BIO250L LAB 2

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Grand Canyon University *

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250L

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Biology

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Jan 9, 2024

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“ Lab 2 Culturing & Aseptic Technique BIO250L ” Student Name: MA ERICKA RENEE SAUVE Access Code (located on the lid of your lab kit): UAWQHC Lab Report Format Expectations Utilize college-level grammar and professional formatting when completing this worksheet. Submissions without proper formatting, all required photos, or sufficient responses will be rejected. Pre-lab Questions 1. This lab includes two experiments that each look at different aspects of culturing microorganisms onto growth media. What concepts does each experiment focus on? Why do you think these concepts are important to the field of microbiology? The first experiment focuses on agar plate preparation and bacterial inoculation. It explains the importance of aseptic techniques and bacteria culture to obtain pure colonies. Yes, this is critical in microbiology for accurately identifying and studying microbial species without contamination. The second experiment focuses on transferring bacteria to stab tubes and agar plates, highlighting different growth environments and patterns. It establishes the abilities required for culturing and observing bacteria in semi-solid and solid media. These concepts are also very important in microbiology for diagnosing infections and researching bacterial behavior. In this lab, the experiments will allow or call for the use of four inoculation tools. List these tools and provide a scenario in which you would use each tool. ○ The loop is used to streak microorganisms on a plate, while the needle is used to pick and transfer colonies to agar plates. Pipettes are used to take liquid samples precisely. Swabs are used to pick microbial colonies from the body's tissues. The spreader is used to spread bacterial colonies evenly on a plate. 2. Describe a piece of equipment used in biotech labs to extend the growth pattern of microbes. Focus particularly on those used with E. coli used to make human insulin. Bioreactors are used to provide optimal environmental conditions for E. coli growth at Biotech. 3. You’re a physician trying to isolate bacterial colonies from the human gut in an attempt to diagnose a gastrointestinal infection. You streak your sample on a growth media containing glucose, amino acids, and salts containing both sulfur and phosphorus with a pH of 7 . You incubate the plates in aerobic conditions at 37 ˚C for three days, at which point you can see clear bacterial colonies forming on the plate. Would you feel confident in stating that you had successfully cultured all the bacteria from your gut sample? Why or why not?

“ Lab 2 Culturing & Aseptic Technique BIO250L ” The method described can culture many types of bacteria present in the gut. However, it may not isolate all bacteria found in the human gut. Therefore, stating that all bacteria from the sample were cultured would not be accurate without further selective and differential culture techniques. EXPERIMENT 1: AGAR PLATE PREPARATION AND BACTERIAL INOCULATION Introduction Questions 1. Describe the objectives of Experiment 1. State the goal of the experiment and the learning outcome that it is trying to convey to you as a microbiology student. The main objective of this exercise is to introduce students to the preparation of agar plates and the inoculation of bacterial cultures. The exercise aims to impart an understanding of culturing techniques and the importance of maintaining a sterile environment. In a laboratory setting, what are three ways you can properly sterilize culturing equipment? ○ Chemical sterilization with appropriate disinfectants. ○ Autoclaving method. ○ Dry heat sterilization at high temperatures. 2. What method of sterilization will you use in this experiment? The method of sterilization used in this experiment is autoclaving, which is a widely accepted and standardized practice. 3. What results do you expect to see with this experiment? For example, do you expect to see growth from all surfaces or just some? Are there particular strains you expect from the surfaces you swabbed? I expect to observe varying bacterial growth from different surfaces depending on the conditions provided with some yielding more colonies and others yielding less. The type of growth will depend on the specific environmental conditions and surfaces swabbed.

“ Lab 2 Culturing & Aseptic Technique BIO250L ” Data and Observations In the table below, report your observed growth corresponding to each plate number and its corresponding source. Then provide a photo of your growth plates. Your data should have a quantitative aspect to it, such as a count of the number of colonies that grew. Your data must also correspond to the photo you include for credit. Table 1: Colony Growth Plate Number Source Growth (Color, Amount, Shape, etc.) 1 Sink Some of them were round and a greater number of them were irregular. Colony with dominant has white color. 2 Throat Irregular with scattered Colony. Color was yellow and small. 3 Shoe Round and irregular with yellow to creamy color. 4 Airborne Regular and closely clustered with yellow color. 5 Control Yellow and white colored bacteria spread on plate. Provide a clear, high-resolution photo of your plates after incubation with the lids removed . For credit on this lab, your photo must show the growth that is reported in your tabulated data and must also include your handwritten name in the background.

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“ Lab 2 Culturing & Aseptic Technique BIO250L ” Results and Discussion 1. Describe what morphological traits (i.e., size, shape, arrangement, color, margin, etc.) you observe from the growth on your plates. Be as specific as possible in the characteristics. I observed Shapes whether it was round or irregular color arrangements whether they were clustered or scattered and their amount whether they were in small amounts or large amounts 2. Based on the morphological qualities you observed on your plates, and using any resources available to you (i.e., a textbook or internet sources such as the CDC website), look up what bacteria you may have found on the surfaces you swabbed. To identify the bacteria present on the surfaces you swabbed, I would compare the observable colony characteristics such as shape, color, size, and texture, with descriptions present in microbiology resources like books or online databases such as the CDC website. Factors like the environment where the swabs were taken can also provide clues about the lifestyle and characteristics of the bacteria present. 3. For the colonies hypothesized in Question 4, are you surprised to find this type of microbe on this surface? No, I am not surprised about the presence of Staphylococcus aureus because it is a common bacteria. 4. Looking at your control, did you perform a proper aseptic technique or were your plates contaminated? If contamination did occur, list the possible sources and how you can prevent contamination in the future. There was no bacterial growth which means proper aseptic techniques were used.

