Culture &Blood culture
Prof. Md. Akram Hossain
Deptt.. Of Microbiology, MMC
Deptt
July 2012
Prof. Muhammad Akram Hossain, Blood
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What is C/S?
It means Culture and Sensitivity
What is Culture?
Growth of microbes in artificial food
(medium) and environment
What is Sensitivity?
To see which antimicrobial will kill the
bacteria
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Culture technique
MATERIALS:
•Specimen
•Sterile cotton swab
•Bacteriological loop
•Suitable media
•Incubator
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Culture technique…
With the inoculating
loop, proceed to streak
in a second and third
streak pattern (see
illustration)..
illustration) Briefly,
isolate bacterial colonies
by pulling several
streaks out from the
original swab inoculum
site.. Use the method
site
shown above to avoid
tearing the agar..
agar
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Culture technique…
Incubate the plate at
37oC for 24-48
hours.
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Sensitivity technique…
Methods:
1.Disk diffusion
(Kirby –Bauer)
2.Broth/Drug
dilution
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What is blood culture?
A blood culture is done when a person has
symptoms of a blood infection, also called
Septicemia. Blood is drawn
bacteremia or Septicemia.
from the person one or more times and is tested
in a laboratory to find and identify any
microorganism present and growing in the
blood.. If a microorganism is found, Sensitivity is
blood
also done
done..
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When blood culture done?
To establish the diagnosis
To identify the causative organism(s)
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When blood culture done?
To establish the diagnosis in suspected
1. septicaemia,
2. endocarditis,
3. bacterial meningitis,
4. pericarditis,
5. septic arthritis,
6. osteomyelitis,
7. pyelonephritis,
8. enteric fever.
fever.
.
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When blood culture done?
To identify the causative organism(s) in
1. severe pneumonia,
2. postpartum fever,
3. pelvic inflammatory disease,
4. cannula sepsis;
sepsis;
5. neonatal epiglottitis
6. sepsis..
sepsis
7. Investigation of patients with PUO
PUO..
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When blood culture done?
1. Septicemia
2. Sepsis
3. Meningitis
4. Enteric fever
5. Endocarditis
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How blood culture differs from other
cultures?
1. Liquid media (broth) is required
2. Amount of sample (Blood) is more 5-5-10
ml ( other e.g. CSF, Urine etc only 1-
1-2
drops)
3. More than 1 sample is to be cultured
4. Timing of collection is important
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When blood is to be collected for culture?
During most blood infections (called intermittent
bacteremia) microorganisms enter the blood at various
time intervals.
intervals.
Blood drawn randomly may miss the microorganisms
microorganisms..
Microorganisms enter the blood 30 30--90 minutes before
the person's fever spikes, collecting the culture just after
the fever spike offers the best likelihood of finding the
microorganism..
microorganism
The second and third cultures may be collected at the
same time, but from different places on the person, or
spaced at 30-30-minute or one-
one-hour intervals, as the
physician chooses
chooses..
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When blood is to be
collected for culture?..
During continuous bacteremia, such as infective
endocarditis, microorganisms are always in the
blood and the timing of culture collection is less
important..
important
Blood cultures should always be collected before
antibiotic treatment has begun
begun..
In Enteric fever-
fever- in the first week of illness
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How many blood cultures?
The physician must also decide how many blood
cultures should be done.
done.
One culture is rarely enough
enough,, but two to three
are usually adequate.
adequate.
Four cultures are occasionally required.
required.
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Types of blood culture
1. Traditional blood culture
• Broth culture or biphasic culture
2. Lytic centrifugation method
3. Automated blood culture
culture--continuous-
continuous-
monitoring blood culture systems
(CMCCS).
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How to collect the blood for culture?
Ten ml (milliliter) of blood is usually needed for each blood
culture bottle
bottle.. (1:10 proportion)
First locates a vein in the inner elbow region.
region. The area of skin
where the blood will be drawn must be disinfected by wiping the
area with alcohol in a circular fashion,
The same pattern of wiping is repeated using an iodine or
iodophor solution
solution..
