key: cord-0923036-mklu5uus
authors: Walker, G. J.; Clifford, V.; Bansal, N.; Ospina Stella, A.; Turville, S.; Stelzer-Braid, S.; Klein, L. D.; Rawlinson, W.
title: SARS-CoV-2 in human milk is inactivated by Holder pasteurization but not cold storage
date: 2020-06-20
journal: nan
DOI: 10.1101/2020.06.18.20134395
sha: 600114ea9a5f967601999f7fe7605c6b9999b222
doc_id: 923036
cord_uid: mklu5uus

As the COVID-19 pandemic evolves, human milk banks worldwide continue to provide donor human milk to vulnerable infants who lack access to mothers own milk. Under these circumstances, ensuring the safety of donor human milk is paramount, as the risk of vertical transmission of SARS-CoV-2 is not well understood. Here, we investigate the inactivation of SARS-CoV-2 in human milk by pasteurisation, and the stability of SARS-CoV-2 in human milk under cold storage (freezing or refrigeration). Following heating to 63{degrees}C or 56{degrees}C for 30 minutes, SARS-CoV-2 replication competent (i.e. live) virus was undetected in both human milk and the control medium. Cold storage of SARS-CoV-2 in human milk (either at 4{degrees}C or -30{degrees}C) did not significantly impact infectious viral load over a 48 hour period. Our findings demonstrate that SARS-CoV-2 can be effectively inactivated by Holder pasteurisation, and confirm that existing milk bank processes will effectively mitigate the risk of transmission of SARS-COV-2 to vulnerable infants through pasteurised donor human milk.

Mr Walker designed the study, collected the data, analyzed the data, created the figure, drafted, and critically reviewed and revised the manuscript. Dr Clifford conceptualized the study, designed the study, drafted, and critically reviewed and revised the manuscript.

Dr Bansal designed the study, and reviewed and revised the manuscript.

Drs Ospina Stella and Turville did critical work to prepare the laboratory to conduct this research, assisted in data collection, and reviewed the manuscript.

Dr Stelzer-Braid collected the data, analyzed the data, and reviewed the manuscript.

Dr Klein designed the study, drafted the initial manuscript, and critically reviewed and revised the manuscript.

Professor Rawlinson conceptualized the study, coordinated and supervised data collection, and critically reviewed and revised the manuscript.

All authors approved the final manuscript as submitted and agree to be accountable for all aspects of the work.

. CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted June 20, 2020. . https://doi.org/10.1101/2020.06.18.20134395 doi: medRxiv preprint ABSTRACT As the COVID-19 pandemic evolves, human milk banks worldwide continue to provide donor human milk to vulnerable infants who lack access to mother's own milk. Under these circumstances, ensuring the safety of donor human milk is paramount, as the risk of vertical transmission of SARS-CoV-2 is not well understood. Here, we investigate the inactivation of SARS-CoV-2 in human milk by pasteurisation, and the stability of SARS-CoV-2 in human milk under cold storage (freezing or refrigeration). Following heating to 63°C or 56°C for 30 minutes, SARS-CoV-2 replication competent (i.e. live) virus was undetected in both human milk and the control medium. Cold storage of SARS-CoV-2 in human milk (either at 4°C or -30°C) did not significantly impact infectious viral load over a 48 hour period. Our findings demonstrate that SARS-CoV-2 can be effectively inactivated by Holder pasteurisation, and confirm that existing milk bank processes will effectively mitigate the risk of transmission of SARS-COV-2 to vulnerable infants through pasteurised donor human milk.

. CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted June 20, 2020. .

As the COVID-19 pandemic evolves, human milk banks worldwide continue to provide donor human milk to vulnerable infants who lack access to mother's own milk. 1 Under these circumstances, ensuring the safety of donor human milk is paramount, as the risk of vertical transmission of SARS-CoV-2 is not well understood. 2 Although case series have reported that SARS-CoV-2 was not detected in breast milk, 3 is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted June 20, 2020. . https://doi.org/10.1101/2020.06.18.20134395 doi: medRxiv preprint of samples was performed on Vero cells in sextuplicate, and the 50% tissue culture infectious dose (TCID50) determined at 72 hours post-infection using standard methods. 6 Freshly expressed milk from two donors and MEM (control) was similarly inoculated with SARS-CoV-2 to assess virus stability under cold storage conditions. Infectious titres of samples were measured at the time of inoculation, and after 48 hours of storage at 4°C or -30°C. This project was approved by the Australian Red Cross Lifeblood HREC (Clifford 27042020).

Following heating to 63°C or 56°C for 30 minutes, SARS-CoV-2 replication competent (i.e. live) virus was undetected in both milk and MEM (Fig. 1A) . After 48 hours there was no reduction in SARS-CoV-2 infectious titre in milk samples kept at 4°C, and a 0.41 log-unit reduction in samples frozen at -30°C (Fig. 1B) . In the control media (MEM), there was a 0.02 and 0.25 log-unit reduction in infectious titres of samples stored at 4°C and -30°C respectively.

Whilst freezing of milk and MEM samples resulted in a slight reduction in infectious titre, viable virus was still recovered after 48 hours of storage.

Milk banks employ many risk mitigation strategies to ensure the safety of donor milk, with donor selection criteria, serological and/or viral nucleic acid screening, validated frozen transport methods, microbial testing, and pasteurisation. 7 For SARS-CoV-2, this includes specific questions to screen for symptoms of COVID-19, deferral of donors with COVID-19 or close contact with COVID-19, and pasteurisation of donor milk. Holder pasteurisation (62.5ºC for 30 minutes) is the most common pasteurisation method among milk banks worldwide, and has been shown to inactivate most enveloped viruses, including MERS-CoV 8 and SARS-CoV-1. 9 Our findings are consistent with a recent study that reported SARS-CoV-2 is inactivated by heat treatment, 10 although inactivation under Holder pasteurisation has not . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted June 20, 2020. . https://doi.org/10.1101/2020.06.18.20134395 doi: medRxiv preprint yet been investigated in human milk. Our findings demonstrate that SARS-CoV-2 can be effectively inactivated by Holder pasteurisation, and confirm that existing milk bank processes will effectively mitigate the risk of transmission of SARS-COV-2 to vulnerable infants through pasteurised donor human milk. This is the first study to assess the stability of SARS-CoV-2 in human milk. We show that cold storage of SARS-CoV-2 in human milk (either by refrigeration or freezing) does not significantly impact infectious viral load over a 48 hour period. Results of the refrigerated samples are consistent with a recent study not involving human milk that found SARS-CoV-2 to be highly stable at 4 º C in the environment. 10 Whilst it is yet to be determined whether SARS-CoV-2 detected in breast milk is infectious, these findings may assist in the development of guidelines around expressing and storing milk from COVID-19 infected mothers. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted June 20, 2020. . https://doi.org/10.1101/2020.06.18.20134395 doi: medRxiv preprint

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The authors thank Christine Sulfaro, Nicole Stevens, Lifeblood Milk donors, and members of the Serology and Virology Division (SAViD) Area Diagnostic Virology Laboratory and Kirby Institute UNSW for their assistance with this study.