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Semmelweis and Lister

develop aseptic techniques, which prevented pathogens from entering the body during childbirth or surgery.

Later, Louis Pasteur developed

techniques for the aseptic transfer of bacteria.

What is an inoculum?

The bacteria transferred to a new medium.the bacteria that are transferred during inoculation are called the inoculum. The new sterile medium receives, or is inoculated with, the inoculum. The inoculum, it is hoped, consists only of the desired species of bacteria and does not include any contaminants.

Which of the following best describes aseptic technique?

To manipulate bacteria without introducing contaminants.

Arrange the steps of loop sterilization in correct order

1)attach burner to gas source.Adjust air intake
2)turn on gas
3)light burner with striker
4)adjust air supply
5)heat loop until entire wire glows red
6)cool loop

A student properly flames her loop but then forgets to let it cool. Instead, she immediately inserts the hot loop into her broth culture. What is the most important issue associated with this mistake?

The hot loop may create aerosols when it touches the culture.When the hot loop touches the liquid broth culture, it will cause some of the broth (and bacteria) to boil briefly, creating a bacteria-containing aerosol. These airborne bacteria could enter the student's respiratory tract or land on her skin. If you hear a hissing sound when you place the loop into the broth culture, you'll know you haven't cooled the loop sufficiently.

If a student forgets to flame the mouth of a broth culture tube before transferring culture to it, what might result?

The medium in the tube might become contaminated.Heating the mouth of the test tube creates an air current that helps prevents airborne bacteria from falling into the tube while the tube's cap is off. If airborne bacteria do fall into the tube, they will contaminate the medium.

The cap of the tube was not handled correctly.

The student placed the cap from the tube on the bench top. This may have contaminated the cap. Once the cap is placed back on the tube, the bacteria on the cap may contaminate the culture within the tube. Always hold caps in the hand that holds the loop. This minimizes the chance that environmental microbes might contaminate the cap.

What error in aseptic technique most likely caused this contaminant colony to grow on this agar slant?

Failure to flame the mouth of tube.this yellow colony most likely originated from an airborne bacterium that landed at the top of the agar. Flaming the mouth of the tube prevents airborne bacteria from entering the tube while it is uncapped. Airborne bacteria may fall anywhere on this slant and grow to form a colony.

This video shows a student incorrectly obtaining bacteria from a colony on a Petri plate. What has he done incorrectly?

The lid is too far away from the plate.the lid should partially cover the plate while the student is removing inoculum, as shown in this photograph. This technique prevents airborne microbes from contaminating the surface of the agar.

A student inoculated an agar slant with inoculum from a colony on a Petri plate. The slant was incubated at an appropriate growth temperature for this species for 2 days. The results are shown in this photograph. Which hypothesis best explains why there is so little growth on the agar?

The loop dug into the agar during inoculation.notice the small gouge in the agar at the bottom of the slant, as indicated by the arrow in this photograph. The student most likely gouged the agar with the loop, depositing a good portion of the inoculating cells under the surface of the agar. After the gouge, very few cells were left on the loop to inoculate the rest of the slant surface.

Transfer to an agar deep involves a slightly different procedure from transfer to an agar slant. Which of the following steps is unique to inoculating an agar deep?

An inoculating needle is used to stab the inoculum into the agar.An inoculating needle is used for inoculating an agar deep. The agar deep medium is stabbed, not streaked. This technique is used in some biochemical tests to place the bacteria within the medium, rather than on the surface.

In the 1850s, Louis Pasteur developed the concept

of the pure culture (an axenic culture)

in the 1880s, Robert Koch developed

the procedure of streaking for isolation.

streaking for isolation

the purpose of this procedure was to obtain isolated colonies

An isolated colony is

composed of millions of cells—all identical and all descendants from a single cell or group of identical cells.

Streak plating is

used to evaluate the purity of a culture
to catalog the diversity of species within a sample
to separate species from a mixed or contaminated sample
to document the characteristic growth pattern of microbe colonies.

Which of the following observations would suggest that a plate was inoculated with a pure (axenic) culture?

Isolated colonies are all white in color and about the same size.Bacterial species tend to produce distinctive-looking colonies. If all colonies on the plate appear identical, then it's likely the inoculum was pure.

Imagine that you forgot to flame the loop before streaking the inoculum from the first quadrant into the second quadrant. What is the most likely consequence of this error?

too much bacterial growth outside the first quadrant.Proper streak plating technique deposits progressively smaller amounts of bacteria in each quadrant. This is done by having the loop pick up only a small amount of bacteria from the previous quadrant's streaks. Flaming the loop between streaks ensures that the loop starts clean and that only this small amount of bacteria is used to inoculate the next quadrant.

A student INCORRECTLY streaks the plate according to the diagram below, creating streaks in the order: a, b, c, d. Which of the following is a streaking error shown in the illustration?

