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P2.051 A Cost-Effective and Simple Alternative Technique For Resuscitation of Freeze-Dried Cultures of Neisseria Gonorrhoeae
  1. M Bala,
  2. V Singh,
  3. M Kakran,
  4. V Ramesh
  1. VMMC & Safdarjung Hospital, New Delhi, India

Abstract

Background Freeze drying (lyophilization) of bacteria is a very well established method for the archiving and long-term storage. The recommended medium for resuscitation of freeze-dried cultures of Neisseria gonorrhoeae is 1 ml of a nutrient blood broth or a rich peptone broth supplemented with 10% blood or nutrient broth. Sheep blood or horse blood is not easily available in most of the labs. and human blood is not recommended. Normal saline (0.9% W/V Sodium chloride) is mostly available in all the labs. This study compared the nutrient blood broth and normal saline for resuscitation of freeze-dried cultures of reference and clinical strains of N. gonorrhoeae and evaluated their performance characteristics for the growth of N. gonorrhoeae strains.

Methods A prospective study was undertaken between January 2011 and December 2012. Ninety three N. gonorrhoeae lyophilized strains including 83 clinical isolates, ATCC 49226 and nine WHO reference strains were tested using both the methods. Reconstituted material from both the techniques was subcultured on to chocolate agar and was incubated for 24–48 hrs at 36°C in a moist atmosphere containing 5 to 10% carbon dioxide. The results were recorded in terms of viable gonococci, colony morphology and colony size.

Results N. gonorrhoeae was successfully isolated from 89 (95.7%) lyophilized strains by both the techniques used for revival. In other 4 cases, it was difficult to state with absolute certainty that the gonococci were nonviable or that unsatisfactory lyophilization practises were responsible for these results as the quality of the lyophilsed strains also determined the growth on revival.

Conclusion Preliminary data suggest that nutrient blood broth and normal saline were equal in performance for revival of lyophilized strains and normal saline could be recommended as convenient and inexpensive alternative for universal use.

  • Freeze-dried cultures
  • Neisseria gonorrhoeae
  • Resuscitation techniques

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