Podcast
Questions and Answers
What is the role of the Lewis b antigen in H.pylori infections?
What is the role of the Lewis b antigen in H.pylori infections?
- It enhances the immune response against H.pylori.
- It serves as a receptor for attachment of H.pylori to gastric cells. (correct)
- It protects gastric cells from bacterial adhesion.
- It is a type of antibiotic used in treatment.
What is a primary characteristic of gnotobiotic animals?
What is a primary characteristic of gnotobiotic animals?
- They require a high level of standard care for maintenance.
- They are selectively bred for resistance to pathogens.
- They have fully developed mucosa-associated-lymphoid tissue.
- They are raised without any exposure to microbes. (correct)
Which statement best describes specific-pathogen-free (SPF) animals?
Which statement best describes specific-pathogen-free (SPF) animals?
- They have never been exposed to any microbes.
- They are living under constant sterile conditions.
- They possess immunity from prior infections by all pathogens.
- They are only free of a particular pathogen while exposed to others. (correct)
What is a significant limitation of using gnotobiotic animals in research?
What is a significant limitation of using gnotobiotic animals in research?
Why is it essential to fully characterize bacterial strains before conducting animal experiments?
Why is it essential to fully characterize bacterial strains before conducting animal experiments?
What ethical consideration must be made when conducting experiments with animal models?
What ethical consideration must be made when conducting experiments with animal models?
What could be a reason for the cost of maintaining gnotobiotic animals?
What could be a reason for the cost of maintaining gnotobiotic animals?
What primary insight have gnotobiotic animals provided in the study of microbes?
What primary insight have gnotobiotic animals provided in the study of microbes?
What does a lower LD50 value indicate about a bacterium?
What does a lower LD50 value indicate about a bacterium?
Why might LD50 and ID50 values lack sensitivity according to the limitations discussed?
Why might LD50 and ID50 values lack sensitivity according to the limitations discussed?
In comparing the infectious nature of two bacteria, which factor must be considered when using ID50 values?
In comparing the infectious nature of two bacteria, which factor must be considered when using ID50 values?
What is a major limitation of using LD50 and ID50 values across different diseases?
What is a major limitation of using LD50 and ID50 values across different diseases?
What is the ID50 value of the bacterium that causes cholera in humans?
What is the ID50 value of the bacterium that causes cholera in humans?
Which bacterium is considered more infectious based on the ID50 values provided?
Which bacterium is considered more infectious based on the ID50 values provided?
What is the typical ID50 value range for bacterial dysentery?
What is the typical ID50 value range for bacterial dysentery?
What does having a high ID50 value suggest about a bacterium's ability to cause disease?
What does having a high ID50 value suggest about a bacterium's ability to cause disease?
What is necessary to achieve a polarized monolayer in tissue culture cells?
What is necessary to achieve a polarized monolayer in tissue culture cells?
What discrepancy may arise when comparing tissue culture cells to in vivo tissue cells?
What discrepancy may arise when comparing tissue culture cells to in vivo tissue cells?
What is often overlooked in the evaluation of polarized monolayers in tissue culture?
What is often overlooked in the evaluation of polarized monolayers in tissue culture?
How can cultured cell lines be effectively utilized despite their limitations?
How can cultured cell lines be effectively utilized despite their limitations?
What is a common mistake made when interpreting results from tissue culture studies?
What is a common mistake made when interpreting results from tissue culture studies?
What specific characteristic is observed in nonconfluent tissue culture cells?
What specific characteristic is observed in nonconfluent tissue culture cells?
What is the role of extracellular matrix substitutes in tissue culture?
What is the role of extracellular matrix substitutes in tissue culture?
Why is it important to confirm the characteristics of polarized cell monolayers?
Why is it important to confirm the characteristics of polarized cell monolayers?
What role does gentamicin play in the invasion assay?
What role does gentamicin play in the invasion assay?
What is the primary measurement obtained from the invasion frequency calculation?
What is the primary measurement obtained from the invasion frequency calculation?
What does a lack of colonies form on agar plates from the second and third sets of wells indicate?
What does a lack of colonies form on agar plates from the second and third sets of wells indicate?
In the context of the gentamicin protection assay, what does the presence of colonies only on plates from the second set of wells suggest?
In the context of the gentamicin protection assay, what does the presence of colonies only on plates from the second set of wells suggest?
Which of the following is NOT a step in the invasion assay procedure?
Which of the following is NOT a step in the invasion assay procedure?
What can be inferred if there is a significant variation in the ratio of gentamicin-resistant CFU to the total CFU?
