Questions and Answers
What is the primary focus of phenotypic methods in microbiology?
Which of the following is NOT a phenotypic morphology characteristic?
What is a disadvantage of using microscopy in microbiology?
What is an advantage of culturing in microbiology?
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Which of the following is a phenotypic biochemical reaction characteristic?
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What is a disadvantage of culturing in microbiology?
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What is the purpose of concentration techniques in pathogen diagnosis?
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Which of the following techniques is used for larval identification?
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What is a common application of flotation techniques in parasitology?
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What is the primary goal of quality examination of parasites?
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Which of the following is NOT a concentration technique used in parasitology?
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What is the primary advantage of using a differential stain over a simple stain?
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What is a drawback of using a simple stain?
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Which of the following stains is an example of a differential stain?
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What is a characteristic of the Hematoxylin and Eosin (H&E) stain?
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What is a disadvantage of using a differential stain?
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What type of components are stained by Hematoxylin?
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What is the charge of the components stained by Eosin?
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Which cellular structure is stained by Hematoxylin?
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What is the reason for different cellular structures having different affinities for H&E stains?
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What type of stain is Eosin?
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What is the main advantage of using Romanowsky stain?
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What types of cellular structures are stained by the Romanowsky stain?
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What is the composition of the Romanowsky stain?
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What is a characteristic of the Romanowsky stain?
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What is the purpose of using the Romanowsky stain?
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What type of stain is used to visualize organisms with impenetrable cell walls?
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Which stain uses a negative staining technique to visualize capsules?
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What is the purpose of heating malachite green dye in the Endospore stain?
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What is the purpose of using a mordant in the Flagella stain?
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Which of the following organisms is typically stained using the Acid fast stain?
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What does Lactophenol Cotton Blue stain target in the cell wall of fungi?
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What is the resulting color of fungal cell walls when stained with Gomori Methenamine Silver (GMS)?
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What type of biological molecules does Periodic acid Schiff (PAS) stain detect?
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Which stain is commonly used to detect Protozoa?
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What is the purpose of using Lactophenol Cotton Blue stain in microbiology?
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What is essential for successfully isolating pathogens from clinical samples?
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What type of media is used for bacterial cultivation?
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What is the purpose of providing optimal atmosphere for bacterial cultivation?
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Why is it important to provide optimal temperature for bacterial cultivation?
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What is the term for the process of isolating bacteria from clinical samples?
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What is an advantage of using culturing in microbiology?
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What is a limitation of culturing in microbiology?
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What is a disadvantage of culturing in terms of time?
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What is a financial disadvantage of culturing in microbiology?
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What can be assessed using culturing in microbiology?
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What is the optimal pH range for the growth of most pathogenic bacteria?
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What is the primary source of carbon and nitrogen for microorganisms?
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What is the term for the gaseous requirements of microorganisms?
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What is the temperature range at which most microorganisms grow best?
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What is the term for the minimum requirements necessary for microbial growth?
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What type of media is used to distinguish different bacterial groups?
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Which type of media is suitable for the growth of viruses?
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What is the purpose of using solid media?
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Which type of media contains inhibitory components for certain groups of microorganisms?
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What is the primary purpose of using liquid media?
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Match the media to correct agar.
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A routine culture sample for gram negative bacteria require a differential and selective medium
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Other phenotypic characteristics include ______. (Hint: total of 5)
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Study Notes
Phenotypic Methods
- Based on observable physical or metabolic characteristics
- Include:
- Microscopic morphology
- Staining characteristics
- Colony morphology
- Environmental growth requirements
- Biochemical reactions
- Antimicrobial susceptibility
Phenotypic Morphology Characteristics
- Shape
- Size
- Structure
- Organization
Phenotypic Biochemical Reaction Characteristics
- Carbon utilization
- Production of:
- Acid
- Gas
- Other
Microscopy
- Advantages:
- Determination of cell/tissue morphology
- Cellular association
- Pathogen morphology
- Impression of disease stage and severity
- Rapid analysis
- Disadvantages:
- Mild/chronic infection may not be detected
- Low sensitivity
- Low specificity
Culturing
- Advantages:
- Isolation of pathogen allows further testing
- Permits full taxonomic identification of pathogen
- Disadvantages:
- Not all pathogens are cultivable
- Requires high expertise
- Requires several media and lab conditions
Concentration Techniques
- Used to increase the concentration of pathogens in a sample for diagnosis
- Commonly employed for diagnosing parasites in feces, vomit, and sputum samples
- Examples of concentration techniques:
- Flotation techniques
- Sedimentation techniques
- Baermann test (specifically used for larval identification)
Quality Examination of Parasites
- Involves determining the type of eggs or larvae present in a sample
Staining Techniques
- Simple stain uses only one dye to identify morphology and cellular arrangement in cells.
