Microbiology Final

Aseptic technique (definition)

A set of procedures employed in the handling of microbes without causing contamination from outside sources.

Inoculum (definition)

A small amount of microorganism used to start a new culture.

Smear prep (definition)

A smear prep is a technique used to produce a sample of microorganisms, typically for microscopy or staining. While there are many variations of the smear prep technique, depending on the final desired use of the sample, the overall key steps are the same for a majority of smear prep protocols.

Colony forming units (CFUs) (definition)

A unit used in microbiology to estimate the number of viable microorganism cells in a sample. Viable is defined as the ability to grow and multiply under the incubation conditions provided.

Colony (definition)

A visible mass of microorganisms formed by replication of a single microorganism on an agar surface, often in a petri dish.

Agar (definition)

Agar media- bacterial culture medium that contains agar as a solidifying agent and is used for growing microorganisms. Agar plate- bacterial culture medium containing agar poured into a petri dish. Agar slant- bacterial culture medium containing agar in a test tube that is allowed to solidify at an angle and forms a solid, slanted surface.

Proper plate labeling/incubation

Agar plates are labeled on the bottom in case the lid becomes separated from the actual culture. Correct labeling of an agar plate with name of organism and agar type. Labelling may also include researcher initials, sampling location, and date. Most bacteria are mesophiles (grow between 20-40°C) and are incubated at 37°C (human body temperature).

Condensation

Agar plates are made by pouring hot, sterile agar into an empty, sterile petri dish and allowing it to cool. Often, condensation will form on the lid. Condensation could fall down onto the agar surface causing contamination or dispersion of any bacterial colonies. To prevent this, plates are always incubated upside down.

Sterile (definition)

An environment devoid of any living cells.

ubiquitous

Bacteria are ubiquitous. This means they are found almost everywhere—soil, water, and even the Earth's crust.

Agar plate (definition)

Bacterial culture medium containing agar poured into a petri dish.

brightfield microscopy

Brightfield microscopes use a combination of glass lenses and light to view the specimen.

Contrast (definition)

Contrast refers to the ability to distinguish objects, such as cells, from the background.

unknown identification (purpose)

Correct identification of an unknown bacterium could help an individual patient, a community during an outbreak, or even the global population during a pandemic.The goals of unknown microbial identification are to examine live organisms of interest, aseptic technique should be followed as closely as possible to prevent the introduction of unwanted or harmful organisms to a user or into an environment, culture, or agar sample.

Differential staining (definition)

Differential staining uses two or more dyes to discriminate between cell types or structures.

Magnification (definition)

Magnification is the apparent increase in image size.

why metric in science?

Metric units are used in scientific research to ensure consistency of collected data. This gives all scientists the same standards of measurement when they conduct experiments.

Staining (purpose)

Microbiological staining increases contrast during microscopic imaging, which allows the observer to visualize microorganism cells. Variations in staining protocols can be used to tell you more about the microorganism you are viewing. Analysis of microbiological staining provides information on general cell morphology (such as cell shape and cell arrangement), or showing individual cell structures such as capsule, cell wall, and inclusions.

sources (most/least reliable)

Most to Least reliable: 1. academic journals 2. sites with references 3. institutional sites without references 4. personal sites and social media posts without references 5. websites trying to sell something

Heat fixation (purpose)

The process of applying gentle heat to a sample. Fixing a specimen preserves various cellular components in a natural state with minimal distortion, while simultaneously killing the specimen and securing it to the slide.

fluid thioglycolate media (FTM) (components)

The top of the fluid in the tube is slightly red, indicating the presence of oxygen. The middle and bottom of the tube are yellowish in color, indicating low to no oxygen present. FTM contains a small amount of agar to localize organisms in places where the conditions are most favorable for growth. Bacterial organisms will prefer to grow in specific locations within a tube of fluid thioglycolate medium.

total magnification (calculation)

Total magnification is the overall enlargement of the image of a specimen. To calculate total magnification, multiply the magnification of the ocular lens (10X) with the magnification of the objective lens.

