I thank Dr. Costa Borges and Embryotool team for this interesting commentary.

 

However, I would comment the authors' sentence:

 

"the installation and use of liquid nitrogen sterilizers is a practice that is still not widespread among IVF laboratories possible due to disbelief in contamination risks or technology insufficiencies"

 

I'm glad to inform the Embryotool team that the technology for sterilizing liquid nitrogen is not anymore insufficient and specific medical devices certified according with 93/42 CEE regulation are available on the market to perform a straightforward washing (as they suggested) with sterile liquid nitrogen (1).

 

Furthermore, the real risk in this pandemic is not related with open or closed vitririfcation systems but with the liquid nitrogen and nitrogen vapor (LN2/NV) themselves. It is well known that they can become contaminated by viruses, bacteria, and fungi (2,3). Airborne contaminants come in contact with LN2/NV and remain cryopreserved (3). 

 

Aerosol and surface stability of Covid-19 has been investigated (4) but the risk of this virus being accidentally cryopreserved in LN2/NV has not yet been studied. The use of contaminated LN2/NV risks virus awakening and the contamination of thawing cells, environment, and operators.

 

In conclusion, we should be focused on the real problem which is LN2/NV contamination that can occur at any step (from manufacturing to final use) and handling, rather than on the hypothetical Covid-19 presence in our gamete and embryos. 

 

Yours sincerely

 

Dr. Lodovico Parmegiani PhD

 

Head of Embryology, NextClinics International

 

Director, IVF Laboratory, GynePro Medical Centers

Via T. Cremona,8 40137 Bologna-Italy

 

 

Conflict of Interest Disclosure: Dr Parmegiani is shareholder of Nterilizer Srl

 

 

References:

 

Parmegiani L, Accorsi A, Bernardi S, Arnone A, Cognigni GE, Filicori M.  A reliable procedure for decontamination before thawing of human specimens cryostored in liquid nitrogen: three washes with sterile liquid nitrogen (SLN2). Fertil Steril. 2012; 98:870-5

Tedder RS, Zukerman MA, Goldstone AH, Hawkins AE, Fielding A, Briggs EM et al. Hepatitis B transmission from contaminated cryopreservation tank. Lancet 1995; 346: 137-140.

Morris GJ. The origin, ultrastructure, and microbiology of the sediment accumulating in liquid nitrogen storage vessels. Cryobiology 2005; 50:231–8.

van Doremalen N, Bushmaker T, Morris DH, Holbrook MG, Gamble A, Williamson BN et al. Aerosol and Surface Stability of SARS-CoV-2 as Compared with SARS-CoV-1. N Engl J Med. 2020

I thank Dr. Costa Borges and Embryotool team for this interesting commentary.

However, I would comment the authors' sentence:

"the installation and use of liquid nitrogen sterilizers is a practice that is still not widespread among IVF laboratories possible due to disbelief in contamination risks or technology insufficiencies"

I'm glad to inform the Embryotool team that the tecnology for sterilizing liquid nitrogen is not anymore insufficient and specific medical devices certified accoding with 93/42 CEE regulation are availble on the market to perform a straightforward washing (as they suggested) with sterile liquid nitrogen (1).

Furthermore, the real risk in this pandemic is not related with open or closed vitririfcation sistems but with the liquid nitrogen and nitrogen vapor (LN2/NV) themeselves. It is well known that they can become contaminated by viruses, bacteria, and fungi (2,3). Airborne contaminants come in contact with LN2/NV and remain cryopreserved (3). 

Aerosol and surface stability of Covid-19 has been investigated (4) but the risk of this virus being accidentally cryopreserved in LN2/NV has not yet been studied. The use of contaminated LN2/NV risks virus awakening and the contamination of thawing cells, environment and operators.

In conclusion, we should be focused on the real problem which is LN2/NV contamination that can occur at any step (from manufacturing to final use) and handling, rather than on the hypotethical Covid-19 presence in our gamete and embryos. 

