School of Biomedical Sciences Overview

Lecture Information

• Instructor: Dr. Eva Hatje

• Course: LQB362 Fundamentals of Microbiology

• Contact Email: e.hatje@qut.edu.au

Acknowledgements

• Acknowledgment of the Turrbal and Yugara people, the traditional owners of the land where QUT is located.

• Respect paid to their Elders, customs, and oral traditions.

Learning Resources

Textbook References

• Microbiology: an Introduction (Thirteenth Edition) by Tortora, G. J., Funke, B. R., & Case, C. L.

  • Chapter 3: Focus on Gram staining

• Brock Biology of Microorganisms (Sixteenth Edition) by Madigan, M. T., et al.

  • Chapter 29: Identification of microorganisms

• Acknowledgment: A/Prof Francesca Frentiu for slide contributions.

Learning Outcomes

1. Steps in identifying infectious agents from clinical specimens, focusing on collection and transport considerations.

2. Microbial identification methods: traditional culture techniques, biochemical tests.

3. Methods for microbial identification: nucleic acid detection, microbial antigens, and host antibodies.

4. Overview of MALDI-TOF technology and its application in diagnostic microbiology.

5. Timeframes for microbiological testing and diagnosis of infectious diseases.

Laboratory Diagnosis of Infectious Diseases

Methods of Diagnosis

• Microscopy

  • e.g., Gram staining

• Culture Techniques

  • Detection and isolation of infectious agents

• Serology

  • Antigen and antibody detection

  • IgM and IgG interpretation for infection status.

• Diagnostic Tests

  • ELISA, PCR processes, fluorescent antibody assays.

Specimen Types and Collection

• Types of Specimens: Blood, urine, CSF, sputum, etc.

  • Collected based on the infection suspected.

• Essential Identifiers on Specimens:

  • Patient’s Full Name, Date of Birth, Collection Time & Date, and Specimen Type.

• Collection Considerations:

  • Aseptic technique to prevent contamination, especially for body sites with resident microbiota.

• Storage and Transport Conditions:

  • Timeliness in processing samples is crucial (e.g., swabs <24 hours; blood <2 hours).

Microbial Identification Techniques

Conventional Techniques

• Gram Staining

  • Classifies bacteria based on cell wall properties.

  • Provides information on Gram reaction, shape, and size.

• Culture Media:

  • Selective and differential media can assist in presumptive or confirmatory identification.

Biochemical Tests

• Several specific tests include:

  • Catalase Test: Differentiates staphylococci (positives) from streptococci (negatives).

  • Oxidase Test: Identifies the presence of cytochrome C oxidase.

  • Indole Test: Differentiates certain bacteria within Enterobacteriaceae.

  • Coagulase Test: Differentiates S. aureus from other staphylococci.

  • Urease Test: Differentiates members of the Proteeae family.

  • Citrate Test: Examines the organism's ability to utilize citrate.

  • Triple Sugar Iron (TSI) Test: Determines fermentation capabilities and sulfur reduction.

Advanced Identification Methods

Automated Systems

• Vitek 2 Identification

  • Automated system using 'cards' for rapid biochemical tests and identification.

Recent Technologies

• MALDI-TOF MS (Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass Spectrometry)

  • Identifies microorganisms by analyzing the ions of their chemical structure against a database for accurate identification.

Nucleic Acid Detection Techniques

Methods of Detection

• Conventional PCR

  • Extract DNA and amplify for identification.

• 16S rRNA PCR

  • Targets the ribosomal RNA gene for species identification.

• Real-Time PCR (qPCR)

  • Combines detection with amplification to quantify microbial loads.

• Reverse Transcriptase PCR

  • For RNA viruses, converts RNA into cDNA for analysis.

Diagnosis in Absence of Culture

• Serological Tests: Detection of antigens/antibodies via assays like ELISA.

• Detection of DNA: Capture specific sequences without needing the organism to be cultured.

Summary of Key Points

• Identification of infectious agents requires systematic collection, transport of samples, and a variety of microbial identification techniques (biochemical, serological, nucleic acid detection).

• Advances like MALDI-TOF MS and automated systems enhance accuracy and efficiency in microbial diagnosis.