No description available.

No description available.

Bacterial contamination of platelet concentrates (PCs) poses the major transfusion-associated infectious risk. Coagulase negative staphylococci (CoNS), the predominant platelet contaminants, are recognized as one of the leading causes of hospital-acquired infections due to their ability to form biofilms (surface-attached aggregates). In this study, 29 CoNS strains were characterized for their growth and biofilm formation abilities in media and PCs. Twenty-five strains were aerobic including Staphylococcus epidermidis, S. capitis, and S. chromogenes, while four were identified as the anaerobe S. saccharolyticus. Biofilm-associated icaA and icaD genes were amplified from eight strains. Interestingly, only six of those strains were biofilm-positive. Sequencing of S. capitis icaD revealed no mutations that could explain differences in biofilm phenotypes. Growth of CoNS in PCs varied significantly between strains. This study provides preliminary evidence that slow-growing biofilm-positive S. epidermidis are more likely to be missed during platelet culture, highlighting the need for improved screening methods.

In atherosclerosis, infiltrating leukocytes and vascular smooth muscle cells (VSMCs) cause progressive vascular narrowing. Platelet-mediated thrombosis ultimately causes complete vessel occlusion, resulting in heart attack or stroke. In animal models and human patients, individually blocking these events is only partially effective. Another therapeutic strategy would be to globally target these multiple cell types. Slit proteins act as developmental neuronal repellents, and Slit2 via interaction with its receptor, Robo-1, impairs inflammatory recruitment of leukocytes and VSMCs. We detected Robo-1 expression in human and murine platelets. Using static and shear assays, we demonstrate that Slit2 impaired platelet adhesion and spreading on fibrinogen, fibronectin and collagen. Slit2 mediated these effects, in part, by suppressing activation of Akt but not Rac1, Cdc42, Erk or p38 MAPK. Slit2 also prevented ADP-mediated granular secretion. In mouse tail-bleeding experiments, Slit2 dose-dependently prolonged bleeding times in vivo. These data suggest a therapeutic role of Slit2 in atherothrombosis.

In atherosclerosis, infiltrating leukocytes and vascular smooth muscle cells (VSMCs) cause progressive vascular narrowing. Platelet-mediated thrombosis ultimately causes complete vessel occlusion, resulting in heart attack or stroke. In animal models and human patients, individually blocking these events is only partially effective. Another therapeutic strategy would be to globally target these multiple cell types. Slit proteins act as developmental neuronal repellents, and Slit2 via interaction with its receptor, Robo-1, impairs inflammatory recruitment of leukocytes and VSMCs. We detected Robo-1 expression in human and murine platelets. Using static and shear assays, we demonstrate that Slit2 impaired platelet adhesion and spreading on fibrinogen, fibronectin and collagen. Slit2 mediated these effects, in part, by suppressing activation of Akt but not Rac1, Cdc42, Erk or p38 MAPK. Slit2 also prevented ADP-mediated granular secretion. In mouse tail-bleeding experiments, Slit2 dose-dependently prolonged bleeding times in vivo. These data suggest a therapeutic role of Slit2 in atherothrombosis.

Bacterial infection is a leading cause of morbidity and mortality arising from platelet transfusions (1, 2). Storage of platelet products at room temperature (20 to 24ºC) provides ideal conditions for bacterial proliferation (1, 3-6). Furthermore, platelets are stored in plasma containing bioavailable iron that bacteria require to survive (7). Thus we hypothesize that the inclusion of iron chelators will bind and remove iron, thereby inhibiting bacterial growth in both culture medium and platelet concentrates. Additionally, we hypothesize that residual red blood cells (RBCs) in platelet units may contribute bioavailable iron that promotes bacterial growth. To test these hypotheses, we first assessed growth of Staphylococcus epidermidis in culture medium after treatment with the iron chelators deferoxamine (DFO) or phytic acid. DFO significantly inhibited bacterial growth in a dose dependent manner (p < 0.009). Conversely, phytate only inhibited bacterial growth at concentrations ≥ 100 mM (p < 0.001); at ≤ 5 mM, phytate supplied S. epidermidis with additional nutrients and significantly promoted growth (p < 0.001). Subsequently, we monitored the change in RBCs over time. Hemolysis, methemoglobin, and iron levels all significantly increased over the 7-day storage period (p < 0.001) releasing bioavailable iron. Indeed, we found that S. epidermidis growth in iron-poor medium drastically increased with the addition of RBCs, thus supporting our second hypothesis. Surprisingly, the inclusion of DFO in minimal medium did not demonstrate a bacteriostatic effect in the presence of RBCs. The inhibitory effect of DFO was likely overcome by iron released from the elevated methemoglobin levels arising from the direct interaction of DFO with hemoglobin. Previous studies demonstrate that methemoglobin releases iron more quickly than normal hemoglobin (8). Lastly, we evaluated the effect of DFO on microbial growth in platelet concentrates using the BacT/ALERT system. The presence of DFO significantly inhibited S. epidermidis growth in buffy coat platelets in a dose dependent manner (p < 0.001). With these findings, the inclusion of iron chelators is a promising approach to preventing transfusion-transmitted bacterial infection and providing patients with a safer platelet product.