Dichotomous key (definition)

Visual flowchart that can be used to map the series of testings that help identify an organism. Each step of a dichotomous key has 2 outcomes, positive or negative, leading to a next set of testing choices. The top of a dichotomous key would consider a broad group of organisms. The possibilities narrow at each level/step decision until arriving at a single possibility at the bottom.

scientific literacy (definition)

knowing how to apply the process of scientific inquiry so that you can find answers for yourself. To be scientifically literate is not to have mastered all the facts and concepts that scientists have discovered and explained.

Mesophile (definition)

organism that grows between moderate temperatures (20-40°C)

Gram staining (purpose, steps)

- Purpose: Gram staining is one of the most common and important bacteriological staining techniques. Gram staining is used for: Visualization of bacterial cells, differentiation of bacterial cell shapes and arrangements, identification of bacterial cell wall structures, differentiation between gram-positive and gram-negative cells. Correct technique and interpretation of a Gram stain can be critical to presumptively identifying a bacterium and can assist in choosing an appropriate antibiotic treatment. - Steps: 1. Primary staining with crystal violet 2. Fixation with Gram's iodine 3. Decolorization with ethanol 4. Secondary staining with safranin

PPE (personal protective equipment)

- lab coat or apron - gloves - goggles or lab safety glasses - closed toe and covered heel shoes - long hair should be tied up to meet proper laboratory safety protocols

basic units for length, weight, volume, and temperature

- length: meters (m) - weight: grams (g) - volume: liters (L) - temperature: degrees Celsius (°C)

types of lab hazards (summary)

- spills - splattering - flames - corrosive chemicals - sharp objects

Characteristics of microbes

-Archeae are typically found in extreme environmental conditions and share some features with Eukarya. -Bacteria have three common shapes; cocci (spheres), bacilli (rods), curviform (spirillum, spirochete, and vibrios). -Protozoa are unicellular eukaryotes that lack cell walls and obtain their nutrients by engulfing food particles. They are typically classified by their means of motility. Protozoa include flagellates, amoeba, ciliates, and non-motile apicomplexans. -Algae are photosynthetic eukaryotes and include unicellular, colonial, and multicellular forms. Algae have cell walls made of cellulose, and most live in an aquatic environment. Algae include green algae, red algae, brown algae, diatoms, euglenoids, and dinoflagellates. -Fungi can be both unicellular (e.g., yeast) and multicellular (e.g., mold) eukaryotes. Fungi have cell walls made of chitin and absorb their nutrients from the environment.

Prokaryotic and Eukaryotic microbes

-Prokaryotic cells do not contain a membrane bound nucleus and whose structure is simpler compared with a eukaryotic cell. -Eukaryotic cells contain a membrane bound nucleus and have a more complex structure with increased intracellular compartmentalization.

Subculturing technique (purpose, key steps)

-Purpose: Subculturing of a bacterial sample is the most commonly used method for obtaining pure cultures of bacterial species from mixed microbial populations. The purpose and methods of microbial isolation are important to the bacterial identification process and help us understand bacterial growth patterns. -Key Steps: 1. Choose carefully when picking which colony/colonies to subculture. Pick an isolated colony (not mixed with or directly adjacent to another colony) to avoid mixing bacterial types and contaminating the pure culture. 2. Use a small amount of colony material, rather than a large glob of bacteria, to reduce risk of cross contamination during sample acquisition. 3. Always use aseptic technique to transfer bacteria between growth media.