The top of the bottle is disinfected using alcohol
alcohol..
Draw the blood and about 10 ml of blood is injected into each
blood culture bottle
bottle..
The type of bottles used will vary based on whether the
physician is looking for bacteria (aerobes or anaerobes), yeast,
mold, or viruses
viruses..
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Standard collection protocol
Collect up to 3-
3-4 sets of blood culture bottles
Two blood culture bottles in each set
Obtain 5 ml blood per bottle
Draw from up to 3 different venipuncture sites
Space blood collections by 1 hour intervals
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New collection protocol
Six bottle method
Collect 6 bottles from one venipuncture (total
40 ml)
Based on mathematical model
Predicts highest sensitivity and Specificity
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Collection…
Suspected sepsis, meningitis, osteomyelitis, arthritis, or acute untreated
bacterial pneumonia
pneumonia::
Obtain two blood cultures from two different sites, such as the left and
right arms
arms..
Fever of unknown origin such as that caused by an occult abscess abscess::
Obtain two blood cultures initially.
initially. If those are negative, obtain two more
24-
24-36 hours later.
later. The yield beyond three or four cultures is virtually nil in
this condition
condition..
Suspected early typhoid fever and brucellosis:
brucellosis:
Obtain four blood cultures over 24- 24-36 hours due to low-
low-grade bacteremia
involved in these rarely seen diseases.
diseases.
Endocarditis (acute infective endocarditis):
endocarditis):
Obtain three blood cultures from three separate venipuncture sites during
the first 1-2 hours and begin therapy.
therapy.
Subacute infective endocarditis
endocarditis::
Obtain three blood cultures within the first 24 hours, ideally within no less
than hourly intervals
intervals.. If all are negative at 24 hours, obtain two more
more.. The
yield beyond five blood cultures in subacute and endocarditis is virtually
nil.
nil. Prof. Muhammad Akram Hossain, Blood
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Prof. Muhammad Akram Hossain, Blood
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Automated blood culture
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Procedure in Microbiology Lab?
Bacteriological Media
Culture in enriched non-
non-selective broth for aerobes,
anaerobes and yeasts.
Bile Broth, Glucose broth, TSI, BHI
Incubation
for 5-
5-7 days, often extended to 14-
14-21 days for suspected
bacterial endocarditis or infection with Brucella sp or yeasts
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Procedure in Microbiology Lab?
Growth is detected
by the presence of turbidity or haemolysis,
Gram stain,
more commonly production of carbon dioxide,
alteration in redox potential or change in pH detected by an automated
monitoring system Incubation
The organism is identified
by Gram stain of the broth,
colony appearance after subculture to solid media and
biochemical and/or antigen tests.
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Interpretation
Most pathogens grow within 1-2 days; days; some organisms may
require longer eg, Candida sp, Brucella sp
sp..
Cultures are usually positive in infective endocarditis (if collected
prior to antibiotic treatment) and early typhoid
typhoid..
Negative cultures do not exclude infection
infection..
Organisms usually regarded as non non--pathogenic contaminants
may cause infection in individuals whose defences are
compromised by
immunosuppression,
trauma, or
the presence of a prosthesis or an indwelling line.
line.
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Interpretation of Positive Blood Cultures
Virtually any organism, including normal flora, can cause bacteremia.
A negative culture result does not necessarily rule out bacteremia; false-
negative results occur when pathogens fail to grow.
A positive culture result does not necessarily indicate bacteremia; false-
positive results occur when contaminants grow.
Gram-negative bacilli, anaerobes, and fungi should be considered
pathogens until proven otherwise.
The most difficult interpretation problem is to determine whether an
organism that is usually considered normal skin flora is a true
pathogen.
From Flournoy DJ and Adkins L, "Understanding the Blood Culture
Report," Am J Infect Control, 1986,14:41-6,
Prof. Muhammad with permission.
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