Bacterial culture Dr Joel Bazira Mbarara University of
Bacterial culture Dr. Joel Bazira Mbarara University of Science and Technology
Bacterial culture is the process of growing microorganisms by: – Taking bacteria from an infection site by specimen collection - in vivo – Growing bacteria in the artificial environment of the laboratory - in vitro
Why grow bacteria? • Obtain definitive identification and characterization • Grow and isolate all bacteria present in an infection • Determine which bacteria is most likely causing infection • Determine which bacteria is likely a contaminant or colonizer
Why grow bacteria 2? Isolating the bacteria enables us to: – Test antimicrobial susceptibility – Measure response to treatment – Characterize the agent – for epidemiological purposes – Store strains for future use including vaccine development
Work flow 1 2 3 4 5 6 7 Specimen collection/ reception Microscopy Inoculation of the media Incubation Plate reading Identification Drug susceptibility testing
Culture methods Streak culture – Isolation of bacteria in pure culture from clinical specimen Lawn culture – Antimicrobial susceptibility testing (disc diffusion), bacteriophage typing Liquid cultures Stroke culture – To obtain pure growth for slide agglutination; biochemical tests Stab culture – Maintenance of stock cultures Pour-plate culture – Quantification of bacteria in liquid cultures, urine sample
Rapid cultivation & automation Instrument-based systems – Periodic and continuous monitoring systems; growth detected by: • Colorimetric or fluorescent detection of CO 2 • Consumption of gasses • Fluorescent detection of growth Bioluminescence assay for viable organisms Colorimetric filtration (urine screening)
Phases of bacterial growth Log of numbers of bacteria Log growth phase Stationary phase Death phase Lag phase 0 5 Time (hr) 10
Measurement of cell growth Measure total counts – Measure both viable and non-viable bacterial cells – Direct microscopy using Gram stain; automated cell counter Measure viable counts – Measure only viable cells – Pour plate cultures to give quantitative number of viable bacteria
Measurement of cell growth Semi-quantitative methods – Give less accurate but working estimate of bacterial load to aid in decision making – Semi-quantitative urine culture; MPN test for water bacteriology Quantitative methods – Give accurate estimate of bacterial number; more exact applications – Vaccine production
Pure cultures • In theory, each colony (~107) bacteria arises from a single bacterium • Each colony consists of a pure clone of cells • Best obtained on solid media; less sure in liquid media
Challenges with Bacterial culture • • Long Turn around time- work flow Labor intensive Technically demanding – experienced staff Quality assurance… Some bacteria do not grow Aerobic vs anearobic culture One bacteria or a group of bacteria Unstable electicity
- Slides: 12