“ Lab 2 Culturing & Aseptic Technique BIO250L ” 5. Compare the colonies that grew on your swab plates to the Airborne Contamination plate. Did any of your experiment plates grow colonies similar to your Airborne Contamination plate? How would you describe the risk of airborne contamination in your experiment? While comparing experiment plates with the Airborne Contamination plate, there were similar colonies that grew, which suggests a risk of airborne contamination. This indicates the significance of aseptic techniques, The extent of similarity between the plates can indicate how significant the risk of airborne contamination was during your experiment. “ EXPERIMENT 2: BACTERIAL TRANSFER TO STAB TUBES AND AGAR PLATES Introduction Questions 1. Describe the objectives of Experiment 2. State the goal of the experiment and the learning outcome that it is trying to convey to you as a microbiology student. the objective of the experiment is to understand bacterial transfer to stab tubes and agar plates, which causes the cultivation of bacteria in different environments. The objective is for students to learn inoculation techniques, observe varying growth patterns, and maintain culture purity, crucial for identifying and studying microorganisms. 2. What results do you expect to see in Experiment 2? Expected outcomes from the experiment are successful bacterial growth in both mediums, with varied patterns in stab tubes depending on oxygen requirements and distinct colonies on agar plates, which indicates effective aseptic technique and bacterial cultivation skills.

“ Lab 2 Culturing & Aseptic Technique BIO250L ” Data and Observations Record your observations in the tables below. Table 2: Initial Reserved Plate Colony Growth Observations Plate Sample Appearance of the original colonies (morphology, etc.) 1 Irregular shape with White and yellow color 2 Irregular shape clustered and yellow in color 3 Regular round Table 3: Final Plate and Stab Tube Growth Observations Sample Form Growth (Y/N) Same Appearance as Initial Plate (Y/N) Successful Transfer? (Y/N) 1 Plate Yes Yes Yes 1 Stab Tube Yes Yes Yes 2 Plate Yes Yes Yes 2 Stab Tube Yes Yes Yes 3 Plate Yes Yes Yes 3 Stab Tube Yes Yes Yes Control Plate No Yes Yes Control Stab Tube No Yes Yes

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“ Lab 2 Culturing & Aseptic Technique BIO250L ” Arrange your plates and tubes as shown below. Remove the lid from the plate and place the stab tube next to the matching plate. Then take a high-resolution photo and insert it below with your handwritten name in the background . For credit on this lab, the growth that you report in the above table must correspond to the photo below , and the photo must be of a high enough resolution to assess the reported data. Results and Discussion 1. Review your data and determine whether or not the growth on your transfer plates have the same colony morphology (i.e., size, shape, arrangement, color, margin, etc.) as your original sample. Discuss this below. The colonies that were observed had similar characteristics such as size, shape, color, and arrangements, which indicates a successful transfer of the same microbial species. However, some colonies on the transfer plates showed different morphologies, which may have been due to possible contamination or differing growth conditions that were selected for different traits or species. 2. If there was any difference in colony morphology, what do you think could have caused this? Differences in results may arise due to unsuccessful transfer or growth of cultures, which could lead to contamination with unexpected characteristics. This, in turn, could result in inaccurate

“ Lab 2 Culturing & Aseptic Technique BIO250L ” findings and misidentification of the species, leading to a waste of time and resources. Therefore, it is crucial to ensure the consistency and purity of cultures to maintain the experiment's validity over extended periods. 3. Imagine you are performing an experiment that requires the transfer of cells to different plates which will grow bacteria over an extended period of time (days or weeks). Describe why an unsuccessful transfer or growth with a different appearance from the initial plates would be a problem in this scenario. If a transfer or growth process is unsuccessful or has unexpected characteristics, it may result in inaccurate outcomes, misidentification of the species, and a waste of time and resources. Therefore, it is crucial to ensure the consistency and purity of cultures to maintain the validity and precision of the experiment. 4. Do some research and describe two or three scenarios in which it would be preferable to use a stab tube vs. a growth plate. ( Hint : What do bacteria use to help them move? Can motility be used to help identify many medically important pathogenic bacteria such as the Enterobacteriaceae family of bacteria?) Stab tubes are useful in specific scenarios compared to growth plates. Firstly, they are ideal for studying the oxygen requirements and motility of bacteria. Bacteria with flagella can be observed easily moving away from the stab line in semi-solid media. This is particularly significant for identifying pathogenic bacteria like those in the Enterobacteriaceae family, where motility is a key diagnostic characteristic. Secondly, stab tubes provide a more stable environment for long-term culture storage as they decrease the rate of desiccation compared to open plates.

BIO250L LAB 2

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