SODIUM POLYANETHOL SULFONATE (SPS) IDENTIFICATION DISKS

Cat. no. Z7381 HardyDisk™ SPS Identification Disks 50 disks/cartridge

INTENDED USE

HardyDisk™ SPS Identification Disks evaluate the ability of an organism to grow in the presence of Sodium Polyanethol Sulfonate. HardyDisk™ SPS Identification Disks can be used to presumptively identify microorganisms including but not limited to Peptostreptococcus anaerobius, Gardnerella vaginalis, and Haemophilus ducreyi.

SUMMARY

Peptostreptococcus belong to the genus of gram-positive anaerobic cocci. These organisms are widely distributed as normal flora in humans and animals and account for up to a third of anaerobic human isolates.(6) The three species of Peptostreptococcus most commonly reported in clinical specimens are P. anaerobius, P. magnus, and P. asaccharolyticus. Specific identification of most anaerobic cocci requires both biochemical tests and gas-liquid chromatography, as these organisms tend to exhibit few visual phenotypic differences.(2)

In contrast to traditional testing procedures, the Sodium Polyanethol Sulfonate (SPS) disk test is a simple and rapid technique used for the presumptive identification of P. anaerobius from other gram-positive anaerobic cocci. The characteristic inhibition by SPS to differentiate P. anaerobius from other gram-positive anaerobic cocci was initially demonstrated by Graves, et al.(1) The SPS disk test has since been demonstrated to have an overall accuracy of 98% in the identification of P. anaerobius(9) Consequently, it is recommended that the SPS disk test be routinely applied to all gram-positive anaerobic cocci isolates to presumptively identify P. anaerobius.(8)

SPS inhibition is also useful in the presumptive identification of microorganisms other than P. anaerobius. Reimer and Reller initially demonstrated that SPS disk susceptibility and alpha-hemolytic streptococci inhibition can be employed to accurately identify Gardnerella vaginalis.(10) Currently the differentiation of G. vaginalis from other catalase-negative, gram-variable, coccobacilli is achieved solely on the susceptibility of G. vaginalis to SPS.(4) An additional study performed by Shawar, et al. confirmed that SPS susceptibility can also be used to differentiate Haemophilus ducreyi from other Haemophilus species.(11)

FORMULA

Each HardyDisk™ SPS Identification Disk is prepared by impregnating 1mg of Sodium Polyanethol Sulfonate onto a 0.25 inch diameter filter paper disk.

STORAGE AND SHELF LIFE

Storage: Upon receipt store at 2-8ºC. away from direct light. The disks should not be used if there are any signs of deterioration or if the expiration date has passed. Product is light and temperature sensitive; protect from light, excessive heat, and moisture.

PRECAUTIONS

PROCEDURE

1. Allow disks to equilibrate to room temperature before use. Prepare a suspension, equivalent to a 0.5 McFarland turbidity standard, of the organism to be tested from a pure 24-48 hour culture.

2. Dip a sterile non-toxic swab into the organism suspension. Rotate the swab several times, pressing firmly on the inside wall of the tube above the fluid level. This will remove excess inoculum from the swab. Evenly inoculate the dried surface of a Brucella Agar with Vitamin K and Hemin plate (Cat. no. A30) to obtain confluent growth.

3. Place one SPS disk on the media surface. With sterile forceps, gently tap the disk to ensure complete contact with the plate surface.

4. Incubate the media at 35ºC. for 24-48 hours. Consult listed references for recommended incubation atmosphere.(2-5)

5. After incubation, examine the media surface and measure the zone diameter surrounding the SPS disk. Zones should be recorded to the nearest whole millimeter.

INTERPRETATION OF RESULTS

When the zone diameter is greater than or equal to 12mm, the organism is considered susceptible to SPS. When the zone diameter is less than or equal to 11mm, the organism is considered resistant to SPS.