- Drawback of simple stain: may not stain all components of the cell, making interpretation difficult.
Differential Stain
- Uses more than one dye to identify morphology and cellular arrangement in cells.
- Allows distinction between different cell types and structures.
- Drawback: requires a multistep method, more time, reagents, and expertise.
Examples of Staining Techniques
- Methylene blue stain
- Gram stain
- Acid fast stain
- Hematoxylin and eosin stain (H&E)
- Negative stain
- Romanowsky stain
- Others...
Hematoxylin and Eosin Stain (H&E)
- Hematoxylin and eosin stain is a common tissue stain used to visualize different cellular structures.
- The stain works due to the varying affinities of cellular structures for the dyes, depending on their chemical composition.
Properties of Hematoxylin
- Hematoxylin is a basic stain that binds to acidic components (negative charged).
- It stains nuclei and chromatin.
Properties of Eosin
- Eosin is an acidic stain that binds to basic components (positively charged).
- It stains muscle, red blood cells, and extracellular components.
Romanowsky Stain
- Also known as commercial Diff-Quick
- A modified Wright-Giemsa stain
- Used to study the adherence of pathogenic bacteria to cells
- Examples of cells stained: blood cells
- Stains chromosomes
- Allows for cytoplasmic detailing
- Composition: Eosin and Methylene blue
- Typically used to stain: muscle, red blood cells, extracellular components
Stains Used in Bacteriology
- Acid fast stain is used to stain organisms with impenetrable cell walls, and it involves the use of heat.
- Examples of organisms that can be stained using acid fast stain include Mycobacterium and Cryptosporidium.
Capsule Stain
- Capsule stain is a negative staining technique.
- It involves contrasting a translucent substance against a darker colored background.
- The darker colored background is achieved using dyes such as nigrosine or Congo red.
Endospore Stain
- Endospore stain is used to dye spores.
- The staining process involves heating malachite green dye.
Flagella Stain
- Flagella stain is used to thicken flagella.
- A mordant is used in the staining process.
Fungal and Parasite Stains
- Lactophenol Cotton Blue stain: specifically stains chitin present in the cell wall of fungi
- Gomori Methenamine Silver (GMS) stain: produces a dark brown staining of the fungal cell wall and surrounding tissue
- Periodic Acid Schiff (PAS) stain: used to detect polysaccharides (e.g. glycogen) and mucosubstances (e.g. glycoproteins, glycolipids, mucins) in tissues
- Wheatley's Trichrome stain: used for the detection of Protozoa
Sample Cultivation
- Many pathogens can be isolated from clinical samples in the laboratory using simple or specialized media.
- The process of isolating bacteria from clinical samples is known as cultivation or isolation of bacteria.
- Media used for cultivation can be either liquid (broth medium) or solid (agar medium).
- Optimal nutrients, atmosphere, and temperature must be provided to successfully isolate the pathogens.
Disadvantages of Culturing
- Culturing is a time-consuming process that can take anywhere from 2 days to over 10 days
- It is an expensive procedure
- Not all bacteria can be cultured, which limits its application
- The process requires in-depth studies on microbial characteristics and genetics
- Culturing is not an effective method for assessing antimicrobial resistance (AMR)
Environmental Factors
- Nutrients, such as glucose and nitrates, are essential for bacterial growth
- pH levels between 7.2-7.4 are optimal for most pathogenic bacteria to thrive
- Ionic strength and osmotic pressure also influence bacterial growth
- Light can affect bacterial growth, although the effect varies among species
- Most bacteria are mesophilic, growing best at temperatures between 30-37 °C
- Bacteria require specific gaseous conditions, including oxygen and carbon dioxide, to grow
Nutritional Factors
- Carbon and nitrogen are obtained from various sources
- Phosphate, sulfate, potassium, magnesium, calcium, and iron are essential nutrients for bacterial growth
- Trace elements, vitamins, and purines/pyrimidines are also necessary for bacterial development
Culture Methods
- Acellular media are used to culture bacteria, fungi, and some parasites
- Cellular media (cell cultures) are used to culture viruses and intracellular bacteria
Types of Media
Physical State
- Solid media: allow for the observation of phenotypic characteristics
- Liquid media: enable the use of different nutrients or the inhibition by antimicrobials
Purpose
- General-purpose media: support the growth of a large number of non-fastidious bacteria
- Selective media: contain inhibitory components to suppress the growth of certain groups of microorganisms
- Differential media: allow for the distinction between different bacterial groups based on their growth patterns on the medium
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Description
This quiz covers phenotypic methods in microbiology, including morphology, staining, colony morphology, environmental growth requirements, biochemical reactions, and antimicrobial susceptibility.