Bacterial Diagnosis - Hunter

Question 1

What is a common symptom for bacterial infections?

Answer 1

Fever.

Question 2

Describe the steps in laboratory diagnosis of bacterial diseases.

Answer 2

1. Physician examines the patient and suspects an infectious diseases.
2. Physician may institute treatment before laboratory confirmation of the diagnosis.
3. Physician collects the appropriate specimen in the appropriate transport medium.
4. Physician provides the preliminary clinical diagnosis to the clinical microbiologist.
5. Clinical microbiologist selects the appropriate diagnostic method or methods, performs tests and reports results.
6. Physician implements appropriate therapy.

Question 3

In what instance would a physician institute treatment before laboratory confirmation of the diagnosis?

Answer 3

Treatment based on clinical diagnosis often comes before laboratory confirmation - especially if suspicion is high for a highly virulent infection such as meningitis.

Question 4

What is the most common reason for failing to establish an etiologic diagnosis or suggesting the wrong diagnosis?

Answer 4

Failure of proper specimen collection. Improperly collected specimens may be contaminated and interfere with the proper diagnosis.

Question 5

What is a special consideration when diagnosing a bacterial infection?

Answer 5

Distinguishing resident or normal flora microbes from those causing infection.

Question 6

What affects specimen collection?

Answer 6

The anatomic site of the infection. Some sites allow for clean collecting while others are more challenging.

Question 7

What is a direct specimen?

Answer 7

This is the type of specimen collected when the microbe are in a sterile site that can be accessed directly.

Question 8

What are some examples of direct specimens?

Answer 8

1. specimen take from a needle aspiration of a deep abscess

2. specimen taken via blood collection

Question 9

What is an indirect specimen?

Answer 9

Specimen taken when microbes are in a sterile site but must be collected through a non-sterile site, for example a urine sample.

Question 10

What is a contaminated specimen?

Answer 10

Specimen taken when microbes are in a site contaminated with normal flora, for example a throat or stool culture.

Question 11

What dictates how the clinical microbiology lab approaches the isolation and identification of the bacterial pathogen?

Answer 11

The specimen type and the presumptive clinical diagnosis.

Question 12

What is the most commonly used tool for specimen collection and what are some concerns about this method?

Answer 12

A sterile swab. It only collects a small amount and it dries out quickly.

Question 13

Why is a transport medium important?

Answer 13

Transport mediums such as a buffered fluid or a semisolid medium prevent drying out, maintain neutral pH and minimizes growth of contaminants.

Question 14

What are 2 types of transport containers?

Answer 14

1. aerobic

2. anaerobic

Question 15

What are some methods of identification of specific microbes in the lab?

Answer 15

1. microscopy
2. broth and agar culture - antibiotic sensitivity testing
3. biochemical characterization
4. antibody detection
5. antigen detection
6. nucleic acid based tests

Question 16

Describe brightfield microscopy.

Answer 16

This is the most common type of microscopy in which the light is focused directly on the specimen.

Question 17

Describe darkfield microscopy.

Answer 17

This is when the central light is blocked and only peripheral light is collected as scatter from the microbes.

Question 18

What types of specimens would you use darkfield microscopy on?

Answer 18

Thin and hard to see specimens such as spirochetes.

Question 19

Describe fluorescence microscopy.

Answer 19

This is similar to dark field except that microbes are labeled with dye that fluoresces when it interacts with light of an appropriate wavelength.

Question 20

What magnification are most bacteria examined with?

Answer 20

100X oil immersion objective.

Question 21

What is a simple stain?

Answer 21

This is a stain that uses a single dye to visualize bacteria.

Question 22

What are differential stains?

Answer 22

These are used to distinguish between different bacterial groups such as gram-pos and gram-neg bacteria. Acid fast is also a differential stain.

Question 23

What is a special stain?

Answer 23

This is a stain used to detect bacterial structures such as capsules, flagella and endospores. An example is India ink - stains carbohydrate moieties.

Question 24

Describe acid fast staining.

Answer 24

1. an acid-fast microbe is stained with a high concentration of dye.
2. The acid-fast microbe will resist the decolorizing step that is usually done with alcohol or dilute acid - hence the name acid fast.

Question 25

Describe bacterial cultures?

Answer 25

1. culture of microbes in nutrient broth or on agar medium is commonly used.
2. a single microbe can grow to amounts that are visible.
3. almost all medically important microbes can be cultured.

Question 26

What is nutrient media?

Answer 26

A type of culture media made from animal or plant products supplemented with a variety of nutrients.

Question 27

What is selective media?

Answer 27

A type of culture media used when specific pathogens are sought in sites with an extensive microbial flora. Usually chemical additives or antimicrobials that inhibit unwanted microbial growth.

Question 28

What is indicator media?

Answer 28

A type of culture media that contains substances designed to demonstrate biochemical or other features of specific pathogens - such as pH indicators of fermentation of specific sugars or red blood cells that can be hemolyzed (Sheep’s blood agar).

Question 29

Describe culture conditions.

Answer 29

Different microbes will need different culture conditions. For example:

1. most aerobic bacteria are maintained in an incubator at 35-37 degrees celsius.
2. some capnophilic require CO2 to grow.
3. anerobic microbes need an oxygen free environment.
4. Some microbes grow best in a broth and some in a semi-solid media.

Question 30

What is antimicrobial sensitivity testing?

Answer 30

This when a culture is grown and then exposed to different antimicrobials to see if it is sensitive or to allow the selection of the most effective chemotherapeutic agent. Both broth dilution tests and agar diffusion tests are routinely used to determine the minimum inhibitory concentration of an antibiotic.

Question 31

Give some examples of other sensitivity testing that helps to identify bacteria.

Answer 31

1. Bile solubility- may or may not be soluble in bile
2. Catalase - pos or neg
3. Coagulase - pos or neg
4. Oxidase - pos or neg

Question 32

What test is used to differentiate fermenters from non-fermenters?

Answer 32

The AP120e system.

Question 33

What is serology?

Answer 33

The detection of antibodies made against microbes. These antibodies can be detected by methods such as ELISA, Western blot and immunofluorescence.

Question 34

What can antibody detection tell you?

Answer 34

First, it can tell you if the body has been exposed to that microbe. Second, it can tell you if the infection is current, recent or a past infection. Current or recent infections are indicated by IgM antibodies. Past infections by IgG.

Question 35

What are the downsides to antibody detection?

Answer 35

1. the antibody response to some bacteria can take several weeks, limiting its usefulness for diagnosis.
2. Antibody detection can be unreliable in immunocompromised patients such as those with SCID or HIV.

Question 36

Describe antigen detection.

Answer 36

Whole bacteria, bacterial antigens, and bacterial toxins released into body fluids can be detected by antibodies using a variety of assays- an example is the rapid strep test.

Question 37

Describe nucleic acid analysis.

Answer 37

Tests - either amplified or non-amplified - for detecting nucleic acid signature of a bacterial pathogen. Non-amplified assays are based on hybridization of nucleic acid to labeled probes - rapid and used when sensitivity is not as important (i.e. identifying if bacteria is present). PCR is an amplified assay and is extremely sensitive but subject to contamination.