EXERCISE 14

Question 1

Principles of Bacterial Cultivation

1. To [?] all bacteria present in a clinical specimen.
2. To determine which of the bacteria that grow are most likely [?] and which are likely [?].
3. To obtain sufficient growth of [?] to allow identification, characterization, and susceptibility testing.

Answer 1

1. grow and isolate
2. causing infection; contaminants or colonizers
3. clinically relevant bacteria

Question 2

Processing of Cultures

1. Select [?] appropriate for the particular specimen type.
2. Determine the [?[ of incubation to recover all organisms of potential significance.
3. Determine which of the [?] on primary media require further characterization.
4. Determine whether [?] are required once the identification of the organism is known.

Answer 2

1. primary culture media
2. temperature and atmosphere
3. isolated recovered
4. antimicrobial susceptibility tests

Question 3

The decision of how far to process the individual specimen cultures must be based on a thorough knowledge of the [?] that apply in any given case.

Answer 3

host-parasite relations

Question 4

“processing control is the restricting the processing and reporting of culture specimens to the production of predictably useful information.”

Answer 4

Barlett call

Question 5

are the culture media that frows the bacteria present in the specimen from the patient

Answer 5

Primary culture media

Question 6

is the cell culture grown on the primary culture media.

Answer 6

Primary culture

Question 7

Selection of Primary Culture Media

Answer 7

1. Agar plates
2. Blood agar medium (✓ Horse blood or sheep blood; ✓ Human blood)
3. MacConkey agar or Eosin methylene blue
4. Broth media
5. Enrichment broth

Question 8

are commonly used

Answer 8

Agar plates

Question 9

is the most common non-selective included in the battery of primary isolation media for virtually every clinical specimen

Answer 9

Blood agar medium

Question 10

agar supplemented with additive such as IsoVitalex

Answer 10

Horse blood or sheep blood

Question 11

is recommended for the recovery of Gardnerella vaginalis, for which, in addition to promoting good growth, hemolysis not seen on sheep blood agar can be observed, providing one clue for the presumptive identification

Answer 11

Human blood

Question 12

is the selective culture medium most commonly used to inhibit gram-positive organisms.

Answer 12

MacConkey agar or Eosin methylene blue

Question 13

should be limited only to those specimens such as body fluids, needle biopsies, or deep tissue aspirations, in which recovery of even a few organisms in low concentration may be significant, or for which the chance of recovery of an anaerobe is reasonable.

Answer 13

Broth media

Question 14

Recovery of the organism in broth culture only after 4-5 days of incubation will have little clinical significance. Except for the recovery of [?] from blood and other specimens and the recovery of [?] in patients suspected of endocarditis.

Answer 14

• Brucella species
• ## Cardiobacterium hominis

Question 15

used to recover pathogenic organisms from specimens, such as feces, in which there is a heavy concentration of commensal organisms

Answer 15

Enrichment broth

Question 16

[?] are held in a prolonged lag phase of growth by the inhibitors in the enrichment broth, allowing relatively few bacterial cells of pathogens (?) to enter uninhibited log phase of growth, during which they can better compete for survival

Answer 16

Commensals

Salmonella and Shigella

Question 17

Specimen Preparation

1. [?] (grinding) of tissue
2. Concentration by [?] of large volumes of sterile fluids, such as ascites (peritoneal) or pleural (lung) fluids
3. [?] of specimens (for legionellae or mycobacteria)

Answer 17

1. Homogenization
2. centrifugation or filtration
3. Decontamination

Question 18

is the process of introducing microorganisms into a culture medium usually using nichrome or platinum inoculating wire or loop

Answer 18

Inoculation

Question 19

are plated quantitatively; everything else is usually plated semi-quantitatively

Answer 19

Urine (Clean-catch midstream urine) and tissues from burn victims

Question 20

Quantitative inoculation is done by either:

