Clinical Microbiology In the Department of Pathology

All serotypes of Shiga toxin-producing E. coli are detected in diarrheal stool by a sensitive multiplex, real-time PCR after enrichment in GN broth.

Molecular Microbiology

For over 100 years, agents of infectious diseases have been identified through their phenotype directly in specimen and after growth in culture.  In the molecular era, there is an opportunity to detect pathogens more rapidly and accurately based on their genetic signatures.  The Stanford Hospital Clinical Microbiology Laboratory offers a growing number of nucleic acid amplification tests (NAATs) for detection and identification of bacterial and fungal pathogens.  These assays improve patient care, reduced antibiotic usage, enhance test utilization, and increase laboratory and hospital efficiency.

Direct bacterial and fungal detection and identification by PCR/gene sequencing:
PCR amplicons of ribosomal RNA genes and other targets are sequenced to detect and identify invasive bacterial and fungal pathogens that are refractory to diagnose by conventional methods.  Sterile specimens such as tissue and body fluids are required for this assay.

Direct Mycobacterium tuberculosis detection by real-time PCR:
Real-time PCR for detection of M. tuberculosis complex targeting the IS6110 and 16S rRNA gene is automatically performed on smear-positive respiratory specimens.  Smear-negative specimens and non-respiratory samples are tested by request.  Genotypic susceptibility testing for rifampin and isoniazid as well as fluoroquinolone and injectable drugs are performed by request at California Department of Public Health, Microbial Diseases Laboratory.

Direct Bordetella pertussis and B. parapertussis detection by real-time PCR:
This test, a real-time PCR assay that detects a unique insertion sequences, is more sensitive than culture for detection of B. pertussis or B. parapertussis in respiratory secretions.  A special collection kit, available from the laboratory, is recommended.

Direct Clostridium difficile detection by real-time PCR:
The clinical laboratory performs a sensitive real-time PCR for rapid and sensitive detection of Clostridium difficile in loose stool specimens.  This test has a turn around time of less than 2 hours allowing rapid diagnosis and isolation of patients with C. difficile infection.

Detection of stool pathogens by real-time PCR:
Shiga toxin-producing E. coli (STEC) are detected by a multiplexed, real-time PCR assay targeting the genes encoding Shiga Toxin-1 and -2.  This assay detects all serotypes of STEC including O157:H7.  Salmonella enterica and Shigella species are detected by a multiplexed, real-time PCR assay.  Both assays are performed after enrichment of stool in selective broth.

Strep Group B screening by real-time PCR:
Streptococcus agalactiae is detected from vaginal/anal swabs by a highly sensitive multiplexed, real-time PCR assay after enrichment in selective broth.

Strep Group A and Group C/G screening by real-time PCR:
Streptococcus pyogenes and Strep group C/G are detected from throat swabs by a highly sensitive multiplexed, real-time PCR assay after enrichment in selective broth.

Identification of cystic fibrosis pathogens by real-time PCR:
A multiplexed, real-time PCR assay is utilized to rapidly and accurately identify Burkholderia cepacia complex, Acinetobacter baumannii, Achromobacter xylosoxidans, and Stenotrophomonas maltophilia and Pseudomonas aeruginosa from solid culture.  

MecA and BlaZ detection by real-time PCR:
All Staphylococcus aureus and clinically important coagulase-negative staphylococcus isolates are tested for the presence of the mecA gene by real-time PCR. This assay is performed on the same day culture is detected. S. lugdunensis isolates are identified by real-time PCR.  Staphylococcus isolates susceptible to penicillin by MIC testing are further subjected to BlaZ PCR.

Mycobacterial culture identification by multiplex, real-time PCR:
Mycobacterial isolates are rapidly identified with a multiplexed, real-time PCR.  Ribosomal RNA gene sequencing is also used as needed.

Bacterial and fungal identification by PCR/gene sequencing:
Isolates not identifiable by conventional microbiology methods are identified by ribosomal RNA genes sequencing.

Pulsed field gel electrophoresis bacterial strain typing:
Bacteria are pelleted, DNA is treated with endonuclease enzymes, the strands are separated, and the DNA strands are subjected to varying waves of electrical current in a polyacrylamide gel. The resulting band patterns are used to define the relatedness among strains. Stanford has experience with numerous species, and we are especially knowledgeable about methicillin-resistant Staphylococcus aureus.


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