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CHAPTER 24 Diagnostic Microbiology and Immunology. Growth-Dependent Diagnostic Methods Isolation of Pathogens from Clinical Specimens. Proper sampling and culture of a suspected pathogen is the most reliable way to identify an organism that causes a disease ( Figure 24.1 ).

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CHAPTER 24

Diagnostic Microbiology and Immunology


Growth dependent diagnostic methods isolation of pathogens from clinical specimens

Growth-Dependent Diagnostic MethodsIsolation of Pathogens from Clinical Specimens



  • Enrichment culture most reliable way to identify an organism that causes a disease (, the use of selected culture media and incubation conditions to isolate microorganisms from samples, is an important part of clinical microbiology.

  • Table 24.1 shows recommended enriched media and selective media for primary isolation of pathogens.

Most clinical samples are first grown on general-purpose media, media such as blood agar that support the growth of most aerobic and facultatively anaerobic organisms.


Differential media are specialized media that allow identification of organisms based on their growth and appearance on the media.


Bacteremia is the presence of bacteria in the blood

  • Septicemia identification of an isolate by observing the color and morphology of colonies of the suspected pathogen growth on various media, as described in is a blood infection resulting from the growth of a virulent organism entering the blood from a focus of infection, multiplying, and traveling to various body tissues to initiate new infections.

Bacteremia is the presence of bacteria in the blood.

  • The selection of appropriate sampling and culture conditions requires knowledge of bacterial ecology, physiology, and nutrition.


Growth dependent identification methods

Growth-Dependent Identification Methods identification of an isolate by observing the color and morphology of colonies of the suspected pathogen growth on various media, as described in

Traditional methods for identifying pathogens depend on observing metabolic changes induced as a result of growth. These growth-dependent methods provide rapid and accurate pathogen identification.


  • Table 24.3 identification of an isolate by observing the color and morphology of colonies of the suspected pathogen growth on various media, as described in gives important clinical diagnostic tests for bacteria.


Antimicrobial drug susceptibility testing

Antimicrobial Drug Susceptibility Testing identification of an isolate by observing the color and morphology of colonies of the suspected pathogen growth on various media, as described in

  • Pathogens should be tested for susceptibility to individual antibiotics to ensure appropriate chemotherapy. This rigorous approach to antimicrobial drug treatment is usually applied only in health care settings.

Antimicrobial drugs are widely used for the treatment of infectious diseases.



Agar media are inoculated by evenly spreading a defined density of a suspension of the pure culture on the agar surface. Filter paper disks containing a defined quantity of the antimicrobial agents are then placed on the inoculated agar.


Antibiograms inhibition zone around each disk is measured. are periodic reports that indicate the susceptibility of clinically isolated organisms to the antibiotics in current local use.


Safety in the microbiology laboratory

Safety in the Microbiology Laboratory inhibition zone around each disk is measured.

Safety in the clinical laboratory requires effective training, planning, and care to prevent the infection of laboratory workers with pathogens.

  • Materials such as live cultures, inoculated culture media, used hypodermic needles, and patient specimens require specific precautions for safe handling.


Immunology and clinical diagnostic methods immunoassays for infectious disease

Immunology and Clinical Diagnostic Methods inhibition zone around each disk is measured. Immunoassays for Infectious Disease



Specific immune responses, particularly antibody major aspects of immunity are summarized in titers and skin tests, can be monitored to provide information about past infections, current infections, and convalescence (Figure 24.11).


Common immunodiagnostic tests for pathogens are shown in table 24 5

Common immunodiagnostic tests for pathogens are shown in major aspects of immunity are summarized in Table 24.5.


Polyclonal and monoclonal antibodies

Polyclonal and Monoclonal Antibodies major aspects of immunity are summarized in

Polyclonal and monoclonal antibodies are used for research and clinical applications.


  • Hybridoma technology ( major aspects of immunity are summarized in Figure 24.12) provides reproducible, monospecific antibodies for a wide range of clinical, diagnostic, and research purposes.


  • Table 24.6 major aspects of immunity are summarized in shows characteristics of monoclonal and polyclonal antibody production.


In vitro antigen antibody reactions serology

In Vitro Antigen-Antibody Reactions: Serology major aspects of immunity are summarized in

The study of antigen-antibody reactions in vitro is called serology.



  • Specificity antigen. Types of antigen-antibody reactions are shown in (Table 24.8) and sensitivity define the accuracy of individual serological tests.


Neutralization antigen. Types of antigen-antibody reactions are shown in (Figure 24.13) and precipitation (Figure 24.14) reactions are examples of antigen-binding tests that produce visible results involving antigen-antibody interactions.


Agglutination

Agglutination antigen. Types of antigen-antibody reactions are shown in

Direct agglutination tests are widely used for determination of blood types (Figure 24.15).


A number of passive agglutination tests are available for identification of a variety of pathogens and pathogen-related products. Agglutination tests are rapid, relatively sensitive, highly specific, simple to perform, and inexpensive.


Fluorescent antibodies

Fluorescent Antibodies identification of a variety of pathogens and pathogen-related products. Agglutination tests are rapid, relatively sensitive, highly specific, simple to perform, and inexpensive.

Fluorescent antibodies are used for quick, accurate identification of pathogens and other antigenic substances in tissue samples and other complex environments.


  • Fluorescent antibody-based methods ( identification of a variety of pathogens and pathogen-related products. Agglutination tests are rapid, relatively sensitive, highly specific, simple to perform, and inexpensive.Figure 24.18) can be used for identification, quantitative enumeration, and sorting of a variety of cell types.


Enzyme linked immunosorbent assay and radioimmunoassay

Enzyme-Linked Immunosorbent Assay and Radioimmunoassay identification of a variety of pathogens and pathogen-related products. Agglutination tests are rapid, relatively sensitive, highly specific, simple to perform, and inexpensive.


  • ELISA identification of a variety of pathogens and pathogen-related products. Agglutination tests are rapid, relatively sensitive, highly specific, simple to perform, and inexpensive. (Figures 24.23, 24.24) and RIA (Figure 24.26) methods are the most sensitive immunoassay techniques.


Both involve linking a detection system, either an enzyme or a radioactive molecule, to an antibody or antigen, significantly enhancing sensitivity.


Immunoblot procedures

Immunoblot Procedures tests have been designed to detect either antibody or antigen in many applications.

Immunoblot (Western blot) procedures (Figure 24.27) are used to detect antibodies to specific antigens or to detect the presence of the antigens themselves.


The antigens are separated by electrophoresis, transferred (blotted) to a membrane, and exposed to antibody.


Molecular and visual diagnostic methods nucleic acid methods

Molecular and Visual Diagnostic Methods radioactive secondary antibodies. Immunoblots are extremely specific, but procedures are complex and time-consuming.Nucleic Acid Methods



To design a nucleic acid probe, a nucleic acid sequence specific for the microorganism of interest must be available.


  • Table 24.9 is in the application of gene amplification (PCR) methods. Various DNA-based methodologies are currently used in clinical, food, and research laboratories. lists pathogens identified with nucleic acid probe and PCR methods.


Diagnostic virology

Diagnostic Virology is in the application of gene amplification (PCR) methods. Various DNA-based methodologies are currently used in clinical, food, and research laboratories.

Virus propagation in vitro can be accomplished only in tissue culture. Therefore, most diagnostic techniques for viral identification are not growth-dependent but routinely rely on immunoassays and nucleic acid–based techniques.

  • Electron microscopy techniques are useful for direct observation of viruses in host samples.


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