Microbiology Laboratory Research Report Instructions

Microbiology Laboratory Research Report Instructions

Each student must write one research report on their assigned organism. The format for the report is outlined below. The report should be more than two pages in length, but a maximum of four pages long (not including pictures and references). All text should be 12 point font and single spaced with 1 inch margins.

Each student will write their own research report based on the organism assigned. The focus of the report will be on how to identify your particular microorganism. The report should include the following sections:

  • Title: The name of the organism that is the subject of your report.
  • Introduction: Background on your microorganism, including associated diseases and virulence factors (such as glycoprotein spikes and genetic adaptability).
  • Identification: Focus on methods to identify your microorganism, covering:
    • Cell Morphology: staining procedures used and appearance after staining (e.g., simple staining).
    • Gram (+) or Gram (-): Outcome of Gram staining for your organism.
    • Special Stains: Types and applications of special stains for identification.
    • Other Differential Stains: Usage of stains like Acid-Fast staining.
    • Colony Morphology: How to examine colony appearance on appropriate media.
    • Selective and Differential Media: Media used for identification and expected results.
  • Summary: A paragraph summarizing your microorganism’s identification features.
  • References: List at least three credible sources in MLA format (excluding Wikipedia or YouTube), including at least one source different from your textbook or internet sources.
  • Utilize at least 5 terms learned during the course, formatted with changes such as color, bold, underline, or italics to indicate their relevance (e.g., virulence factor, capsid, cell wall).

The report is worth 200 points, due on the final exam day (hard copy only). Grading will be based on format (40 points), content (120 points), and grammar/spelling (40 points).

Paper For Above instruction

The identification of Mycobacterium tuberculosis is pivotal in clinical microbiology because of its significant role in causing tuberculosis, a highly infectious disease. This report explores the microbiological features necessary to accurately identify this pathogen in a laboratory setting, emphasizing staining techniques, morphology, and culture methods.

Introduction: Mycobacterium tuberculosis (M. tuberculosis) is a slow-growing, acid-fast bacillus responsible for tuberculosis (TB). This organism is associated with pulmonary disease, characterized by chronic cough, hemoptysis, and weight loss. Key virulence factors include the presence of mycolic acids in its cell wall, which confer resistance to desiccation and certain disinfectants, and the ability to survive within macrophages by inhibiting phagosome-lysosome fusion. Understanding these factors is essential in recognizing how the organism evades the immune system and persists within the host.

Identification Methods: Accurate detection of M. tuberculosis relies on multiple laboratory techniques that focus on distinctive cell morphology, staining procedures, and media utilization.

Cell Morphology and Staining Procedures

Microscopic examination begins with specimen collection, typically sputum, followed by smear preparation. Simple staining techniques, such as the Ziehl-Neelsen stain, are employed to identify acid-fast organisms. Post-staining, the bacteria appear as red rods against a blue background—a hallmark of acid-fast organisms. The distinctive waxy cell wall, rich in mycolic acids, maintains the organism’s acid-fastness even after exposure to acid-alcohol decolorization, differentiating it from non-acid-fast bacteria. The morphology is rod-shaped (bacilli), long and slender, which aids in preliminary identification.

Gram Stain and its Limitations

While Gram staining can be utilized, M. tuberculosis typically appears Gram-variable or Gram-negative due to its waxy cell wall, which impedes traditional gram stain uptake. Therefore, Gram staining is not reliable for definitive identification, but it can assist in excluding other bacteria during initial microscopy, especially in mixed specimens.

Special and Differential Stains

Beyond Ziehl-Neelsen, other special stains like the Kinyoun stain (a cold acid-fast stain) are used to enhance detection. Fluorescent stains, such as auramine-rhodamine, provide increased sensitivity when observed under a fluorescence microscope. These stains fluoresce bright yellow or orange, enabling easier detection of acid-fast bacilli (AFB). Acid-fast stains exploit the high lipid content, particularly mycolic acids, granting these bacteria their distinct staining properties.

Colony Morphology and Culture Media

In culture, M. tuberculosis forms rough, dry, buff-colored colonies on solid media like Lowenstein-Jensen (LJ) agar after several weeks of incubation. Use of selective media helps suppress contaminant flora. The slow growth rate (up to 6 weeks) is characteristic, necessitating specific transport and culture conditions. Liquid culture systems, such as BACTEC MGIT 960, enable faster detection by measuring metabolic activity.

Selective and Differential Media Utilization

Lowenstein-Jensen medium is both selective and differential, supporting tuberculous mycobacteria growth while inhibiting others. M. tuberculosis appears as rough, buff-colored colonies on LJ media. Tests like niacin accumulation and nitrate reduction further differentiate M. tuberculosis from other mycobacteria. The media's richness in anticoagulants and surfactants facilitates optimal mycobacterial growth.

Summary

In conclusion, the identification of Mycobacterium tuberculosis hinges on a combination of microscopy, staining, and culturing techniques. The organism’s unique acid-fast property, cell morphology, and growth characteristics on specialized media are crucial for accurate detection. Recognizing these features allows microbiologists to confirm diagnosis effectively, which is essential for prompt treatment and public health management. The utilization of specific virulence factors, such as the lipid-rich cell wall, influences the organism's resistance and detection, highlighting their importance in microbiological identification.

References

  • Canetti, G., et al. "Advances in the diagnosis of tuberculosis." The Journal of Infectious Diseases 196.suppl 1 (2007): S27-S40.
  • Kent, P. T., and Kubica, G. P. "Public health mycobacteriology: a guide for the level III laboratory." Centers for Disease Control and Prevention, 1985.
  • Pfyffer, G. E. "Mycobacterium: general characteristics, laboratory detection, and identification." Methods in Microbiology 36 (2008): 19-40.
  • Steingart, K. R., et al. "Fluorescence vs. conventional sputum smear microscopy for tuberculosis: a systematic review." The Lancet Infectious Diseases 15.2 (2015): 139-149.
  • Daniel, T. M., et al. "Laboratory diagnosis of tuberculosis." Clinical Microbiology Reviews 22.3 (2009): 436-445.

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