Streak d does NOT draw inoculum from streak c. For a plate to be streaked correctly, streak d must draw inoculum from streak c. In this example, streak d draws inoculum from streak a instead. This will draw too much bacteria into quadrant 4 and produce few, if any, isolated colonies.

Which of the following images shows how to properly open a Petri plate to streak the agar?

Lifting the lid of the Petri plate only enough to get the loop inside keeps the agar protected. Most of the lid remains right above the agar, and this prevents airborne microbes from falling into your culture.

Where should you label your Petri plate with information about the culture?

All labeling should be done on the bottom portion of the agar plate. This ensures that the written information stays with your culture, even if the lid gets accidentally exchanged with another plate.

Imagine you're streaking a plate. You've just finished the first streak (streak a). Arrange the next steps of the procedure in their correct order.

1)flame loop & let cool
2 lift lid of petri plate
3 streak across streak a & into quadrant 2
4 streak within quadrant 2
5 close petri plate lid & rotate plate

A student streaks a plate using inoculum from a broth culture tube. The culture tube was labeled "Escherichia coli," indicating that the culture was composed of a single bacterial species. Which of the following best explains the unexpected plate result shown here after incubation?

The culture in the tube was contaminated.E. coli colonies should all have the same color, size, shape, and margins. However, there are colonies on this plate that have different color, size, and shapes. This means that another bacterial species is present—a contaminant. Notice, too, that both colonies are growing along the streak lines. This suggests that both species came from the broth tube. Airborne contaminants introduced during streaking would, in contrast, be more randomly scattered about the plate.

If a correctly streaked plate were INCORRECTLY incubated right side up (instead of upside down), which of the following plate results would you most likely see?

During incubation, water tends to condense in the lid of Petri plates. If the plate is incubated in a right-side-up position, this condensation will "rain" onto the surface of the agar. The water on the agar will create a soupy mess of bacteria instead of isolated colonies. Incubating the plate upside down keeps water condensation in the lid and away from your culture.

A student performs a streak plate using inoculum from a broth culture tube. The culture tube was labeled "Escherichia coli," indicating that the culture was composed of a single bacterial species. Which of the following best explains this unexpected plate result seen after incubation?

The plate was exposed to airborne contaminants.When a plate is exposed to airborne contaminants, the contaminating microbes settle all over the plate. After incubation, the colonies from these contaminants will be all over the plate also, not just following the streak lines.

A student performs a streak plate using inoculum from a broth culture tube. The culture tube was labeled "Escherichia coli," indicating that the culture was composed of a single bacterial species. Which of the following best explains this unexpected plate result seen after incubation?

The loop wasn't flamed between quadrant streaks.You want to spread only a tiny fraction of the bacteria in one quadrant to the next, so it's important to flame and cool the loop between each streaking step. This ensures thinning of the bacteria with each streak, which ultimately produces isolated colonies after incubation.

A streak plate was prepared from an unknown culture. Which of the following best explains this plate result seen after incubation?

The unknown culture contains two species of bacteria.There are multiple distinct colonies along the streak lines consistently in each quadrant. This suggests that more than one microbe was present in the initial inoculum.

Observe the micrograph and draw conclusions about the pictured bacterium. cell body that is long and squiggly with flagella at both ends of the body

the indicated bacterium is amphitrichous flagella
the indicated bacterium is motile

Observe the micrograph and draw conclusions about the indicated bacterium. Round cell body surround by capsule

the indicated structure could serve as a virulence factor
is resistant to drying (desiccation)
is a capsule
can be used for adhesion

What can you conclude about the pictured bacteria? Cell body is retangular and in a row

if a gram stain was performed on this bacterium within 24 hrs of a fresh culture, it would most likely be gram positive
the bacteria are bacilli
structure is an endospore

Observe the micrograph and draw conclusions about the indicated bacterium. cell body is round with what looks like cilia all around the body

bacterium has a peritrichous flagella movement
bacterium is motile

What type of inoculating instrument is used to transfer a bacterial culture to an agar deep?

Inoculating needle

Which type of media should be inoculated using a zig-zag motion across the surface of the media?

Agar slant

Which of the following describes the growth of Serratia marcescens in nutrient broth?

The growth was distributed both on the surface and throughout the broth, creating a turbid appearance.

After removing the cap from the culture tube it is recommended that the neck of the tube is briefly passed through the flame of the Bunsen burner. What is the purpose of this step?

This creates an upward movement of airflow out of the tube that prevents contaminating organisms from traveling into the tube from the outside air

After incubating your culture for two days, you suspect that your culture is contaminated. Which method(s) would help you determine if this is the case?

Microscopic examination of cellular morphology
Differential staining techniques, such as the Gram stain
Subculture onto an agar plate

After incubating an agar slant containing S.marcescens, you observe growth lacking orange-red pigmentation. Is this growth necessarily that of a contaminating organism?

No, young S.marcescens sometimes have not accumulated sufficient pigment to appear orange-red

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