MLT 207 Les 8 pt 2

Introduction

• Discussing microbiological testing for bacterial identification starting with hemolysis.

Slide 8: Initial Testing for Gram-negative Bacteria

• Flaws of the current material considered.

  • Identifying Bacteria: Determining if bacteria are Gram-positive or Gram-negative.

  • Result: Gram-negative confirmed.

• Next Steps:

  • Procedure for non-fermenters: would use oxidase test.

  • Proceeding with the oxidase test for the current Gram-negative.

• Interpretations:

  • If oxidase positive, identify as a Gram-negative rod, not Enterobacteriaceae.

  • If oxidase negative, identify as an Enterobacteriaceae.

  • Further testing based on fermentation status:

  • Lactose Fermenters (LF)

  • Non-lactose Fermenters (NLF)

• Lactose Fermentation: Confirmed the current specimen is a lactose fermenter.

  • Next Test: Likely to perform the indole test.

Slide 9: Testing for Gram-positive Bacteria

• Current Observation: Growth on CNA but not on McConkey indicates Gram-positive.

• First Test for Gram-positive: Catalase test.

  • Results:

  • If catalase negative, might be Streptococcus.

  • If catalase positive, proceed to differentiate Staphylococcus vs. Micrococcus.

• Differentiation Tests:

  • Use oxidase test:

  • If oxidase positive, identify as Micrococcus.

  • If negative, identify as Staphylococcus.

• Staphylococcus Testing: Perform Coagulase test:

  • If positive, identify as Staphylococcus aureus.

  • If negative, may suggest other Staphylococcus species.

• Streptococcus Testing: For hemolysis; check type:

  • Hemolysis Types:

  • Beta hemolysis

    • First test is the Bacitracin susceptibility (trace susceptibility).

Slide 10: Gram-negative Further Investigation

• Observation: Growth on McConkey, no growth on CNA, confirmed Gram-negative.

• Determine Fermentation Status: Identify as a non-lactose fermenter (NLF).

• If oxidase positive, consider it a non-lactose fermenter not in Enterobacteriaceae.

• Swarming Growth: Uniquely observed in Proteus spp.

  • Next Steps: Check for indole:

  • If positive, identified as Proteus vulgaris.

  • If negative, identified as Proteus mirabilis.

• Swarming Descriptor: Recognizes growth patterns assist in identification.

Slide 11: Confirmations with Gram Positive Bacteria

• Observation Reiteration: Confirm Gram-positive growth on CNA.

• Catalase Test Results:

  • If positive, consider Micrococcus or Staphylococcus.

  • If negative, likely Streptococcus.

• Hemolysis Type Inquiry: Identify type as Alpha hemolytic.

• Alpha Hemolytic Testing:

  • Use Optochin disc test:

  • If susceptible, identify as Streptococcus pneumoniae.

  • If resistant, consider other options.

  • Fastidious organisms often grown on chocolate agar, requiring separate testing.

Overall Approach to Identification

• Initial Assessment: Conduct Gram stain to determine Gram reaction.

• Testing Protocols:

  • Gram-positive: Start with catalase.

  • Gram-negative: Start with oxidase.

• Recognize hemolytic behavior is primarily associated with Gram-positive organisms.

• Noteworthy Exception: Only use oxidase in Gram-positive to differentiate between Staphylococcus and Micrococcus.

Practical Considerations and Tips

• Examine growth samples after a 24-hour incubation period for proper identification.

• Practice memorizing flowchart approaches and results for accurate diagnosis.

• Understand the importance of specimen type for accurate identification:

  • Staphylococcus saprophyticus is notably a concern in young females with UTIs (often referred to as "honeymoon cystitis").

  • Other age demographics may not necessitate concern about saprophyticus.

• Contamination Considerations: High encounters with Staphylococcus epidermidis may indicate poor specimen collection techniques.

  • Focus on educational improvements if trends indicate contamination.

• Interpretation of Results: The importance of the clinical context should guide identification and testing protocols.