How do bacteria behave during buffy coat platelet production?

Collected in March 2017

What is this research about?

Platelets are important for controlling bleeding and healing wounds. Platelet transfusions can restore normal platelet levels in patients who have platelet disorders or who have undergone hemorrhaging or chemotherapy. One of the major problems in transfusion medicine is the bacterial contamination of platelet units, which causes transfusion reactions, sometimes with fatal outcomes. Bacteria that are normally found on the skin are the main contaminants.

Platelets are collected from donors by apheresis (platelets are obtained directly from the circulating blood of a single donor, using an apheresis machine) or by the buffy-coat method (whole blood is collected from four donors and processed to isolate the platelets, which are then combined into a single platelet unit for transfusion). The buffy-coat method involves an overnight hold of the donated blood before processing and a resting period of two hours of the whole-blood-derived buffy-coats. The apheresis method does not include these two holding steps.

Interestingly, bacterial contamination is less common in platelet units obtained by the buffy-coat method than in those obtained by apheresis. Because the reasons behind this pattern were unclear, our researchers investigated whether the extra holding steps in the buffy-coat production method eliminate bacteria. Understanding the mechanisms of bacterial survival during the buffy-coat preparation method would enable the identification of the steps that can be optimized to minimize the risk of platelet contamination, increasing the safety of platelet supply for Canadians.

In Brief: Bacterial contamination of platelet units is a serious health concern. This study shows how bacterial growth is affected by the buffy coat platelet production process.

What did the researchers do?

To mimic bacterial contamination of platelets, the researchers added bacteria to units of whole blood donated for research and measured bacteria levels throughout the platelet buffy-coat process. Eight different bacterial types, some of which were reported platelet contaminants, were used to contaminate whole blood. The researchers wanted to test whether the overnight hold and two-hour resting time altered bacterial growth or survival. Also, they tracked bacteria during the manufacturing process to examine how bacteria are divided among the different fractions (such as red blood cells and plasma). Another goal of this study was to monitor changes in platelet quality markers as possible indicators of bacterial contamination of the platelet product.

Whole blood units were collected from different donors and one type of bacteria was added at a known concentration to one of the four units.
Standard procedure for platelet buffy-coat production method was followed.
Bacterial concentration was measured during different stages of production.
Platelet quality was checked by examining levels of the platelet activation marker CD62P and the platelet response to dynamic light scattering on days 1 and 5 of storage.

What did the researchers find?

Bacterial growth during the overnight-hold step varied among the bacteria types; some types were eliminated by this step whereas others multiplied to high concentrations or stayed alive but did not multiply. Bacteria segregated differently in the different fractions, with more bacteria ending up in the cellular fractions (red blood cells and buffy-coat fraction) than in plasma during the buffy-coat procedure. This segregation contributed to a reduction in bacterial concentration in the final platelet unit. Bacterial levels were reduced by the filtration of the platelet units to remove white blood cells. The quality markers that were checked (CD62P and dynamic light scattering) did not consistently indicate bacterial contamination of the platelets.

How can you use this research?

The researchers had shown that the overnight hold of whole blood during the buffy-coat process eliminated some types of bacteria but not others.

Platelet units are currently screened for bacteria by removing a small volume from the unit and transferring it into a liquid culture bottle that is then incubated to detect bacterial presence. Platelet units contaminated with bacterial types that proliferate during the overnight-hold step would allow sufficient levels of bacteria in the final product of platelets which means that there is a high possibility of bacteria being detected by routine screening methods. On the other hand, the bacterial types that do not multiply are reduced significantly in the final platelet unit compared to whole blood due to their segregation in the different fractions. This imposes a risk of not detecting bacterial contamination during the platelet screening process (false negative test result), which might result in transfusing a contaminated unit to a patient. Based on that, thought should be given to screening whole blood samples obtained after the overnight hold before the buffy-coat manufacturing process to help improve the safety of platelets.

In conclusion, this research aimed to explore the reason(s) for the lower bacterial contamination rates associated with the buffy-coat method. The researchers showed that the overnight hold of whole blood decreases the numbers of some bacteria, which could be contributing to the lower contamination pattern, along with other factors during the manufacturing process.

About the research team: This research was conducted in the laboratory of Dr. Sandra Ramirez-Arcos, a development scientist with the product and process development group at the Canadian Blood Services Centre for Innovation. Dr. Mariam Taha is a former PhD student at Dr. Ramirez-Arcos’ laboratory. Dr. Qi-Long Yi, is a senior biostatistician at Canadian Blood Services. Craig Jenkins is a senior manager- product and process development at Canadian Blood Services’ Centre for Innovation and Dr. Peter Schubert is a research associate at the Canadian Blood Services Centre for Innovation and a clinical associate professor in the department of pathology and laboratory medicine at the University of British Columbia. The research was done in collaboration with Dr. Miloslav Kalab, an honorary research scientist at Agriculture and Agri-Food Canada. Dr. Elisabeth Maurer is the founder of LightIntegra Technology Inc.

This ResearchUnit is derived from the following publication:

1. Taha M, Kalab M, Yi QL, Maurer E, Jenkins C, Schubert P, Ramirez-Arcos S. Bacterial Survival and Distribution During Buffy Coat Platelet Production. Vox Sang 2016. 111(4):333-340.

Acknowledgements: Dr. Mariam Taha received a Graduate Fellowship awarded by Canadian Blood Services and Health Canada. This research received financial support from Health Canada and Canadian Blood Services, and was also partially funded by LightIntegra Technology Inc., who provided the machine (ThromboLUX) and the reagents for the machine and helped in analysing the dynamic light scattering data. Canadian Blood Services is funded by the federal government (Health Canada) and provincial and territorial ministries of health. The views herein do not reflect the views of the federal, provincial, or territorial governments of Canada. Canadian Blood Services is grateful to the blood donors who made this research possible.

This ResearchUnit was prepared by Dr. Mariam Taha.

Keywords: Bacterial contamination, platelets, buffy-coat, platelet production

Want to know more? Contact Dr. Sandra Ramirez-Arcos at sandra.ramirez@blood.ca.

ResearchUnit is a knowledge mobilization tool developed by Canadian Blood Services’ Centre for Innovation