Bacterial contamination of platelet concentrates (PCs) poses the major transfusion-associated infectious risk. Coagulase negative staphylococci (CoNS), the predominant platelet contaminants, are recognized as one of the leading causes of hospital-acquired infections due to their ability to form biofilms (surface-attached aggregates). In this study, 29 CoNS strains were characterized for their growth and biofilm formation abilities in media and PCs. Twenty-five strains were aerobic including Staphylococcus epidermidis, S. capitis, and S. chromogenes, while four were identified as the anaerobe S. saccharolyticus. Biofilm-associated icaA and icaD genes were amplified from eight strains. Interestingly, only six of those strains were biofilm-positive. Sequencing of S. capitis icaD revealed no mutations that could explain differences in biofilm phenotypes. Growth of CoNS in PCs varied significantly between strains. This study provides preliminary evidence that slow-growing biofilm-positive S. epidermidis are more likely to be missed during platelet culture, highlighting the need for improved screening methods.

Bacterial infection is a leading cause of morbidity and mortality arising from platelet transfusions (1, 2). Storage of platelet products at room temperature (20 to 24ºC) provides ideal conditions for bacterial proliferation (1, 3-6). Furthermore, platelets are stored in plasma containing bioavailable iron that bacteria require to survive (7). Thus we hypothesize that the inclusion of iron chelators will bind and remove iron, thereby inhibiting bacterial growth in both culture medium and platelet concentrates. Additionally, we hypothesize that residual red blood cells (RBCs) in platelet units may contribute bioavailable iron that promotes bacterial growth. To test these hypotheses, we first assessed growth of Staphylococcus epidermidis in culture medium after treatment with the iron chelators deferoxamine (DFO) or phytic acid. DFO significantly inhibited bacterial growth in a dose dependent manner (p < 0.009). Conversely, phytate only inhibited bacterial growth at concentrations ≥ 100 mM (p < 0.001); at ≤ 5 mM, phytate supplied S. epidermidis with additional nutrients and significantly promoted growth (p < 0.001). Subsequently, we monitored the change in RBCs over time. Hemolysis, methemoglobin, and iron levels all significantly increased over the 7-day storage period (p < 0.001) releasing bioavailable iron. Indeed, we found that S. epidermidis growth in iron-poor medium drastically increased with the addition of RBCs, thus supporting our second hypothesis. Surprisingly, the inclusion of DFO in minimal medium did not demonstrate a bacteriostatic effect in the presence of RBCs. The inhibitory effect of DFO was likely overcome by iron released from the elevated methemoglobin levels arising from the direct interaction of DFO with hemoglobin. Previous studies demonstrate that methemoglobin releases iron more quickly than normal hemoglobin (8). Lastly, we evaluated the effect of DFO on microbial growth in platelet concentrates using the BacT/ALERT system. The presence of DFO significantly inhibited S. epidermidis growth in buffy coat platelets in a dose dependent manner (p < 0.001). With these findings, the inclusion of iron chelators is a promising approach to preventing transfusion-transmitted bacterial infection and providing patients with a safer platelet product.

Thesis (MAppSc)--University of South Australia, 1997

Thesis (MAppSc)--University of South Australia, 1998