Bacteriology-31 Non Culture Based Methods For Clinical Diagnosis*

Front 1

What are some immunologic techniques for identification of infectious agents in patients

Back 1

Fluorescent antibody test (direct or indirect method)
Immunochromatographic methods (lateral flow or pregnancy test format)
Enzyme immunoassay (EIA) or ELISA
Latex agglutination

Front 2

When is the Fluorescent antibody (FA) test for cell-associated antigens used

Back 2

For visualization of agent in clinical material or tissue culture by microscopy. Can use direct (labeled primary antibody) or indirect (labeled secondary antibody, which is more sensitive)

Front 3

Fluorescent antibody test (FA) is used for detection of what

Back 3

Pneumocystis
Legionella
Bordetella
Chlamydia
Agents that are difficult or time-consuming to culture

Front 4

What are immunochromatographic methods, such as lateral flow or the pregnancy test, use for

Back 4

Rapid testing for strep throat; Shiga toxin in fecal extracts

Front 5

What is ELISA used for

Back 5

If enzyme labeled secondary antibody develops color, we know the antibody has recognized and bound the antigen we are looking for

Front 6

What is latex agglutination used for

Back 6

Rapid identification of pathogens in CSF; Group B strep in vaginal secretions; Clostridium difficile toxin in fecal extracts; Rotovirus in feces

Front 7

What are the two nucleic acid-based techniques for identification of infectious agents in patient material

Back 7

Nucleic acid probes and Amplification techniques

Front 8

What are the basic components of probe-based detection systems

Back 8

A probe or single stranded DNA or RNA (many copies needed)
A target of nucleic acid
Hybridization conditions
A method for detection of hybridized product

Front 9

What are the advantages of nucleic acid probes
What is the main disadvantage

Back 9

High specificity (few false positives)
Very rapid (same day results)
Requires little specialized laboratory equipment
The main disadvantage is the low sensitivity

Front 10

What are some methods for improving the sensitivity of nucleic acid probes

Back 10

Amply the number of targets
Add enzymes that make more targets
Enhance the identification of pairs with enzyme-linked anti-hybrid antibodies

Front 11

What are the best applications for nucleic acid probes

Back 11

Identify organisms that grow very slowly (TB)
For organisms that are expensive or labor intensive to culture (viruses; Chlamydia)
For organisms that are non culturable (leprosy, syphilis)

Front 12

What are the components needed for PCR

Back 12

Primers; Target DNA; Temperature-stable DNA polymerase; Thermocycling device

Front 13

What are some pathogens for which the PCR reaction has been developed

Back 13

Mycobacterium tuberculosis
Chlamydia trachomatis
Lyme disease (Borrelia burgdorferi)
HIV-1
HTLV-1

Front 14

What are the advantages of PCR

Back 14

High specificity and sensitivity
Minute sample can be used
Can detect the presence of microorganisms that cannot be cultured
Speed

Front 15

What are the limitations of PCR

Back 15

Potential for false positives through contamination
Potential for false negatives through amplification inhibitors
No antimicrobial susceptibility patterns provided
Requires special expensive equipment

Front 16

What is real-time PCR

Back 16

PCR where the DNA synthesized can be measured during the reaction using SYBRgreen or DNA probes

Front 17

What are the advantages of Real time PCR

Back 17

Allows user to quantitate product
Eliminates electrophoresis step
Allows for detection of multiple products

Front 18

What are the limitations of Real time PCR

Back 18

Requires expensive instrumentation
Requires synthesis of fluorescent probes

Front 19

What are the applications of Real time PCR

Back 19

Can test for multiple targets at one time, and can estimate viral load (HIV)

Front 20

What are the main characteristics of RT PCR

Back 20

The target sequence is RNA. Can identify RNA based viruses, determine what genes are being expressed (to determine viral load), and detect expression of genes in latent infections (CMV)

Front 21

What are some general considerations in detection of immune response against infectious agents

Back 21

Time to seroconversion
Discrimination of old versus new infections (antibody class)
Four-fold rise between acute and convalescent sera

Front 22

What are some means of detection of pathogen-specific antibodies

Back 22

Precipitation reactions (immunodiffusion and CIE)
Agglutination reactions
EIA, ELISA
Western Blot
RIA
Complement fixation

Front 23

What is molecular subtyping used for

Back 23

Estimating the degree of relatedness of bacterial isolates of the same or closely related species

Front 24

In pulse field gel electrophoreis (PFGE), what indicates relatedness

Back 24

Three band differences or less

Front 25

What species are tested in Pulsenet's database

Back 25

Shigella species
Salmonella enterica
E.coli O157:H7
Campylobacter
Listeria monocytogenes

Front 26

What is serotyping used for

Back 26

Distinguishing closely related species or strains of the same species. Includes the O antigen, capsular antigens, flagellar capsular types, and outer membrane proteins