Osteomyelitis (OM), the most common manifestation of bone infection, is characterized by the inflammatory destruction of bone and bone marrow. Despite antibiotic and surgical treatment, OM can advance to a chronic infection, where it can arise intermittently over a period of years. In the US, an aging population; an increasing prevalence of compounding factors; and improvements in diagnosis have resulted in an increase in the incidence of this chronic state. While Gram positive organisms are the most common causative agent of initial infections, the percentage of Gram negative species increases in reoccurring bone infections. Pathogens have a multitude of virulence factors to take advantage of the pathological features of OM and establish persistent infection, including biofilm formation and multi-drug resistance genes.
The aim of this dissertation was to determine the ability of Gram negative bacteria to internalize into bone cells. Employing the hFOB, MLO-A5, and MLO-Y4 cell lines as cell models, the Gram negative species, Proteus mirabilis, Serratia marcescens, Pseudomonas aeruginosa, and Acinetobacter baumannii, were recovered from exposed cells, following an internalization assay. P. mirabilis and S. marcescens were selected for further characterization of the internalization process, which was both time- and concentration-dependent for these two species, regardless of cell type. Confocal analysis demonstrated the presence of internalized bacteria within all three cell types. Inhibition of cellular endocytic mechanisms reduced internalized bacteria recovered, for both species, indicating that bacterial internalization is, at least in part, cell mediated. While the presence of internalized P. mirabilis did not impact cell viability, measured either by LDH release or MTT activity, the presence of S. marcescens both increased LDH release and reduced MTT activity, indicating a loss of cell viability in response to the microorganism. Despite this difference in cytotoxicity, both bacterial species were recovered 24 h post-infection, suggesting that bacteria could propagate persistent, reoccurring infections by hiding intracellularly and re-infecting proximate cells upon release from bone cells. While relapsing or chronic bone infections are typically attributed to biofilm, the results herein imply that the ability of Gram negative bacteria to internalize into bone cells may be as relevant to chronic OM as biofilm-associated bacteria.
|School:||University of the Sciences in Philadelphia|
|School Location:||United States -- Pennsylvania|
|Source:||DAI-B 80/08(E), Dissertation Abstracts International|
|Subjects:||Cellular biology, Microbiology|
|Keywords:||Bacterial internalization, Bone cells, Bone infection, Gram negative bacteria, Osteomyelitis|
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