NOVOBIOCIN TEST

Effective Date: 
Fri, 08/09/2013
Reviewed: 
Tue, 01/10/2017
Policy: 

 

Purpose and Scope:

The HardyDisk™ Novobiocin Differentiation Disks are useful in presumptively distinguishing S. saprophyticus from other CoNS. Other human staphylococcal species that are novobiocin-resistant (S. cohnii, S. xylosus, S. pulvereri) are rarely isolated from patients.

Coagulase-negative staphylococci (CoNS) are among those organisms that have traditionally been considered skin contaminants, and their recovery from cultures doesn't always indicate presence of disease. Therefore, little attention had been paid to the pathogenic potential of this group of bacteria until recently. By the mid-1970's, microbiologists were becoming aware that CoNS could indeed be pathogenic, especially in compromised hosts.

 Today, S. saprophyticus has proven to be an important uropathogen. It is second only to E. coli as the most common cause of cystitis and acute urinary tract infection (UTI) in healthy, young adult women. S. saprophyticus tends to adhere to uroepithelial cells more often and more successfully than other staphylococcal species, this is believed to partially explain the organism's frequent role in urinary tract infections.

 

Reagents and Supplies:

  • HardyDisk™ Novobiocin Differentiation Disks 5ug
  • Mueller Hinton plates
  • BD  BBL Prompt Inoculation System

 

Reagent Storage

  • Upon receipt store at -20 to +8 degrees C. away from direct light.
  • The disks should not be used if there are any signs of deterioration, discoloration, or if the expiration date has passed.
  • Protect from light, excessive heat, and moisture.
  • The expiration date applies to the product in its intact packaging when stored as directed.

 

Quality Control:

 

QC is performed on each day the test is performed. Use procedure with stock culture of Staph saprophyticus ATCC 15305  (Novobiocin resistant) and Staph epidermidis ATCC 12228  (Novobiocin sensitive).

Record QC results on Novobiocin Test QC Log.

 

Procedure

  1. Allow disks to equilibrate to room temperature.
  2. Using a pure 18-24 hour culture, prepare a suspension using the Prompt inoculation system. (See Kirby Bauer Sensitivities Procedure SOP Micro-06 for instructions on using Prompt Inoculation system.)
  3. Inoculate Mueller Hinton Agar  plate with a sterile swab to obtain confluent growth. (See Kirby Bauer Sensitivities Procedure SOP Micro-06 for instructions on inoculating Mueller Hinton agar.)
  4. Aseptically apply one novobiocin disk onto the inoculated agar surface and lightly press down to ensure full contact with the medium.
  5. Incubate aerobically for 18-24 hours at 35-37 degrees C.
  6. Measure (in millimeters) the diameter of the zone of inhibition around the novobiocin disk, and record as susceptible or resistant.

Sensitive - A zone of inhibition greater than 16mm.
Resistant - A zone of inhibition less than or equal to 16mm.

 

Limitations

It is recommended that biochemical and/or serological tests be performed on colonies from pure culture for complete identification.

The novobiocin disk is not helpful and can give misleading results if it is performed on isolates other that those from urinary specimens.

Occasional human isolates that are not S. saprophyticus, S. cohnii subsp., or S. xylosis may also be resistant to novobiocin.

 

 

 

 

 

 

REFERENCES

HardyDisk™ Novobiocin Differentiation Disks Package Insert Copyright 2013

Murray, P.R., et al. 2003. Manual of Clinical Microbiology, 8th ed. American Society for Microbiology, Washington, D.C.