Evidence of Staphylococcus Aureus Deformation, Proliferation, and Migration in Canaliculi of Live Cortical Bone in Murine Models of Osteomyelitis

Abstract

While Staphylococcus aureus osteomyelitis is considered to be incurable, the major bacterial reservoir in live cortical bone has remained unknown. In addition to biofilm bacteria on necrotic tissue and implants, studies have implicated intracellular infection of osteoblasts and osteocytes as a mechanism of chronic osteomyelitis. Thus, we performed the first systematic transmission electron microscopy (TEM) studies to formally define major reservoirs of S. aureus in chronically infected mouse (Balb/c J) long bone tissue. Although rare, evidence of colonized osteoblasts was found. In contrast, we readily observed S. aureus within canaliculi of live cortical bone, which existed as chains of individual cocci and submicron rod-shaped bacteria leading to biofilm formation in osteocyte lacunae. As these observations do not conform to the expectations of S. aureus as non-motile cocci 1.0–1.5 µm in diameter, we also performed immunoelectron microscopy (IEM) following in vivo BrdU labeling to assess the role of bacterial proliferation in canalicular invasion. The results suggest that the deformed bacteria: 1) enter canaliculi via asymmetric binary fission; and 2) migrate toward osteocyte lacunae via proliferation at the leading edge. Additional in vitro studies confirmed S. aureus migration through a 0.5 µm porous membrane. Collectively, these findings define a novel mechanism of bone infection, and provide possible new insight as to why S. aureus implant related infections of bone tissue are so challenging to treat.

Surface topography of silicon nitride affects antimicrobial and osseointegrative properties of tibial implants in a murine model

“…Staphylococcus aureus is responsible for the majority of chronic osteomyelitis cases; and these bone infections remain a major challenge in orthopaedics . Additionally, these infections are considered to be incurable due to biofilm dwelling bacteria that persist within Staphylococcus abscess communities (SACs or Brodie's abscesses) and deep within cortical bone; and costing billions of dollars each year . Of particular concern is the community‐associated methicillin‐resistant S. aureus (MRSA) strain USA300 LAC, which now accounts for the majority of orthopaedic infections .…”

Gram‐Negative Bacteria Are Internalized Into Osteocyte‐Like Cells

“…Once freed, these bacteria can cause a relapse in infection. Interestingly, de Mesy Bentley et al demonstrated the presence of significant colonies of S. aureus within the canaliculi of live cortical bone. While they show evidence of invasion of the canaliculi by infecting bacteria, it is also possible that some of those bacteria found within the canalicular system arose from infected bone cells.…”

“…Staphylococcus aureus, the most common pathogen identified in orthopaedic infections, 4 has been shown to invade osteoblasts in vitro, [5][6][7][8][9] as well as other eukaryotic cells. [10][11][12][13][14][15][16][17] In vivo, S. aureus has been found internalized in osteoblasts, both in an experimental chick embryo infection model 18 and in a patient with OM, 19 as well as closely associated with bone cells within the canaliculi of bone, both in a chronically infected mouse model, 20 as well as in a patient with an infected diabetic foot ulcer. 21 It has been suggested that such an evasive strategy would allow a pathogen a location safe from both the immune system and those antibiotics incapable of crossing the cell membrane.…”

New developments and future challenges in prevention, diagnosis, and treatment of prosthetic joint infection

“…The failure rate of PJI treatment remains high despite surgical debridement, prosthetic exchange, and targeted systemic antimicrobial agents. Emerging antibiotic resistance, the formation of biofilm, and migration of bacteria to immune‐privileged locations such as the osteocyte lacuna‐canalicular network all contribute to the challenge of treating these infections 105–107 . A summary of promising treatment strategies in PJI is shown in Table 3.…”