Chapter 17

Chapter 17: Procedures for Identifying and Diagnosing Infections

Specimen Collection

  • Definition: The process of sampling body sites or fluids to identify suspected infectious agents.

  • Key Points:

    • Results depend significantly on the methods used for specimen collection, handling, transport, and storage.

    • Aseptic procedures are imperative to avoid contamination.

General Guidelines for Specimen Collection

  • Refer to TABLE 17.1 for detailed methods and recommendations:

    • Abscess of skin or membrane or decubitus ulcer

    • Procedure: Debride the surface with a sterile wipe; suction the fluid using a sterile needle and syringe.

    • Note: Aspiration of tissue samples is preferred to using a swab; transfer to an anaerobic transport system as soon as possible.

    • Anaerobic cultures

    • Collection: Sample from deeper tissue below the surface using a sterile syringe.

    • Important: The sample must not be exposed to air and must be transported immediately in an anaerobic system to the lab.

    • Blood

    • Preparation: Clean the skin area with iodine; utilize Vacutainer or Bactec blood collection tubes.

    • Inoculation: Blood culture bottles should be inoculated immediately following collection.

    • Bone marrow

    • Procedure: Prepare the site for surgical incision; obtain samples using a special needle.

    • Typical sites: Sample is often taken from the sternum or ilium and placed in a blood bottle for culturing.

    • Cerebrospinal fluid (CSF)

    • Procedure: Remove the CSF aseptically via lumbar puncture.

    • Handling: Place CSF into sterile tubes; do not refrigerate; transport immediately to the lab.

    • Feces

    • Procedure: Collect a small specimen into a sterile container; cover and transport.

    • Note: Refrigerate samples held for over an hour. Cultures typically aim to rule out enteric pathogens; special kits are available to detect cysts, trophozoites, and larvae of intestinal worms.

    • Urine

    • Two collection methods:

      1. Clean midstream catch:

        • Clean external genitalia, separate labia (for females) or retract the foreskin (for males) and urinate briefly, then collect 100-200 cc sample midstream.

        • Important: Samples not processed promptly must be refrigerated.

      2. Sterile catheter method:

        • Insert a sterile catheter into the bladder, draining urine into a sterile container.

        • Clean and rinse the urethral opening; use only a specimen catheter for sampling bladder infections; indwelling catheters typically contain microbes.

Clinical Microbiology

  • Identification Methods for Unknown Microbes:

    • Phenotypic Methods:

    • Microscopic morphology: Fresh or stained microorganisms from specimens; assess shape, size, stain reaction, and cell structures.

    • Macroscopic morphology: Evaluates colony appearance including texture, size, shape, pigment, and growth requirements.

    • Physiological/biochemical characteristics: Detection of specific enzymes or metabolic pathways.

Biochemical Phenotypic Methods

  • MALDI-TOF:

    • Definition: A method of automated identification utilizing the unique combination of proteins and nucleic acids in microorganisms.

Genotypic Methods

  • Definition: Techniques that assess the genetic make-up of microorganisms.

  • Advantages:

    • No culture necessary.

    • Methods are precise, automated, and yield quick results.

    • Polymerase Chain Reaction (PCR):

    • Valuable for DNA and RNA (RT-PCR) analysis.

    • High sensitivity and specificity are critical for detecting pathogens, including HIV, Lyme disease, human papillomavirus, tuberculosis, and hepatitis.

Immunologic Methods

  • Definition: Analysis of blood (serology) and other fluids, focusing on antibody-antigen reactions.

  • Serology:

    • In vitro diagnostic testing of serum.

    • Visual reactions from antibody-antigen interactions include:

    • Precipitates.

    • Color changes.

    • Release of radioactivity.

    • High specificity and sensitivity for detecting or identifying unknown antibodies using known antigens, or vice versa.

Serological Testing

  • Clumping: Indicates a positive test result.

  • No Clumping: Indicates a negative test result.

Point-of-Care and Rapid Diagnostic Tests

  • Advantages:

    • Minimal training required for administration and evaluation.

    • No need for access to hospitals or laboratories.

    • Currently available Rapid Diagnostic Tests (RDTs) include those for pathogens such as:

    • Streptococcus.

    • Staphylococcus.

    • HIV.

    • Hepatitis B and C.

    • SARS-CoV-2.

Clinical Case Example

  • Patient Profile:

    • 73-year-old patient presenting with knee pain.

  • Procedure:

    • Joint fluid was drained and cultured.

    • Initially identified the population of bacteria as Francisella tularensis (a Tier 1 Selective Agent).

  • Outcome:

    • 24 individuals had been exposed to this pathogen and began antibiotic treatment.

    • Subsequent rRNA sequencing re-identified the pathogen as Veillonella parvula.