The presence of bacteria in the blood is called Bacteraemia
Bacteraemia occurs in
Typhoid fever
Brucellosis
Leptospirosis
Endocarditis.
Severe and life-threatening form of bacteraemia is called Septicaemia
Septic shock is due to
Gram negative bacilli
This is a clinical term used to describe severe life-threatening bacteraemia in which multiplying bacteria release toxins into the blood stream and trigger the production of cytokines, causing fever, chills, toxicity, tissue anoxia, reduced blood pressure, and collapse. Septic shock is usually a complication of septicaemia with Gram negative bacilli, and less frequently, Gram positive organisms. Prompt treatment is essential. Bacteraemia usually occurs when pathogens enter the bloodstream from abscesses, infected wounds or burns, or from areas of localized disease as in pneumococcal pneumonia, meningitis, pyelonephritis, osteomyelitis, cholangitis, peritonitis, enterocolitis and puerperal sepsis. There is usually a high white cell count with neutrophilia, left shift of the neutrophils, and often toxic granulation

4. PATHOGENS ISOLATED FROM BLOOD CULTURES
BACTERIA
E. coli
S. epidermidis
Neisseria meningitidis
Salmonella Typhi
S. aureus
FUNGI
Candida albicans
yeasts
Histoplasma capsulatum
NOTE:
Blood does not have a normal microbial flora.

5. Day 1 Collect blood and inoculate culture media
Collection of blood:
Blood should be collected before antimicrobial treatment has started
Collect the blood as the temperature of patient begins to rise
At least two specimens (collected at different times) should be cultured.
About 20 ml of blood is taken from adults.
Blood from neonates should be collected from a peripheral vein not from the umbilical vein.
1–2 ml of blood is enough to detect the presence of bacteria in children’s blood.

6. Choice of culture media
Diphasic blood culture medium
Columbia agar and broth
Commercially produced culture media
NOTE: Choice of media depends on the bacterial disease that is suspected. Some bacteria grow well in a particular medium, others do not.

7. Aseptic blood collection and dispensing technique
Wash hands.
Disinfect the venepuncture site with 70% ethanol and 2% tincture of iodine.
Decontaminate blood culture bottle tops.
Using a sterile syringe, withdraw 20ml blood.
Dispense 10 ml into the culture medium.
Incubate the inoculated media as soon as possible.
Incubate upto 4 weeks when brucellosis is suspected
Incubate upto 14 days for anaerobic infections

8. Aseptic Technique

9. Examining the blood culture:
1- CENTRIFUGE:
a sample of EDTA anticoagulated venous blood or heparinized capillary blood and make smears of the buffy coat layers.
2- STAIN:
_ Gram smear: for Gram positive and Gram negative bacteria detection
_ Ziehl-Neelsen smear: To detect AFB (acid fast bacteria)
_ Giemsa or rapid Field’s smear: To detect borreliae, or parasites such as trypanosomes, malaria parasites, and microfilariae.
3- DRY the smears, fix with absolute methanol for 2 minutes and stain by the appropriate staining technique.

10. Day 2 and Onwards 3- REPORT Diphasic culture (Columbia agar and broth)
Check for microbial growth, indicated by colonies growing on the agar slope, usually beginning at the agar-broth interface.
Colonial appearances
Colonies of staphylococci, S. Typhi, brucellae, and most coliforms can usually be seen easily, whereas colonies of S. pneumoniae, Neisseria species, S. pyogenes, and Y. pestis are less easily seen.
Pseudomonas and Proteus species produce a film of growth on the agar.
When growth is present:
– Subculture on blood agar, chocolate agar, and MacConkey agar.
– Incubate the blood agar and MacConkey agar plates aerobically and the chocolate agar plate in a carbon dioxide atmosphere (candle jar).
– Examine a Gram stained smear of the colonies.
Depending on the bacteria seen, test the colonies further (e.g. for coagulase, catalase, oxidase, urease, and motility).

11. Subculture on lactose egg yolk milk agar and incubate anaerobically
Large Gram positiverods
(C. perfringens)
Subculture the colonies on Kligler iron agar
motile, urease and oxidase negative Gram negative rods are isolated
Suspect Brucella species and send for identification.
Mark it as ‘High Risk’.
catalase positive,Gram negative coccobacilli are isolated

12. Examine toluidine blue smear (diphasic culture)
Examine thioglycollate culture
Subculture and examine microscopically
Incubate subculture anaerobically
NOTE: When there is no growth, wash slope of diphasic culture. Reincubate cultures and subculture.

13. Contamination of Blood Culture
Collection of blood
During subculturing
Frequent contaminants include:
Commensal staphylococci, micrococci, and diphtheroids
Contaminants from the environment
such as species of Bacillus or Acinetobacter.

14. Other possibilities
Occasionally in immunocompromised patients, organisms usually considered ‘contaminants’ may be pathogenic, especially fungi.
Contamination is indicated when an organism is recovered from only one bottle when it should have grown in both thioglycollate broth and the diphasic culture medium or when a mixed microbial flora is isolated.

15. Another use…
Severe and often fatal reactions can be caused by the transfusion of contaminated blood.
The bacteriological investigation of a transfusion reaction is as follows:

16. Visible signs Varied subculturing Tests
Appearing unusually dark in colour
Containing small clots
The plasma appearing red, or unusually turbid (examine after centrifuging a sample of the blood).
Visible signs
At room temperature
At 4°C
Varied subculturing
Motility Test
Gram stained smear of the plasma
Tests
Motility is interpreted using high dry magnification to locate the bacteria within the drop of water. If they swim randomly and "against the current" of water streaming across the slide surface, they are positive for motility. If they seem to be buffeted around, all moving the same direction and at the same speed, there is no motility. Instead, this type of movement is the result of the bacteria being knocked about by water molecules, called Brownian movement.