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Diagnostic Microbiology Laboratory (DML)

The Diagnostic Microbiology Laboratory (DML) routinely performs a variety of procedures to isolate and identify important pathogenic bacteria and fungi from wildlife.  Microbes are identified based upon morphological characteristics, biochemical/physiological properties, molecular assays (e.g., PCR), and DNA sequence analysis.

DML Capabilities

Biochemical tests are one method used to identify bacteria at the NWHC. (Public domain.)

Diagnostic Bacteriology:

  • Routine culture of aerobic and anaerobic bacteria using a variety of standard culture media
  • Selective culture for slow-growing or fastidious bacteria such as Mycoplasma, Francisella tularensis (tularemia), etc.
  • Identification of bacterial isolates through the use of physiological tests and DNA sequencing
  • Identification of unculturable or difficult-to-culture bacteria (e.g., Mycobacterium) in tissue samples by pan-bacterial PCR and DNA sequencing of the 16S rRNA gene
  • Detection of Clostridium piliforme (Tyzzer’s disease) by PCR
  • Samples suspect for certain organisms such as Chlamydia/Chlamidophila (chlamydiosis), Leptospira (leptospirosis), and Francisella tularensis (tularemia), are submitted to a specialty laboratory

 

Conventional PCR and DNA sequencing
Conventional PCR and DNA sequencing are used to detect unculturable microorganisms in diagnostic samples. (Public domain.)

 

Diagnostic Mycology:

  • Routine culture and identification of filamentous fungi and yeasts using a variety of culture media
  • Selective culture for slow-growing and difficult-to-isolate fungi such as dermatophytes, Batrachochytrium (chytridiomycosis), Ophidiomyces ophiodiicola (snake fungal disease) and other reptile-infecting fungi, Pseudogymnoascus destructans (white-nose syndrome), etc.
  • Identification of fungal isolates based on morphology and DNA sequencing
  • Identification of unculturable or difficult-to-culture fungi in tissue samples by pan-fungal PCR and DNA sequencing of the internal transcribed spacer (ITS) region of the rRNA gene complex
  • Detection of Batrachochytrium dendrobatidis and Batrachochytrium salamandrivorans (chytridiomycosis) by real-time PCR
  • Detection of Ophidiomyces ophiodiicola (snake fungal disease) by real-time PCR
  • Detection of Pseudogymnoascus destructans (white-nose syndrome) by real-time PCR

 

 

Microbial Toxins:

Routine culture
Routine microbial culture using a variety of culture media. (Public domain.)
  • Identification of Clostridium botulinum neurotoxin types C and E using the mouse protection assay

Additional Characterizations of Isolates:

  • Strain characterization of bacteria and fungi for epidemiological investigations by multilocus sequence typing
  • Identification of Pasteurella multocida non-capsular serotypes
  • Identification of Salmonella serotypes is performed by a reference laboratory
  • Characterization of novel bacterial and fungal pathogens

Other Capabilities:

  • Detection of circoviruses by PCR (see Diagnostic Virology Laboratory (DVL) for additional viral diagnostic tests)
  • Confirmation of host-species identity through sequencing of DNA barcoding regions
  • Development and optimization of molecular assays for detection of bacterial and fungal pathogens

DML Capabilities

Biochemical tests are one method used to identify bacteria at the NWHC. (Public domain.)

Diagnostic Bacteriology:

  • Routine culture of aerobic and anaerobic bacteria using a variety of standard culture media
  • Selective culture for slow-growing or fastidious bacteria such as Mycoplasma, Francisella tularensis (tularemia), etc.
  • Identification of bacterial isolates through the use of physiological tests and DNA sequencing
  • Identification of unculturable or difficult-to-culture bacteria (e.g., Mycobacterium) in tissue samples by pan-bacterial PCR and DNA sequencing of the 16S rRNA gene
  • Detection of Clostridium piliforme (Tyzzer’s disease) by PCR
  • Samples suspect for certain organisms such as Chlamydia/Chlamidophila (chlamydiosis), Leptospira (leptospirosis), and Francisella tularensis (tularemia), are submitted to a specialty laboratory

 

Conventional PCR and DNA sequencing
Conventional PCR and DNA sequencing are used to detect unculturable microorganisms in diagnostic samples. (Public domain.)

 

Diagnostic Mycology:

  • Routine culture and identification of filamentous fungi and yeasts using a variety of culture media
  • Selective culture for slow-growing and difficult-to-isolate fungi such as dermatophytes, Batrachochytrium (chytridiomycosis), Ophidiomyces ophiodiicola (snake fungal disease) and other reptile-infecting fungi, Pseudogymnoascus destructans (white-nose syndrome), etc.
  • Identification of fungal isolates based on morphology and DNA sequencing
  • Identification of unculturable or difficult-to-culture fungi in tissue samples by pan-fungal PCR and DNA sequencing of the internal transcribed spacer (ITS) region of the rRNA gene complex
  • Detection of Batrachochytrium dendrobatidis and Batrachochytrium salamandrivorans (chytridiomycosis) by real-time PCR
  • Detection of Ophidiomyces ophiodiicola (snake fungal disease) by real-time PCR
  • Detection of Pseudogymnoascus destructans (white-nose syndrome) by real-time PCR

 

 

Microbial Toxins:

Routine culture
Routine microbial culture using a variety of culture media. (Public domain.)
  • Identification of Clostridium botulinum neurotoxin types C and E using the mouse protection assay

Additional Characterizations of Isolates:

  • Strain characterization of bacteria and fungi for epidemiological investigations by multilocus sequence typing
  • Identification of Pasteurella multocida non-capsular serotypes
  • Identification of Salmonella serotypes is performed by a reference laboratory
  • Characterization of novel bacterial and fungal pathogens

Other Capabilities:

  • Detection of circoviruses by PCR (see Diagnostic Virology Laboratory (DVL) for additional viral diagnostic tests)
  • Confirmation of host-species identity through sequencing of DNA barcoding regions
  • Development and optimization of molecular assays for detection of bacterial and fungal pathogens
Real-time PCR
Real-time PCR is used to confirm the presence of certain pathogenic microorganisms in samples.(Public domain.)