Acanthamoeba griffini Sawyer (ATCC® 30731)

Strain Designations: S-7  /  Depositor: TK Sawyer  /  Biosafety Level: 1

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Strain Designations S-7
Application
Molecular characterization of corneal pathogen
Biosafety Level 1
Isolation
shallow-water, beach-bottom sample, New London, CT, 1962
Product Format frozen
Type Strain no
Comments
species description
Group I introns
Subgenus systematics based upon SSU rDNA sequence data
Molecular characterization of corneal pathogen
phylogeny
Medium Medium 997: Fresh water ameba medium
Growth Conditions
Temperature: 25.0°C
Duration: grown with bacteria
Protocol: ATCCNO: 30011 SPEC: This strain is distributed as a dried preparation. See the general procedures for opening a dried vial. Aseptically add 1 ml of sterile distilled water to the inner shell vial, remove the filter paper aseptically with a pair of forceps, and place it in the center of an agar plate of ATCC medium 997. Add the liquid remaining in the vial to the plate and spread it evenly over the surface of the plate. Incubate the plate at 25C. Trophozoites (amebae) should be evident within 2-3 days.
Subcultivation
Protocol: ATCCNO: 30011 SPEC: This strain is distributed as a dried preparation. See the general procedures for opening a dried vial. Aseptically add 1 ml of sterile distilled water to the inner shell vial, remove the filter paper aseptically with a pair of forceps, and place it in the center of an agar plate of ATCC medium 997. Add the liquid remaining in the vial to the plate and spread it evenly over the surface of the plate. Incubate the plate at 25C. Trophozoites (amebae) should be evident within 2-3 days.
Cryopreservation
Cryoprotective Solution

DMSO                                                                                    1.5 ml

Dryl's solution (or similar)                                                  8.5 ml

1.     Mix the components in the order listed. When the medium is added to the DMSO the solution will warm up due to chemical heat.

2.    Harvest cells from a culture which is at or near peak density by adding 5 ml ATCC medium 5080 (Dryl's solution) and washing cells into suspension.  Rub the surface of the plate with a spread bar to detach adhering trophozoites.

3.     Adjust the concentration of cells to at least 2 x 106/ml in fresh medium.

4.     Mix the cell preparation and the cryoprotective solution in equal portions.

5.     Dispense in 0.5 ml aliquots into 1.0 - 2.0 ml sterile plastic screw-capped cryules (special plastic vials for cryopreservation).

6.     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.)  

7.     Ampules are stored in either the vapor or liquid phase of a nitrogen refrigerator.

8.     To establish a culture from the frozen state place the vial in a 35°C water bath.  Immerse the vial to a level just above the surface of the frozen material. Do not agitate the vial.   Immediately after thawing, aseptically transfer the contents of the ampule to the center of a fresh plate of ATCC medium 997.  Distribute the material evenly over the plate using a spread bar.

9.     Wrap the entire edge of the plate with parafilm and incubate upright at 25°C.

Follow the protocol for maintenance of culture.

Name of Depositor TK Sawyer
Chain of Custody
ATCC <<--TK Sawyer<<--J. Griffin
Year of Origin 1962
References

Gast RJ, et al. Discovery of group I introns in the nuclear small subunit ribosomal RNA genes of Acanthamoeba. Nucleic Acids Res. 22: 592-596, 1994. PubMed: 8127708

Sawyer TK. Acanthamoeba griffini, a new species of marine amoeba. J. Protozool. 18: 650-654, 1971.

Nerad TA, et al. Acanthamoeba pearcei n. sp. (Protozoa: Amoebida) from sewage contaminated sediments. J. Eukaryot. Microbiol. 42: 702-705, 1995. PubMed: 8520585

Gast RJ, et al. Subgenus systematics of Acanthamoeba: Four nuclear 18S rDNA sequence types. J. Eukaryot. Microbiol. 43: 498-504, 1996. PubMed: 8976608

Ledee DR, et al. Acanthamoeba griffini, molecular characterization of a new corneal pathogen. Invest. Ophthalmol. Vis. Sci. 37: 544-550, 1996. PubMed: 8595954

Daggett PM, et al. Distribution and possible interrelationships of pathogenic and nonpathogenic Acanthamoeba from aquatic environments. Microb. Ecol. 8: 371-386, 1982.

Daggett PM, et al. A molecular approach to the phylogeny of Acanthamoeba. Biosystems 18: 399-405, 1985. PubMed: 4084681

Stothard DR, et al. Fluorescent oligonucleotide probes for clinical and environmental detection of Acanthamoeba and the T4 18S rRNA gene sequence type. J. Clin. Microbiol. 37: 2687-2693, 1999. PubMed: 10405422

Khan NA, et al. Proteases as markers for differentiation of pathogenic and nonpathogenic species of Acanthamoeba. J. Clin. Microbiol. 38: 2858-2861, 2000. PubMed: 10921939

Schroeder JM, et al. Use of subgenic 18s ribosomal dna pcr and sequencing for genus and genotype identification of acanthamoebae from humans with keratitis and from sewage sludge. J. Clin. Microbiol. 39: 1903-1911, 2001. PubMed: 11326011

Kong HH, et al. Mitochondrial DNA restriction fragment length polymorphism (RFLP) and 18S small-subunit ribosomal DNA PCR-RFLP analyses of Acanthamoeba isolated from contact lens storage cases of residents in southwestern Korea. J. Clin. Microbiol. 40: 1199-1206, 2002. PubMed: 11923331

Ledee DR, et al. Advantages of using mitochondrial 16S rDNA sequences to classify clinical isolates of Acanthamoeba. Invest. Ophthalmol. Vis. Sci. 44: 1142-1149, 2003. PubMed: 12601042

Marciano-Cabral F, Cabral G. Acanthamoeba spp. as agents of disease in humans. Clin. Microbiol. Rev. 16: 273-307, 2003. PubMed: 12692099

Cross References

Nucleotide (GenBank) : U07412 nuclear SSU rRNA gene

Nucleotide (GenBank) : AF019060 Acanthamoeba hatchetti 2AX1 18S ribosomal RNA gene, partial sequence.

Nucleotide (GenBank) : U02540 Acanthamoeba griffini S-7 ATCC 30731 nuclear small subunit ribosomal gene, group I intron.

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.
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