What can be inferred if there is a significant variation in the ratio of gentamicin-resistant CFU to the total CFU?
In which situation would centrifugation steps be necessary during the invasion assay?
In which situation would centrifugation steps be necessary during the invasion assay?
What defines a bacterial strain as a mutant in the context of this assay?
What defines a bacterial strain as a mutant in the context of this assay?
What is one of the main advantages of using organ cultures over tissue culture cells?
What is one of the main advantages of using organ cultures over tissue culture cells?
Why might organ cultures be difficult for long-term experiments?
Why might organ cultures be difficult for long-term experiments?
What specific example of organ culture research was mentioned?
What specific example of organ culture research was mentioned?
Why have researchers turned to developing artificial organ cultures?
Why have researchers turned to developing artificial organ cultures?
What process occurs in making artificial skin equivalents?
What process occurs in making artificial skin equivalents?
What has the popularity of cosmetic surgery provided for scientific research?
What has the popularity of cosmetic surgery provided for scientific research?
What has caused a decrease in the availability of fallopian tube and uterine tissues?
What has caused a decrease in the availability of fallopian tube and uterine tissues?
What is the outcome of keratinocyte seeding in artificial skin development?
What is the outcome of keratinocyte seeding in artificial skin development?
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Study Notes
Animal Models in Bacterial Infection Research
- Leb mice, carrying the human Lewis b antigen, are used to study Helicobacter pylori infections.
- Lewis b antigen acts as a receptor for H. pylori attachment to gastric cells and mucin.
- Germ-free animals, raised in sterile environments, lack bacteria and have underdeveloped mucosal-associated lymphoid tissue (MALT).
- These animals are expensive to maintain and lack immunity due to prior exposure.
- Specific-pathogen-free animals are raised in an environment free of a specific pathogen but exposed to other microbes.
- They are useful for studying host response without pre-existing immunity.
- Gnotobiotic animals are crucial for understanding immunity in "truly naive" animals and the nature of commensalism.
Measuring Bacterial Infection in Animal Models
- Ethical considerations are crucial when using animal models in research, as they involve bacterial infections.
- Justification for animal experiments:
- Characterize bacterial strains before conducting experiments
- The lower the LD50 (lethal dose) or ID50 (infectious dose), the more lethal or infectious the bacterium.
- LD50 and ID50 offer valuable measures of virulence but have limitations:
- They lack sensitivity and may not always reflect individual virulence determinants.
- They require a large number of animals for testing.
- They provide a relative measure of virulence when comparing strains or mutants, but can be misleading when comparing different diseases.
- For example, cholera has a higher ID50 than bacterial dysentery, but cholera can be fatal while dysentery is less lethal.
Tissue Culture Models
- Tissue culture cells can be used to study bacterial infections but have some limitations:
- Protein distribution can differ from in vivo, as cells lack differentiated surfaces.
- Polarization of monolayers may not accurately reflect in vivo tissue.
- Though they have limitations, tissue culture cells are useful tools for discovering new phenomena.
- They can generate hypotheses that can later be tested in animals.
- Results from tissue culture cells should not be directly extrapolated to human diseases.
Invasion Assay
- A gentamicin protection assay is used to measure bacterial invasion.
- Gentamicin kills extracellular bacteria and does not penetrate mammalian cells, allowing the measurement of internalized bacteria.
- Invasion frequency is determined by comparing gentamicin-resistant CFU to cell-associated CFU.
Interpretation of Invasion Assay
- Bacterial mutants defective in adhesion factors will show no colonies on agar plates for the second and third sets of wells.
- Mutants able to adhere but defective in invasion factors will produce colonies on plates from the second set but not the third.
- Comparing adherence and invasion frequencies for wild-type and mutant bacteria helps determine the nature of the defective virulence factor.
Organ Cultures
- Offer a better model of natural infection than tissue culture cells as they contain multiple cell types, including immune cells.
- They provide a more accurate representation of what occurs in vivo but can be difficult to obtain and maintain.
- Ex vivo organ cultures using human gastrointestinal tissue are useful for studying pathogens like Campylobacter jejuni and its flagellum-mediated adherence.
Source of Tissues and Organs
- Skin cells for research are readily available due to cosmetic surgery.
- Fallopian tube and uterine tissues are available from hysterectomies, but availability has decreased.
- Obtaining tissues like liver or heart is limited.
Artificial Organ Cultures
- Artificial organ cultures are being developed to overcome limitations of donor-derived organ cultures.
- Examples include artificial skin equivalents cultured from human foreskin fibroblasts and keratinocytes.
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