Giardia intestinalis (Lambl) Alexeieff (ATCC® 50803)

Organism: Giardia intestinalis (Lambl) Alexeieff  /  Depositor: FD Gillin

Strain Designations WB clone C6
Application
Enteric Research
Food and waterborne pathogen research
Biosafety Level 2
Isolation
clone of strain WB (ATCC 30957), 1983
Product Format frozen
Type Strain no
Genome Sequenced Strain

Yes

Comments
Excellent cyst-formation under axenic conditions
Variable surface antigen
differential mRNA processing
Genome sequencing strain
Medium Medium 2155: LYI Giardia Medium (filtered)
Growth Conditions
Temperature: 35.0°C
Duration: axenic; anaerobic
Cryopreservation

1.   Harvest cells from a culture that is at or near peak density. To detach cells from the wall of the culture tubes place on ice for 10 minutes.  Invert tubes several times until the majority of the cells are in suspension.  Centrifuge tubes at 800 x g for 5 minutes. 

2.   Adjust the concentration of cells to 2 x 107/ml in fresh medium.

3.  Before centrifuging prepare a 24% (v/v) solution of sterile DMSO in fresh medium containing 8% (w/v) sucrose.  The solution is prepared as follows:

a) Add 10.5 g sucrose to 10 ml of fresh medium and filter sterilize through a 0.2 mm filter;

b) Add 2.4 ml of DMSO to an ice cold 20 x 150 mm screw-capped test tube;

c) Place the tube on ice and allow the DMSO to solidify (~5 min) and then add 7.6 ml of ice cold medium prepared in step 3a.  The final concentration will be 24% (v/v) DMSO and 8% (w/v) sucrose;

d) Invert several times to dissolve the DMSO;

e) Allow to warm to room temperature.

4.   Mix the cell preparation and the cryoprotective agent, prepared in step 3, in equal portions. Thus, the final concentration will equal 12% (v/v) DMSO + 4% sucrose (w/v) and 107 cells/ml. The time from the mixing of the cell preparation and DMSO stock solution before the freezing process is begun should be no less than 15 min and no longer than 30 min.

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 the vials in a controlled rate freezing unit.  From room temperature cool at -1°C/min to -40°C.  If the freezing unit can compensate for the heat of fusion, maintain rate at        -1°C/min through the heat of fusion.  At -40°C plunge 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.  The frozen preparations should be stored in either the vapor or liquid phase of a nitrogen refrigerator. Frozen preparations stored below -130°C are stabile indefinitely. Those stored at temperatures above -130°C are progressively less stabile as the storage temperature is elevated.

8.   To establish a culture from the frozen state place an ampule in a water bath set at 35°C. Immerse the vial just to a level just above the surface of the frozen material. Do not agitate the vial.

9.   Immediately after thawing, do not leave in the water bath, aseptically remove the contents of the ampule and inoculate a 16 x 125 mm screw-capped test tube containing 13 ml ATCC Medium 2695.

10.          Incubate the culture on a 15° horizontal slant at 35°C.

Name of Depositor FD Gillin
Special Collection NCRR Contract
Year of Origin 1983
References

Aley SB, et al. Killing of Giardia lamblia by cryptdins and cationic neutrophil peptides. Infect. Immun. 62: 5397-5403, 1994. PubMed: 7960119

Reiner DS, et al. A lipoprotein-cholesterol-albumin serum substitute stimulates Giardia lamblia encystation vesicle formation. J. Eukaryot. Microbiol. 42: 622-627, 1995. PubMed: 7581338

Aley SB, Gillin FD. Giardia lamblia: post-translational processing and status of exposed cysteine residues in TSA 417, a variable surface antigen. Exp. Parasitol. 77: 295-305, 1993. PubMed: 8224085

Reiner DS, et al. Giardia lamblia: absence of cyst antigens and reduced secretory vesicle formation and bile salt uptake in an encystation-deficient subline. Exp. Parasitol. 77: 461-472, 1993. PubMed: 7504633

Das S, Gillin FD. Giardia lamblia: increased UDP-N-acetyl-D-glucosamine and N-acetyl-D-galactosamine transferase activities during encystation. Exp. Parasitol. 83: 19-29, 1996. PubMed: 8654548

Knodler LA, et al. Novel protein-disulfide isomerases from the early-diverging protist Giardia lamblia. J. Biol. Chem. 274: 29805-29811, 1999. PubMed: 10514458

Eckmann L, et al. Nitric oxide production by human intestinal epithelial cells and competition for arginine as potential determinants of host defense against the lumen-dwelling pathogen Giardia lamblia. J. Immunol. 164: 1478-1487, 2000. PubMed: 10640765

Que X, et al. Developmentally regulated transcripts and evidence of differential mRNA processing in Giardia lamblia. Mol. Biochem. Parasitol. 81: 101-110, 1996. PubMed: 8892309

Abel ES, et al. Possible roles of protein kinase A in cell motility and excystation of the early diverging eukaryote Giardia lamblia. J. Biol. Chem. 276: 10320-10329, 2001. PubMed: 11104758

Marti M, et al. Conformationally correct expression of membrane-anchored Toxoplasma gondii SAG1 in the primitive protozoan Giardia duodenalis. Infect. Immun. 70: 1014-1016, 2002. PubMed: 11796643

McArthur AG, et al. The evolutionary origins of eukaryotic protein disulfide isomerase domains: new evidence from the Amitochondriate protist Giardia lamblia. Mol. Biol. Evol. 18: 1455-1463, 2001. PubMed: 11470836

Morrison HG, et al. Giardia lamblia expresses a proteobacterial-like DnaK homolog. Mol. Biol. Evol. 18: 530-541, 2001. PubMed: 11264404

Ellis JG IV, et al. Potential involvement of extracellular signal-regulated kinase 1 and 2 in encystation of a primitive eukaryote, Giardia lamblia. Stage-specific activation and intracellular localization. J. Biol. Chem. 278: 1936-1945, 2003. PubMed: 12397063

Ey PL, Darby JM. VSP417-6, a variant-specific surface protein encoded at a sixth locus within the vsp417 gene subfamily of Giardia intestinalis. Int. J. Parasitol. 32: 425-436, 2002. PubMed: 11849639

Langford TD, et al. Giardia lamblia: identification and characterization of Rab and GDI proteins in a genome survey of the ER to Golgi endomembrane system. Exp. Parasitol. 101: 13-24, 2002. PubMed: 12243734

Sanchez LB, et al. Cloning and sequencing of an acetyl-CoA synthetase (ADP-forming) gene from the amitochondriate protist, Giardia lamblia. Gene 233: 225-231, 1999. PubMed: 10375639

Henze K, et al. Unique phylogenetic relationships of glucokinase and glucosephosphate isomerase of the amitochondriate eukaryotes Giardia intestinalis, Spironucleus barkhanus and Trichomonas vaginalis. Gene 281: 123-131, 2001. PubMed: 11750134

Sun C-H, et al. A novel Myb-related protein involved in transcriptional activation of encystation genes in Giardia lamblia. Mol. Microbiol. 46: 971-984, 2002. PubMed: 12421304

Cross References

Nucleotide (GenBank) : AACB02000000 Giardia lamblia ATCC 50803 strain WB C6, whole genome shotgun sequencing project.