Quadrant streak plate method (purpose, disadvantages, key steps)

-Purpose: The quadrant streak plate technique is a common microbiology isolation procedure for bacteria in culture, but does have two noted drawbacks. First, it can be difficult to master the technique, requiring practice to achieve consistent and acceptable results. Secondly, you cannot accurately count bacteria from a quadrant streak plate. The purpose and methods of microbial isolation are important to the bacterial identification process and help us understand bacterial growth patterns. -Disadvantages: First, it can be difficult to master the technique, requiring practice to achieve consistent and acceptable results. Secondly, you cannot accurately count bacteria from a quadrant streak plate. -Key Steps: 1. Use aseptic technique to transfer a bacterial sample to an agar plate. 2. Streak the bacterial sample across one quadrant (approximately ¼ of the agar surface) in a back-and-forth motion. 3. Re-sterilize the loop and then drag the loop through the previously inoculated quadrant to pick up a small fraction of sample to the next quadrant surface. Repeat until four quadrants are completed in the same manner. 4. Incubate the agar plate for 24 hours to grow isolated bacterial colonies.

Smear prep (order steps)

1. Appropriately label slide 2. Apply bacteria to the slide using aseptic technique 3. Air drying 4. Heat fixation 5. Apply staining method

Aseptic technique steps-broth to agar (summary)

1. Pick up and heat loop 2. Pick up culture, remove cap, heat mouth of tube 3. Insert loop into culture 4. Heat mouth of culture tube, replace cap, place in rack 5. Lift lid and inoculate sterile agar plate 6. Heat loop and return to rack

evaluating a scientific claim (5 questions)

1. does it seem logical? 2. is the source credible? 3. are scientific studies cited? 4. are the experiments well designed? 5. do data match conclusions?

Four steps of the scientific method (list)

1. hypothesis 2. experiment 3. analysis 4. conclusion

Time and order of sequences/steps (hand washing)

1. remove any items on your wrists or fingers 2. turn on warm water 3. wet your hands 4. apply soap to hands 5. use nail stick or nail brush to clean nails 6. rub wet, soapy hands for at least 20 seconds 7. rinse hands 8. dry hands with a clean paper towel 9. use paper towel to turn off water 10. throw away paper towel

Pure culture (definition)

A bacterial culture containing only one strain of microorganism.

Reading graph (also in Scientific Thinking in Everyday Life article)

A graph is a visual summary of data. Most have a horizontal (X) axis and a vertical (Y) axis. Typically, the independent variable does on the horizontal axis and the dependent variable foes on the vertical axis.

Broth culture (definition)

A liquid nutrient medium used for the propagation of microorganisms.

Vegetative cell (definition)

A metabolically active and growing cell.

Types of microbes

Fungi, algae, protozoa, bacteria, and archeae - bacteria - algae-Spirogyra - protozoa-Amoeba - fungi-Saccharomyces cerevisiae (yeast) and Aspergillus (mold)

Inoculating tools (uses)

Inoculating tools must be sterilized both before and after transfer by heating to red hot in the flame of a Bunsen burner or in a microincinerator. The inoculating tool must be allowed to cool before picking up cells to be transferred, or they will be destroyed in the process. Inoculating loops are used to transfer an inoculum between tubes of broth or onto the surface of an agar slant or plate.

Bergey's Manual of Determinative Bacteriology (definition)

Internationally recognized reference book and collection of datasets for bacterial classification and identification.

Resolution (definition)

Resolution is the ability to distinguish fine detail.

SPC key steps

The SPC technique can be described as having four key steps:1.Perform a serial dilution series of a bacterial culture into a set of blank liquid media to reduce numbers.2.Transfer an aliquot of prepared dilutions to solid agar media.Spread the transferred samples across the agar surface using a spreader (often called a "hockey stick").Incubate agar plate, allowing for growth of colonies and further visual counting.

Goal of Microscopy

The goal of microscopy is to create a magnified image of objects too small to be seen with the eye alone.

Why is hand washing important

Your hands may come in contact with infectious organisms, irritating chemicals, stains, and other potentially hazardous materials. Your hands may also be the source of contamination. This can lead to poor lab experiment results. It is important to wash your hands prior to a lab exercise, after a lab exercise, and nay other times your hands have been in contact with potential hazards to maintain your own safety and the safety of others around you.

Hypothesis (definition)

a formulated statement of predicted behaviors