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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5512_Библиотеки_им_академика_М_И_Перельмана.pdf
Introduction to Diagnostic Micbiolo
Inoculation of Mulple Tubes
Neisseria, Haemo
hilus)
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FIGURE 19.18
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Introduction to Diagnostic Micbiolo
Js19
Aerobic Culture
Capnophilic Culture
CO2-enriched atmosphere
2•
Candle Jar Technique
Anaerobic Culture
Thiogoate Bro Method
Clostridium, Actinomyces)
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Medical Laborato Technology: Volume 2
Columbia CNA ar:
Columbia Neomycin ar:
Kanamycin- VancomycinAgar: It
It
It
Procedure
It
Caution
°
Introduction to Diagnostic Micbiolo
Js21
Brewer Anaerobic Jar and Gas-P tem for Anaerobic Culture
quality control
°
repeated Refrigeration
and desiccation
°
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FIGURE 19.19
(e)
Introduction to Diagnostic Micbiolo
Js23
Short-term preservation can be accomplished by inoculating on agar slants (Figure 19.19)
for aerobes or in agar-stab cultures for the facultative anaerobes. In case of obligate anaer
obes, stab the medium and lay a layer of sterile mineral oil on the top. Fastidious anaer
obes are also cultured in thioglycollate broth with a layer of sterile mineral oil on the top. If
acid is a metabolic product of the organism being preserved, the medium employed should
be well-buered. During incubation, a core of growth develops around the line of stab. All
cultures are refrigerated after incubation.
Ideally, an initial presumptive diagnosis is made by the physician following physical exam
ination of the patient and a study of the clinical history. This is followed by the collection of
appropriate specimens and sending them to the laboratory for diagnosis. Communication
between the physician and the laboratory is established through the request form, which
should also bear the note from the physician regarding his presumptive diagnosis. The lab
oratory rst performs any direct examination that is appropriate for the given specimen.
the same time, the technician inoculates the specimen to the primary media. Very often the
direct examination guides the technician to choose the proper media and culture conditions
for the laboratory propagation of the infectious agent.
Some of the primary culture media recommended for dierent specimens are listed in
Table 19.3. The primary culture media are usually those best suited for the growth of the sus
pected pathogenic organisms. Streaking on the primary plates allows the tecician to isolate
the causative organism from a mixed population. Often a selective medium is also included
to suppress the growth of commensal normal ora present in the specimen. Some specimens,
however, are normally sterile (e.g., CSF and blood); in these cases there is no need for a selec
tive medium and growth of any organism in the primary culture is considered to be alarming
and requires further work up.
The primary culture may occasionally provide suicient information for the conrmatory
identication of organisms based on their colony character, smell, and ability to swarm or
dierential reactions on the medium. But usually, further tests will be needed using the pure
culture of the organism. This is attained by the secondary culture. Secondary cultures must
be pure cultures resulting from inoculation of single colonies on the primary culture into
additional media. All biochemical and serological tests leading to conrmatory identication
are done with the secondary culture. As therapy is dependent on the correct identication of
the oending organism, the importance of the isolation of a pure culture cannot be overem
phasized. Biochemical and serologic testing must be performed on pure culture to be valid. In
addition, antimicrobial susceptibility testing is only meaningful with a pure culture.
Three conditions have to be met before the pathogen can be successfully cultured in the
laboratory:
• Appropriate handling of the specimen is critical so that the organism is viable when
inoculated. Prompt inoculation on appropriate media or use of a suitable transport
media is recommended. Extreme care is needed to handle such delicate organisms
as spp., which cannot stand drastic temperature change or drying. Thus,
specimens (e.g., CSF) suspected of having spp. should never be refrigerated.
On the other hand, to avoid overgrowth of the pathogen or normal ora organisms,
urine should be refrigerated if a delay is anticipated.
• Appropriate culture media must be chosen for the organism to grow. Some organisms
caot be cultured in the laboratory (e.g., requiring the
use of alternative methods like serodiagnosis. Media used for various specimens, and
conditions in which they are kept for the identication of specic pathogenic bacteria,
will be described in subsequent pages.
Medical Laborato Technology: Volume 2
°
°
Primary Culture
pyogenes pneumoniae,
2
pyogenes pneumoniae.
Haemophilus
Clostridium Bacteroides
Salmonella Shigella
Salmonella Shigella
(Salmonella-Shigella
°
) Neisseria
2
a Culture Me
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mination of ary Culture
The primary plate provides considerable information that can assist in an early identication
of the pathogen. This includes growth rate, spreading habit, smell, size of the colony, and
appearance of the colony, growth on selective media and limited biochemical reactions on
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FIGURE 19.20
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inhibitory discs
Medical Laborato Technology: Volume 2
Secondary Culture
Carbohate Feentation Test
biochemical test
dierential media