Virulence of Bacteria Colonizing Vascular Bundles in Ischemic Lower Limbs.

BACKGROUND: We documented previously the presence of bacterial flora in vascular bundles, lymphatics, and lymph nodes of ischemic lower limbs amputated because of multifocal atheromatic changes that made them unsuitable for reconstructive surgery and discussed their potential role in tissue destruction. The question arose why bacterial strains inhabiting lower limb skin and considered to be saprophytes become pathogenic once they colonize deep tissues. Bacterial pathogenicity is evoked by activation of multiple virulence factors encoded by groups of genes. METHODS: We identified virulence genes in bacteria cultured from deep tissue of ischemic legs of 50 patients using a polymerase chain reaction technique. RESULTS: The staphylococcal virulence genes fnbA (fibronectin-binding protein A), cna (collagen adhesin precursor), and ica (intercellular adhesion) were present in bacteria isolated from both arteries and, to a lesser extent, skin. The IS256 gene, whose product is responsible for biofilm formation, was more frequent in bacteria retrieved from the arteries than skin bacteria. Among the virulence genes of Staphylococcus epidermidis encoding autolysin atlE, icaAB (intercellular adhesion), and biofilm insert IS256, only the latter was detected in arterial specimens. Bacteria cultured from the lymphatics did not reveal expression of eta and IS256 in arteries. The Enterococcus faecalis asa 373 (aggregation substance) and cylA (cytolysin activator) frequency was greater in arteries than in skin bacteria, as were the E. faecium cyl A genes. All Pseudomonas aeruginosa virulence genes were present in bacteria cultured from both the skin and arteries. Staphylococci colonizing arterial bundles and transported to tissues via ischemic limb lymphatics expressed virulence genes at greater frequency than did those dwelling on the skin surface. Moreover, enterococci and Pseudomonas isolated from arterial bundles expressed many virulence genes. CONCLUSIONS: These findings may add to the understanding of the mechanism of development of destructive changes in lower limb ischemic tissues by the patient's, but not hospital-acquired, bacteria, as well as the generally unsatisfactory results of antibiotic administration in these cases. More aggressive antibiotic therapy targeted at the virulent species should be applied.

Slime production by Staphylococcus aureus and Staphylococcus epidermidis strains isolated from patients with diabetic foot ulcers.

Slime production is a very important factor related to biofilm formation. The objective of the present study was to determine the frequency of slime production by Staphylococcus aureus and Staphylococcus epidermidis strains recovered from 50 patients with diabetic foot ulcers. Slime production was determined using the Congo red agar (CRA) method and compared with immunocytochemistry for the production of polysaccharide intercellular adhesin (PIA). Out of 55 S. aureus strains, 69% produced slime as shown by the CRA method. Of them, 84.2% also produced PIA. Of 17 CRA-negative strains, 70.6% produced PIA. Out of 20 S. epidermidis strains, 75% were CRA positive and 93.3% produced PIA. All CRA-negative S. epidermidis produced PIA. In conclusion, PIA production is a very common trait of S. aureus and S. epidermidis isolates obtained from diabetic foot ulcer patients.

Epidemiology and prevalence of methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis in patients with diabetic foot ulcers: focus on the differences between species isolated from individuals with ischemic vs. neuropathic foot ulcers.

We examined whether foot ischemia or neuropathy with diabetic foot ulcer (DFU) promote selection of staphylococci species, evaluated frequency of MRSA and MRSE among strains yielded from patients with DFU and assessed multidrug resistance of isolates. Patients with DFU and foot osteomyelitis were divided into ischemic foot ulcer (IFU, n=21) and neuropathic foot ulcer (NFU, n=29) groups. Frequency of Staphylococcus epidermidis yielded from curettage of IFU was higher compared with NFU (P<0.05). S. epidermidis was also more frequently isolated from the toe web surface of patients with IFU compared with NFU (55% vs. 17.9%, respectively) and healthy volunteers (HV, n=20) (17.6%, P<0.05). These mostly MRSE strains (83.3-100%) originating from DFU patients were multidrug resistant (88.8%). Also, most of MRSA isolates were multidrug resistant (70.3%). Higher rates of MSSA from DFU patients than HV showed resistance to antimicrobials. This is the first report indicating that diabetic patients with IFU differ with NFU patients in higher frequency of S. epidermidis skin colonization and ulcer infection. We suggest that IFU should be defined as separate disease state of DFU and S. epidermidis should be appreciated as a nosocomial pathogen.