Diagnosing infection tutorial Flashcards
Learning outcomes
- Identify the range of diagnostic tests used in microbiology and when to request the different types in common situations.
- Explain the concepts of test sensitivity and specificity, and the impact of the patient group/population tested on predictive values of tests.
- Describe the difference between Gram stain and microscopy and culture, and the implications of the Gram stain result in terms of preliminary identification of the pathogen and empirical antibiotic therapy -Demonstrate an understanding of the key elements of prescribing an antimicrobial, including: obtaining microbiological cultures Understand how inflammatory markers and other investigations are used to diagnose and monitor the results
- Understand how to request and interpret basic diagnostic tests that can guide antimicrobial therapy (e.g. microbiology)
The diagnostic pathway
- Patient signs and symptoms lead us to…
- A differential diagnosis. We use this to choose….
- Diagnostic investigations. For infections these can be microbiological or non-microbiological (e.g imaging, bloods etc). The investigations seek to….
- Confirm, or refute the items on our differential diagnosis list.
Range of microbiological tests
- Immediate: Point Of Care Tests POCT: 0-15 mins
- Super-Quick lab tests: Microscopy with or without staining: 1-2 hours
- Quick lab tests: Nucleic acid amplification, automated serological tests: 2-24 hours.
- Standard microbiological tests: Culture based methods: 18-48 hours
- Slow tests: Epidemiological typing, Whole Genome Sequencing, samples to reference labs, slow to grow organisms: 2-10 days
The time it takes from taking a sample from a patient to getting a result from the lab is called the Turnaround Time or TAT.Note timings are end to end-assay times in the lab and often a small proportion of the Turnaround Time of a test. This also explains the wide variety in TAT.
For culture based results remember that some bacteria grow more quickly than others. This can sometimes be affected by the quantity of organisms in the sample to start with.
Gram staining
Staining is necessary as bacterial cells are essentially transparent. The principal stains are the crystal violet and iodine which are applied first. These bind to cell wall structures (peptidoglycan) of bacteria. The acetone washes away all of this stain from gram negative bacteria, but it is retained in gram positive bacteria. This is because the gram positive species have thicker peptidoglycan walls, and the gram negative species have an outer cell membrane which is washed away by the acetone. The neutral red is a counter stain. The gram positive species are holding onto the violet/ iodine stain so they don’t take up the counter stain and remain purple in colour. The gram negative species take up the counter stain and show up pink-red.
So afterstaining we can see a shape and we categorise bacteria according to this shape-ball like cocci or longer, thinner rods.
Prevalence, sensitivity and specificity
- Prevalence: the proportion of people in the population who have the disease. (50% in our example)
- Sensitivity: the ‘True positive’ rate [A/ (A+C) x 100= 90%]
- The probability that a test will be positive in a person who has the disease
- A sensitive test will produce very few ‘false negatives’ (negative test results in people withthe disease)
- Specificity: the ‘True negative’ rate [ D/ (B+D) x 100= 95%]•The probability that a test will be negative in a person who does not have the disease
- A specific test will produce very few ‘false positives’ (positive test results in people withoutthe disease)
- Sensitivity and specificity are characteristics of the test, irrespective of what population it is used in.
PPV and NPV
- Positive Predictive Value: the probability that the person with a positive test result has the disease
- Negative Predictive Value: the probability that the person with a negative test result does not have the disease
