Vittaforma corneae (Shadduck et al.) Silveira and Canning (ATCC® 50505)

Depositor: ES Didier  /  Biosafety Level: 2

Permits and Restrictions

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Deposited As Nosema corneum Shadduck et al.
Biosafety Level 2
Isolation Corneal biopsy material from HIV-seronegative adult human male with keratitis, 1989
Product Format frozen
Storage Conditions Frozen Cultures:
-70°C for 1 week; liquid N2 vapor for long term storage

Freeze-dried Cultures:
2-8°C

Live Cultures:
See Protocols section for handling information
Type Strain no
Comments
Molecular comparison with other microspordian species
Molecular comparison of isolates from different animals
Discrimination between different species
Growth Conditions
Temperature: 37°C
Atmosphere: 5% CO2
Cell Line: ATCC® CRL-1634™
Alternate Cell Line: ATCC® CCL-34™ (kidney, canine) or CCL-26™ (kidney, monkey)
Cryopreservation Harvest and Preservation
  1. To harvest the culture, detach any remaining tissue culture cells (infected and uninfected) by scraping the surface of the flask with a cell scraper.
  2. Transfer the cell suspension (including parasites) to 15 mL plastic centrifuge tubes. Centrifuge at 1300 x g for 10 min.
  3. Remove all but 0.5 mL of the supernatant from each tube, resuspend the cell pellets, and pool them to a single tube.
  4. Pass the resulting cell suspension through a syringe equipped with a 27 gauge 1/2 in needle to break up any remaining cells. Adjust the parasite concentration to 2.0 - 4.0 x 107 cells/mL with fresh medium or PBS.
    NOTE: If the concentration of parasites is too low, centrifuge at 1300 x g for 10 min and resuspend in the volume of fresh medium or PBS required to yield the desired concentration.
  5. Prepare a cryoprotective solution containing 20% (v/v) DMSO and 20% (v/v) HIFBS in fresh medium or PBS.
  6. Mix the cell preparation and cryoprotective solution in equal portions. The final concentration will be 1.0 - 2.0 x 107 cells/mL, 10% DMSO, and 10% HIFBS. The time from the mixing of the cell preparation and cryoprotective solution to the start of the freezing process should be no less than 15 min. and no more than 60 min.
    NOTE: To prevent culture contamination, penicillin-streptomycin solution (ATCC® 30-2300) may be added to a final concentration of 50 to 100 I.U./mL penicillin and 50 to 100 µg/mL streptomycin.
  7. Dispense in 0.5 mL aliquots to 1.0-2.0 mL sterile plastic screw-capped cryules (special plastic vials for cryopreservation).
  8. Place vials in a controlled-rate freezing unit. From room temperature, cool at -1°C/min to -40°C. If freezing unit can compensate for the heat of fusion, maintain rate at -1 C/min through heat of fusion. At -40°C plunge ampules into liquid nitrogen. Alternatively, place the vials in a Nalgene 1°C freezing apparatus. Place the apparatus at -80°C for 1.5 to 2 hours and then plunge ampules into liquid nitrogen. (The cooling rate in this apparatus is approximately -1°C/min.)
  9. Store frozen ampules in either the vapor or liquid phase of a nitrogen refrigerator.
  10. To thaw a frozen ampule, place it in a 35°C water bath such that the lip of the ampule remains above the water line. Thawing time is approximately 2 to 3 minutes.  Do not agitate the ampule.  Do not leave ampule in water bath after thawed.
  11. Immediately after thawing, aseptically transfer contents to a T-25 tissue culture flask containing a fresh monolayer of cells (ATCC® CRL-1634™) and 10 mL ATCC 30-2003 with 3% (v/v) HIFBS.
  12. Outgas the flask for 10 seconds with a 95% air, 5% CO2 gas mixture.
  13. Incubate in a 37°C CO2 incubator with the cap screwed on tightly.
Name of Depositor ES Didier
Special Collection NCRR Contract
Chain of Custody
ATCC <-- ES Didier <-- J.A. Shadduck
Year of Origin 1989
References

Didier ES, et al. Identification and characterization of three Encephalitozoon cuniculi strains. Parasitology 111: 411-421, 1995. PubMed: 11023405

Didier ES, et al. A microsporidian isolated from an AIDS patient corresponds to Encephalitozoon cuniculi III, originally isolated from domestic dogs. J. Clin. Microbiol. 34: 2835-2837, 1996. PubMed: 8897194

Didier ES, et al. Diagnosis of disseminated microsporidian Encephalitozoon hellum infection by PCR - southern analysis and successful treatment with albendazole and fumagillin. J. Clin. Microbiol. 34: 947-952, 1996. PubMed: 8815114

Didier ES, et al. Characterization of Encephalitozoon (Septata) intestinailis isolates cultured from nasal mucosa and bronchoalveolar lavage fluids of two AIDS patients. J. Eukaryot. Microbiol. 43: 34-43, 1996. PubMed: 8563708

Shadduck JA, et al. Isolation of a microsporidian from a human patient. J. Infect. Dis. 162: 773-776, 1990. PubMed: 2117629

Mittleider D, et al. Sequence survey of the genome of the opportunistic microsporidian pathogen, Vittaforma corneae. J. Eukaryot. Microbiol. 49: 393-401, 2002. PubMed: 12425527

Molestina R, Becnel JJ, Weiss LM. Culture and Propagation of Microsporidia. In Microsporidia: Pathogens of Opportunity, First Edition, Chapter 18: pp. 457-467, 2014. Hoboken, NJ: John Wiley & Sons, Inc.

Notice: Necessary PermitsPermits

These permits may be required for shipping this product:

  • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
Basic Documentation