Answer 20

a. Plate count/ viable cell count either through pour plate or spread plate method
b. by means of a quantitative inoculating loop (1:100 or 1:1000)

Question 21

original streak line is cross-struck with an ordinary loop to produce isolated, countable colonies

Answer 21

quantitative inoculating loop (1:100 or 1:1000)

Question 22

is applied by swabbing a dimesized area or placed a drop of liquid specimen on the plate; (streaking for isolation)

Answer 22

Quadrant streak

Question 23

Quadrant streak

Answer 23

Semiquantitative by using an ordinary loop

- Quadrant streak (streaking for isolation. In

Question 24

Semiquantitative by using an ordinary loop: Quadrant streak (streaking for isolation)

1. The original inoculums are then [?] with an ordinary nichrome inoculating loop.
2. The loop is then [?] and quadrant two is struck by pulling the loop through quadrant one a few times and streaking the rest area.
3. The loop is then flamed again and quadrant three is streaked by going into quadrant two a few times and [?] the rest of the area.
4. Finally, quadrant four is streaking by pulling the loop over the [?] without flaming.

Answer 24

1. cross-struck
2. flipped over or flamed
3. streaking
4. rest of the agar

Question 25

Inoculation procedure seems simple but like media preparation, it must be performed [?].

Answer 25

cautiously

Question 26

For the past 150 years, bacteria have been investigated primarily in

Answer 26

liquid batch cultures

Question 27

Contrary to most expectations, these cultures are not homogeneous mixtures of singlecell bacteria, because [?] eventually develop in most liquid batch cultures

Answer 27

free-floating bacterial aggregates

Question 28

These aggregates share characteristics with biofilms, such as [?]. We investigated how aggregates develop and what influences this development in
liquid batch cultures of Pseudomonas aeruginosa.

Answer 28

increased antibiotic tolerance

Question 29

We focused on how the method of inoculation

affected aggregation by assessing [?] using confocal laser scanning microscopy.

Answer 29

aggregate frequency and size

Question 30

Several traditional methods of initiating an overnight bacterial culture, i.e., inoculation directly from frozen cultures, inoculation using agar-grown cells, or inoculation using cells grown in liquid cultures, were investigated. We discovered a direct link between the [?] and the size and frequency of [?] in liquid batch cultures, with inoculation directly from a plate resulting in the most numerous and largest aggregates.

Answer 30

inoculation method; biofilm aggregates

Question 31

These large aggregates had an overall impact on the cultures’ subsequent tolerance toward [?], indicating that the inoculation method has a profound impact on antibiotic tolerance.

Answer 31

tobramycin

Question 32

We also observed a mechanism whereby preformed aggregates recruited single cells from the surrounding culture in a [?] building up aggregated biomass in the culture. This recruitment was found to rely heavily on the exopolysaccharide Psl.

Answer 32

“snowball effect”

Question 33

Additionally, we found that both Escherichia coli and Staphylococcus aureus produced aggregates in liquid batch cultures. Our results stress the importance of [?] throughout experiments and the substantial impact [?] in liquid batch cultures may have on the outcomes of microbiological experiments.

Answer 33

inoculation consistency; aggregate development

Question 34

[?] are fundamental to the field of microbiological research. These cultures are normally thought of as [?] of single cell bacteria; the present study shows that this is not always true.

Answer 34

Pure liquid cultures; homogeneous mixtures

Question 35

Bacteria may aggregate in these liquid cultures. The aggregation can be induced by the method chosen for [?]. The presence of aggregates can significantly change the outcomes of experiments by altering the [?] of the cultures

Answer 35

inoculation; phenotype

Question 36

The study found a mechanism whereby preformed aggregates can recruit surrounding single cells in a form of snowball effect, creating [?] in the cultures.

Answer 36

more and larger aggregates

Question 37

Once formed, these aggregates are [?]. Aggregates in liquid cultures may be an immense unseen challenge for microbiologists

Answer 37

hard to remove