Yours sincerely

Dr. Lodovico Parmegiani PhD

Head of Embryology, NextClinics International

Director, IVF Laboratory, GynePro Medical Centers
Via T. Cremona,8 40137 Bologna-Italy

Conflict of Interest Disclosure: Dr Parmegiani is shareholder of Nterilizer Srl

References:

1. Parmegiani L, Accorsi A, Bernardi S, Arnone A, Cognigni GE, Filicori M.  A reliable procedure for decontamination before thawing of human specimens cryostored in liquid nitrogen: three washes with sterile liquid nitrogen (SLN2). Fertil Steril. 2012; 98:870-5
2. Tedder RS, Zukerman MA, Goldstone AH, Hawkins AE, Fielding A, Briggs EM et al. Hepatitis B transmission from contaminated cryopreservation tank. Lancet 1995; 346: 137-140.
3. Morris GJ. The origin, ultrastructure, and microbiology of the sediment accumulating in liquid nitrogen storage vessels. Cryobiology 2005; 50:231–8.
4. van Doremalen N, Bushmaker T, Morris DH, Holbrook MG, Gamble A, Williamson BN et al. Aerosol and Surface Stability of SARS-CoV-2 as Compared with SARS-CoV-1. N Engl J Med. 2020 Mar 17.

To the editor. I read with interest the article by Molina et al. in vol. 106 N° 1 / July 2016 of this journal in which the authors suggested that cryostorage containers “should be emptied and cleaned periodically because of the risk of lost straws or small particles of contaminated material” and that “periodic chemical sterilization of liquid nitrogen (LN2) containers is recommended for oocyte and embryo banking”(1). Regarding procedures for decontamination I would like to suggest that the easiest way to sterilize the inner walls and the liquid nitrogen inside tanks is by ultraviolet (UV) radiation. As is common practice in today’s IVF labs (2- 5), by using specifically designed devices available on the open market which ensure sterilization and certification of LN2 and N vapours (6). These devices also allow the sanification of dewars containing the LN2. The simplicity of this technology permits the decontamination of the storage containers with temporary removal of specimens but not requiring that dewars are emptied of LN2. Thus, it is possible to perform a periodic decontamination of cryostorage containers via UV radiation simply and without the use of chemicals. Thank you for the opportunity to clarify this point. Lodovico Parmegiani M.Sc.; Ph.D. GynePro Medical Centers Bologna, ITALY E-mail: l.parmegiani@gynepro.it DISCLOSURE STATEMENT L. Parmegiani holds an international patent (PCT/IB2009/007801: Device and method for sterilizing liquid nitrogen by ultraviolet radiation). REFERENCES 1. Molina I, Mari M, Martínez JV, Novella-Maestre E, Pellicer N, Pemán J. Bacterial and fungal contamination risks in human oocyte and embryo cryopreservation: open versus closed vitrification systems. Fertil Steril 2016;106:127-132 2. Parmegiani L, Cognigni GE, Bernardi S, Cuomo S, Ciampaglia W, Infante FE, Tabarelli de Fatis C, Arnone A, Maccarini AM, Filicori M, Efficiency of aseptic open vitrification and hermetical cryostorage of human oocytes. Reprod Biomed Online 2011; 23: 505–512. 3. Parmegiani L, Accorsi A, Bernardi S, Arnone A, Cognigni GE, Filicori M, A reliable procedure for decontamination before thawing of human specimens cryostored in liquid nitrogen: three washes with sterile liquid nitrogen (SLN2). Fertil Steril 2012; 98: 870–875. 4. Parmegiani L, Accorsi A, Cognigni GE, Bernardi S, Troilo E, Filicori M. Sterilization of liquid nitrogen with ultraviolet irradiation for safe vitrification of human oocytes or embryos. Fertil Steril 2010; 94: 1525-1528. 5. Parmegiani L, Cognigni GE, Filicori M. Ultra-violet sterilization of liquid nitrogen prior to vitrification. Hum Reprod 2009; 24:11. 6. Parmegiani L., Troilo E., Arnone A., Maccarini A.M., Rastellini A., Lanzilotti S., Bernardi S. Aseptic procedures for vitrification, warming and cryostorage. Current Trends in Clinical Embryology 2014; 1: 17- 22 . doi: 10.11138/cce/2014.1.1.017