Bioburden after Staphylococcus aureus inoculation in type 1 diabetic rats undergoing internal fixation.

SUMMARY: Fracture stabilization in the diabetic patient is associated with higher complication rates, particularly infection and impaired wound healing, which can lead to major tissue damage, osteomyelitis, and higher amputation rates. With an increasing prevalence of diabetes and an aging population, the risks of infection of internal fixation devices are expected to grow. Although numerous retrospective clinical studies have identified a relationship between diabetes and infection, currently there are few animal models that have been used to investigate postoperative surgical-site infections associated with internal fixator implantation and diabetes. The authors therefore refined the protocol for inducing hyperglycemia and compared the bacterial burden in controls to pharmacologically induced type 1 diabetic rats after undergoing internal fracture plate fixation and Staphylococcus aureus surgical-site inoculation. Using an initial series of streptozotocin doses, followed by optional additional doses to reach a target blood glucose range of 300 to 600 mg/dl, the authors reliably induced diabetes in 100 percent of the rats (n = 16), in which a narrow hyperglycemic range was maintained 14 days after onset of diabetes (mean ± SEM, 466 ± 16 mg/dl; coefficient of variation, 0.15). With respect to their primary endpoint, the authors quantified a significantly higher infectious burden in inoculated diabetic animals (median, 3.2 × 10 colony-forming units/mg dry tissue) compared with inoculated nondiabetic animals (7.2 × 10 colony-forming units/mg dry tissue). These data support the authors' hypothesis that uncontrolled diabetes adversely affects the immune system's ability to clear Staphylococcus aureus associated with internal hardware.

The nature of innate and adaptive interleukin-17A responses in sham or bacterial inoculation

Streptococcus pyogenes is the causative agent of numerous diseases ranging from benign infections (pharyngitis and impetigo) to severe infections associated with high mortality (necrotizing fasciitis and bacterial sepsis). As with other bacterial infections, there is considerable interest in characterizing the contribution of interleukin-17A (IL-17A) responses to protective immunity. We here show significant il17a up-regulation by quantitative real-time PCR in secondary lymphoid organs, correlating with increased protein levels in the serum within a short time of S. pyogenes infection. However, our data offer an important caveat to studies of IL-17A responsiveness following antigen inoculation, because enhanced levels of IL-17A were also detected in the serum of sham-infected mice, indicating that inoculation trauma alone can stimulate the production of this cytokine. This highlights the potency and speed of innate IL-17A immune responses after inoculation and the importance of proper and appropriate controls in comparative analysis of immune responses observed during microbial infection.

Re-inoculation of autoclaved soil as a non-sterile treatment for xenobiotic sorption and biodegradation studies

Autoclaved soil is commonly used for the study of xenobiotic sorption and as an abiotic control in biodegradation experiments. Autoclaving has been reported to alter soil physico-chemical and xenobiotic sorption characteristics such that comparison of autoclaved with non-autoclaved treatments in soil aging and bioavailability studies may yield misleading results. Experiments could be improved by using autoclaved soil re-inoculated with indigenous microorganisms as an additional or alternative non-sterile treatment for comparison with the sterile, autoclaved control. We examined the effect of autoclaving (3 x 1 h, 121°C, 103.5 KPa) on the physico-chemical properties of a silt loam soil (pH 7.2, 2.3% organic carbon) and the establishment of indigenous microorganisms reintroduced after autoclaving. Sterilisation by autoclaving significantly (p ≤ 0.05) decreased pH (0.6 of a unit) and increased concentrations of water-soluble organic carbon (WSOC; nontreated = 75 mg kg^-1; autoclaved = 1526 mg kg^-1). The initial first-order rate of ¹⁴C-2,4-dichloro-UL-phenol (2,4-DCP) adsorption to non-treated, autoclaved and re-inoculated soil was rapid (K₁ = 16.8-24.4 h^-1) followed by a slower linear phase (K₂). In comparison with autoclaved soil (0.038% day^-1), K₂ values were higher for re-inoculated (0.095% day^-1) and nontreated (0.181% day^-1) soil. This was attributed to a biological process. The Freundlich adsorption coefficient (K(f)) for autoclaved soil was significantly (p ≤ 0.05) higher than for re-inoculated or non-treated soil. Increased adsorption was attributed to autoclaving-induced changes to soil pH and solution composition. Glucose-induced respiration of autoclaved soil after re-inoculation was initially twice that in the non-treated control, but it decreased to control levels by day 4. This reduction corresponded to a depletion of WSOC. 2,4-DCP mineralisation experiments revealed that the inoculum of nonsterile soil (0.5 g) contained 2,4-DCP-degrading microorganisms capable of survival in autoclaved soil. The lag phase before detection of significant 2,4-DCP mineralisation was reduced (from 7 days to ≤3 days) by pre-incubation of re-inoculated soils for 7 and 14 days before 2,4-DCP addition. This was attributed to the preferential utilisation of WSOC prior to the onset of 2,4-DCP mineralisation. Cumulative ¹⁴CO₂ evolved after 21 days was significantly lower (p ≤ 0.05) from non-treated soil (25.3%) than re-inoculated soils (ca 45%). Experiments investigating sorption-biodegradation interactions of xenobiotics in soil require the physico-chemical properties of sterile and non-sterile treatments to be as comparable as possible. For fundamental studies, we suggest using re-inoculated autoclaved soil as an additional or alternative non-sterile treatment.

Inoculation with Metal-Mobilizing Plant-GrowthPromoting Rhizobacterium Bacillus sp. SC2b and Its Role in Rhizoremediation

A plant growth-promoting bacterial (PGPB) strain SC2b was isolated from the rhizosphere of Sedum plumbizincicola grown in lead (Pb)/zinc (Zn) mine soils and characterized as Bacillus sp. based on (1) morphological and biochemical characteristics and (2) partial 16S ribosomal DNA sequencing analysis. Strain SC2b exhibited high levels of resistance to cadmium (Cd) (300 mg/L), Zn (730 mg/L), and Pb (1400 mg/L). This strain also showed various plant growth-promoting (PGP) features such as utilization of 1-aminocyclopropane-1-carboxylate, solubilization of phosphate, and production of indole-3-acetic acid and siderophore. The strain mobilized high concentration of heavy metals from soils and exhibited different biosorption capacity toward the tested metal ions. Strain SC2b was further assessed for PGP activity by phytagar assay with a model plant Brassica napus. Inoculation of SC2b increased the biomass and vigor index of B. napus. Considering such potential, a pot experiment was conducted to assess the effects of inoculating the metal-resistant PGPB SC2b on growth and uptake of Cd, Zn and Pb by S. plumbizincicola in metal-contaminated agricultural soils. Inoculation with SC2b elevated the shoot and root biomass and leaf chlorophyll content of S. plumbizincicola. Similarly, plants inoculated with SC2b demonstrated markedly higher Cd and Zn accumulation in the root and shoot system, indicating that SC2b enhanced Cd and Zn uptake by S. plumbizincicola through metal mobilization or plant-microbial mediated changes in chemical or biological soil properties. Data demonstrated that the PGPB Bacillus sp. SC2b might serve as a future biofertilizer and an effective metal mobilizing bioinoculant for rhizoremediation of metal polluted soils.