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Assessment of Collargol and AgNO3 EC50 on Tetrahymena thermophila CU428 strain used in the gene expression study

Name: 
Angela Piersanti
Team: 
Cristina Miceli and Sandra Pucciarelli – University of Camerino, Italy
Host: 
Katre Juganson – National Institute of Chemical Physics and Biophysics, Estonia
Date: 
Saturday, May 30, 2015 to Saturday, June 6, 2015
Report: 

During this STSM I reported the results of the differential gene expression analysis of RNA-seq samples from cultures of Tetrahymena thermophila CU428 strain stressed for 24 hours with silver nanoparticles and adequate controls to experts in nanoparticles' effects, i.e. Mrs Juganson and her colleagues at the “National Institute of Chemical Physics and Biophysics” in Tallinn. The analyzed samples had been exposed to sub- lethal concentration of toxicants: Collargol 20 mg Ag/L and AgNO3 1.52 mg Ag/L according to the toxicity test results on the same Tetrahymena thermophila strain:

EC50 values (95% confidence intervals)

 

Collargol (mg Ag/L)

AgNO3 (mg Ag/L)

2h

71.6

(67.3-75.1)

1.81

(1.70-1.96)

24 h

78.5

(75.6-81.9)

2.59

(2.35-2.84)

 

Collargol solubility was reported to be 7.6% in deionized water at 10 mg Ag/L (Bondarenko et al. 2013)1; thus it was estimated that at 20 mg Ag/L of Collargol the concentration of Ag-ion equals 20*0.076=1.5 mg Ag/L.

Actually the CU428 strain that we used for gene expression analysis was more sensitive, as was shown by the results of the toxicity test that we performed in the current STSM:

EC50 values (95% confidence intervals)

 

Collargol (mg Ag/L)

AgNO3 (mg Ag/L)

2h

28.1

(25.2-32.4)

0.96

(0.87-0.98)

24 h

38.2

(33.1-45.8)

1.05

(1.02-1.17)

We also used several techniques, such as measuring the absorbance of T. thermophila culture (at 600 nm), counting the cells using haemocytometer, and microscopic evaluation to confirm that incubation for 24 h in MQ water medium does not significantly harm the cells.

Moreover, new unpublished studies demonstrate that while in pure water after 24 h the concentration of Ag- ions released from 20 mg Ag/L of Collargol is similar to the concentration that was used in Ag-ion control, the amount of ions released after 2-h incubation is much lower than what was previously assessed.

The results of the test conducted during current STSM will help to make a decision whether it is necessary to sequence more RNA samples and at which test conditions.

According to the point of view of nanoparticles experts it would be more significant, to assess the mechanism in term of toxicity, to sequence RNA samples exposed to the same concentration but a different time point, e.g. after 2-h exposure. That is because the cells seem to adapt to such an exposition after a while and start to recover.

 

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1Bondarenko, O., Ivask, A., Kaekinen, A., Kurvet, I., and Kahru, A. (2013) Particle-Cell Contact Enhances Antibacterial Activity of Silver Nanoparticles, Plos One 8.

 

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