LIMITATIONS

P. micros and P. prevotii produce small zones of inhibition, usually less than 10mm. Occasionally isolates can produce zones larger than 12mm, which will be interpreted as sensitive.(8)

H. ducreyi is susceptible with a zone greater than or equal to 12mm; all other Haemophilus species are resistant and will have zones less than 12mm. Capnocytophaga is the only other genus of gram-negative rods that are susceptible to SPS, but they are ALA-positive. The addition of 0.002% Tween® 80 may aid in the dispersion of the cells.(4,11)

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, other culture media, swabs, applicator sticks, incinerators, and incubators, etc., as well as serological and biochemical reagents, are not provided.

QUALITY CONTROL

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Peptostreptococcus anaerobius
ATCC ® 27337
F 24-48hr 35°C Anaerobic Zone of inhibition greater than or equal to 12mm
Gardnerella vaginalis
ATCC ® 14018
F 24-48hr 35°C CO 2 ** Zone of inhibition greater than or equal to 12mm
Bacteroides fragilis
ATCC ® 25285
F 24-48hr 35°C Anaerobic Zone of inhibition less than 12mm

User Quality Control

** Atmosphere of incubation is enriched with 5-10% CO2.

PHYSICAL APPEARANCE

HardyDisk™ SPS Identification Disks are 0.25 inch (in diameter) filter paper disks with the letters SPS on both sides, and should appear white in color.

SPS Resistant

Showing SPS Sensitivity
Gardnerella vaginalis (ATCC® 14018) growing with a zone around a HardyDisk™ SPS Identification Disks (Cat. no. Z7381), demonstrating sensitivity. Incubated in CO2 for 48 hours at 35ºC.

SPS Resistant

Showing SPS Resistance
Bacteroides fragilis (ATCC® 25285) growing around a HardyDisk™ SPS Identification Disks (Cat. no. Z7381), demonstrating resistance. Incubated anaerobically for 48 hours at 35ºC.

REFERENCES

1. Graves, M.H., et al. 1974. Sodium polyanethol sulfonate sensitivity of anaerobic cocci. Applied Microbiology.; 27: 1131-1133.

2. Jorgensen., et al. Manual of Clinical Microbiology, American Society for Microbiology, Washington, D.C.

3. Tille, P., et al. Bailey and Scott's Diagnostic Microbiology, C.V. Mosby Company, St. Louis, MO.

4. York, M.K., M.M. Traylor. 2002. Sodium polyanethol sulfonate disk test. Clinical Microbiology Procedures Handbook, Vol. I & II. American Society for Microbiology, Washington, D.C.

5. Koneman, E.W., et al. Color Atlas and Textbook of Diagnostic Microbiology, J.B. Lippincott Company, Philadelphia, PA.

6. Brook, I. 1988. Recovery of anaerobic bacteria from clinical specimens in 12 years at two military hospitals. J. Clin. Microbiol.; 26:1181-1188.

7. Schwarz, D. et al. 1926. Anaerobic streptococci: their role in puerperal infection. South Medical Journal.; 19:470-479.

8. Summanen, Paula, et al. 1993. Wadsworth Anaerobic Bacteriology Manual, 5th ed. Star Publishing, Belmont, CA.

9. Wideman, P.A, et al. 1976. Evaluation of the sodium polyanethol sulfonate disk test for the identification of Peptostreptococcus anaerobius. J. Clin. Microbiol.; 4:330-334.

10. Reimer, L.G. and L.B. Reller. 1985. Use of a sodium polyanethol sulfonate disk for the identification of Gardnerella vaginalis. J. Clin. Microbiol.; 21:146-149.

11. Shawar R., J. Sepulveda, and J.E. Clarridge. 1990. Use of the RapID-ANA system and sodium polyanethol sulfonate disk susceptibility testing in identifying Haemophilus ducreyi. J. Clin. Microbiol.; 28:108-111.

12. Catlin, B.W. 1992. Gardnerella vaginalis: characteristics, clinical considerations, and controversies. Clinical Microbiology Review.; 5: 213-237.


ATCC is a registered trademark of the American Type Culture Collection.
Tween is a registered trademark of ICI Americas, Inc.

081516gr