Bartonella Henselae Survives after the Storage Period of the Red Blood Cell Units.
Authors: Magalhaes Renata Ferreira , Pitassi Luiza Helena Urso , Salvadego Marizete , Moraes Aparecida Machado , Barjas-Castro Maria Lourdes , Velho Paulo Eduardo Ferreira 
Published: October 2019

Abstract
Introduction: The genus Bartonella comprises a unique group of emerging, Gram-negative, facultative intracellular bacteria, which can cause Carrión’s disease, trench fever, cat scratch disease and bacillary angiomatosis. Man is a reservoir host of Bartonella species. Ten to 15% of the populations in areas that are hyperendemic for Carrión’s disease are asymptomatic carriers of B. bacilliformis. In addition, positive serologic tests for B. henselae were found in 2 to 6% of Americans, 48% of Swiss, 19% of Germans, and 13.7% of Brazilians. Blood donors can be asymptomatic carrier of Bartonella spp. and the risk for transmission by transfusion should be considered. The aim of this study was to demonstrate that B. henselae remains viable in red blood cell (RBC) units at the end of the storage period.
Methods: Two RBC units from healthy blood donors were collected in CPDA1 and each one split into two portions via a sterile connecting device. One bag from each split unit received experimentally B. henselae (Adolpho Lutz Institute, Sao Paulo, Brazil) and the second one served as a control. A brain and heart infusion (BHI) suspension of B. henselae colonies (Houston 1, American Type Culture Collection, Rockville, MD, ATCC 49882T) was used to obtain equivalence with tube 10 of McFarland scale, which determined an initial suspension with approximately 3×109 colony-forming units (CFU)/mL. This bacterial suspension was inoculated (0,33mL of the initial suspension) in one split RBC unit using a Sampling Site Coupler. Thus approximately 5,5×106 CFU of B. henselae were obtained per mL of RBC concentrate. All split units were stored (4 ±2°C) for 35 days. Aliquots were collected weekly using, Sample Site Coupler for, culture in dish with chocolate agar (incubated at 37°C in environment with 5% CO2), samples for blood culture bottles Bact/Alert (BioMérieux, Inc, USA) and Karnovisky medium for future evaluation by transmission electron microscopy.
Results: All dishes with chocolate agar culture medium from infected units (seeded on days D0, D7, D14, D21, D28 and D35) presented growth of exuberant and mucoid colonies, after the fourth day of incubation. The electron microscopy from these samples showed typical Gram-negative cell wall in the interior of red blood cells. Samples from infected units presented negative blood culture bottles. All cultures from control units had negative results.
Discussion: Bartonella ssp. is fastidious bacteria. Incubation for a short period of time and in standard medium does not allow for B. henselae growth as observed in the bottle culture of this study. Cultivation in blood-enriched media and incubation in environment with high CO2 levels of infected smears resulted positive. In conclusion, this study has demonstrated that B. henselae remain viable in RBC units during the storage period, at 4±2°C. These data reinforce the possibility of infection by transfusion of blood units collected from asymptomatic blood donors. Receptors of blood transfusion many times are or may become immunocompromissed, with risk of developing severe forms of bartonelloses.