Trypanosoma cruzi Survival following Cold Storage: Possible Implications for Tissue Banking Diana L. Martin 1 *, Brook Goodhew 1 , Nancy Czaicki 2 , Kawanda Foster 3 , Srijana Rajbhandary 4 , Shawn Hunter 5 , Scott A. Brubaker 6 1 Centers for Disease Control and Prevention, Atlanta, Georgia, United States of America, 2 University of California, Berkeley, California, United States of America, 3 University of Georgia, Athens, Georgia, United States of America, 4 Center for Medical Technology Policy, Baltimore, Maryland, United States of America, 5 Center for Tissue Innovation and Research, Kettering, Ohio, United States of America, 6 American Association of Tissue Banks, McLean, Virginia, United States of America Abstract While Trypanosoma cruzi, the etiologic agent of Chagas disease, is typically vector-borne, infection can also occur through solid organ transplantation or transfusion of contaminated blood products. The ability of infected human cells, tissues, and cellular and tissue-based products (HCT/Ps) to transmit T. cruzi is dependent upon T. cruzi surviving the processing and storage conditions to which HCT/Ps are subjected. In the studies reported here, T. cruzi trypomastigotes remained infective 24 hours after being spiked into blood and stored at room temperature (N = 20); in 2 of 13 parasite-infected cultures stored 28 days at 4uC; and in samples stored 365 days at 280uC without cryoprotectant (N = 28), despite decreased viability compared to cryopreserved parasites. Detection of viable parasites after multiple freeze/thaws depended upon the duration of frozen storage. The ability of T. cruzi to survive long periods of storage at +4 and 280uC suggests that T. cruzi-infected tissues stored under these conditions are potentially infectious. Citation: Martin DL, Goodhew B, Czaicki N, Foster K, Rajbhandary S, et al. (2014) Trypanosoma cruzi Survival following Cold Storage: Possible Implications for Tissue Banking. PLoS ONE 9(4): e95398. doi:10.1371/journal.pone.0095398 Editor: Zoran Ivanovic, French Blood Institute, France Received February 5, 2014; Accepted March 26, 2014; Published April 23, 2014 This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication. Funding: This research was supported by a grant from the Scientific and Technical Affairs Committee of the American Association of Tissue Banks (AATB, www. aatb.org). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: hzx3@cdc.gov Introduction Trypanosoma cruzi is a parasite that causes human Chagas disease (American trypanosomiasis). Transmission typically occurs in areas of Latin America where substandard housing provides a habitat for the triatomine bug vectors that deposit the parasite in fecal matter during a nighttime blood meal. The parasite can enter the bloodstream through the skin via any break in the skin or through contact with a mucous membrane. In the vertebrate host, the parasite exists in both intracellular and extracellular forms. Extracellular trypomastigotes can invade virtually any nucleated cell type. Once inside the cell, trypomastigotes transform to amastigotes and replicate [1]. Amastigotes transform back to trypomastigotes after approximately nine rounds of replication over 4–7 days and escape the cell. The released trypomastigotes can be taken up by a triatomine vector during a blood meal or can propagate the infection in vivo by infecting other host cells. The acute phase of infection lasts for 1–2 months, during which the parasite has a broad tissue distribution and parasitemia is patent. After the acute phase, parasites persist primarily, but not exclusively, in muscle tissue, and the predominant clinical pathology is cardiomyopathy. Although infection of the host is life-long, parasites are rarely seen in the blood during chronic infection, and even sensitive polymerase chain reaction (PCR) assays only detect parasites in the blood of up to 66% of chronically-infected individuals [2]. To date, 23 cases of vector-borne infection have been identified in the United States (U.S.) [3]. Other potential routes of human transmission in the United States include congenital infection and infection via transplantation and transfusion [4,5]. An estimated 300,000 T. cruzi-infected individuals reside in the U.S. [6]. In the U.S., nine cases of T. cruzi transmission from solid organ donation have been documented [5,7,8]. Eight reported cases of T. cruzi transmission via blood transfusion have been reported; blood donor screening was implemented in the United States in 2007 [9]. The risk of transmission through transplantation of tissues from infected donors is unexplored. Tissue banks oversee the donation of a number of non-solid-organ tissues such as skin, long bone, tendons, ligaments, cornea, heart valves, musculoskeletal tissue, and nerve tissue. Unlike solid organ donation, recovery of tissue may take place up to 15–24 hours after asystole, and many tissues are processed and stored prior to transplantation [10]. While some tissues undergo minimal processing and are stored in cryoprotec- tant to preserve function (e.g. reproductive tissue and heart valves), other tissues undergo more extensive processing and cold storage in the absence of cryoprotectant prior to transplantation. Some tissue, such as musculoskeletal tissue, can be stored at ,240uC in the absence of cryoprotectant for up to 5 years per American Association of Tissue Bank (AATB) Standards [10]. In early 2009, the U.S. Food and Drug Administration (FDA) issued a draft Guidance for Industry (DHHS/FDA/CBER 2009) that suggested all donors of human cells, tissues, and cellular and tissue-based products (HCT/Ps) be screened and tested for antibodies to T. cruzi. This document identified T. cruzi as a ‘‘relevant communi- cable disease agent’’ and described ‘‘current data are insufficient to PLOS ONE | www.plosone.org 1 April 2014 | Volume 9 | Issue 4 | e95398