Hereditary hemochromatosis promotes colitis and colon cancer and causes bacterial dysbiosis in mice.

Hereditary hemochromatosis (HH), an iron-overload disease, is a prevalent genetic disorder. As excess iron causes a multitude of metabolic disturbances, we postulated that iron overload in HH disrupts colonic homeostasis and colon-microbiome interaction and exacerbates the development and progression of colonic inflammation and colon cancer. To test this hypothesis, we examined the progression and severity of colitis and colon cancer in a mouse model of HH (Hfe-/-), and evaluated the potential contributing factors. We found that experimentally induced colitis and colon cancer progressed more robustly in Hfe-/- mice than in wild-type mice. The underlying causes were multifactorial. Hfe-/- colons were leakier with lower proliferation capacity of crypt cells, which impaired wound healing and amplified inflammation-driven tissue injury. The host/microflora axis was also disrupted. Sequencing of fecal 16S RNA revealed profound changes in the colonic microbiome in Hfe-/- mice in favor of the pathogenic bacteria belonging to phyla Proteobacteria and TM7. There was an increased number of bacteria adhered onto the mucosal surface of the colonic epithelium in Hfe-/- mice than in wild-type mice. Furthermore, the expression of innate antimicrobial peptides, the first-line of defense against bacteria, was lower in Hfe-/- mouse colon than in wild-type mouse colon; the release of pro-inflammatory cytokines upon inflammatory stimuli was also greater in Hfe-/- mouse colon than in wild-type mouse colon. These data provide evidence that excess iron accumulation in colonic tissue as happens in HH promotes colitis and colon cancer, accompanied with bacterial dysbiosis and loss of function of the intestinal/colonic barrier.

New markers for sepsis caused by Pseudomonas aeruginosa during burn infection.

INTRODUCTION: Sepsis is a leading cause of mortality in burn patients. One of the major causes of sepsis in burn patients is Pseudomonas aeruginosa. We hypothesized that during dissemination from infected burn wounds and subsequent sepsis, P. aeruginosa affects the metabolome of the blood resulting in changes to specific metabolites that would serve as biomarkers for early diagnosis of sepsis caused by P. aeruginosa. OBJECTIVES: To identify specific biomarkers in the blood after sepsis caused by P. aeruginosa infection of burns. METHODS: Gas chromatography with time-of-flight mass spectrometry was used to compare the serum metabolome of mice that were thermally injured and infected with P. aeruginosa (B-I) to that of mice that were neither injured nor infected, mice that were injured but not infected, and mice that were infected but not injured. RESULTS: Serum levels of 19 metabolites were significantly increased in the B-I group compared to controls while levels of eight metabolites were significantly decreased. Thymidine, thymine, uridine, and uracil (related to pyrimidine metabolism), malate and succinate (a possible sign of imbalance in the tricarboxylic acid cycle), 5-oxoproline (related to glutamine and glutathione metabolism), and trans-4-hydroxyproline (a major component of the protein collagen) were increased. Products of amino acid metabolism were significantly decreased in the B-I group, including methionine, tyrosine, indole-3-acetate, and indole-3-propionate. CONCLUSION: In all, 26 metabolites were identified, including a unique combination of five metabolites (trans-4-hydroxyproline, 5-oxoproline, glycerol-3-galactoside, indole-3-acetate, and indole-3-propionate) that could serve as a set of biomarkers for early diagnosis of sepsis caused by P. aeruginosa in burn patients.

Effects of powdered rifampin and vancomycin solutions on biofilm production of staphylococcus aureus on orthopedic implants.

PURPOSE: Hardware infections in orthopedic surgery, specifically those involving biofilm producing bacteria, are troublesome and are highly resistant to systemic antibiotics. The purpose of this study was to demonstrate the power of rifampin and vancomycin solutions in inhibiting as well as eliminating in vitro on staphylococcus aureus (S. aureus) biofilm in vitro on stainless-steel implants. METHODS: A suspension of either S. aureus or a S. aureus containing a plasmid that cods for the green fluorescence protein containing fluorescent protein plasmid was applied to 1 × 1cm sterile stainless steel orthopedic plating material (coupon). Biofilm development was confirmed by; the quantitative assay (colony forming unit [CFU/coupon]) and visualized using confocal laser scanning microscopy. With this established method of biofilm development, we determined the minimum biofilm inhibitory concentration (MBIC) and the minimum biofilm eradication concertation (MBEC) of Rifampicin and Vancomycin. To determine the MBIC, stainless steel plates were subjected to different concentrations of antibiotic solution and inoculated with overnight cultures of S. aureus. After 24 h of incubation at 37 °C, the biofilms on the untreated and antibiotic-treated coupons were quantified. To determine the MBEC, partial S. aureus biofilms were developed on the coupons and then treated with the different concentrations of each antibiotic for 24 h. The number of bacteria within the control untreated as well as treated coupons was determined. RESULTS: Both rifampin and vancomycin solutions inhibited biofilm production of S. aureus on stainless steel mediums; the MBIC for rifampin and vancomycin were 80 ng/mL and 1 µg/mL respectively. The MBEC for Rifampicin was similar to the MBIC. However, the MBEC for Vancomycin was 6 mg/ml. CONCLUSIONS: When applied to orthopedic stainless steel hardware in vitro, solutions of rifampin and vancomycin powder separately or in combination can completely prevent and eliminate biofilm produced by S. aureus. LEVEL OF EVIDENCE: II.

Application of Lactobacillus gasseri 63 AM supernatant to Pseudomonas aeruginosa-infected wounds prevents sepsis in murine models of thermal injury and dorsal excision.

Introduction. Severely burned patients are susceptible to bacterial infection within their burn wounds, which frequently leads to sepsis, multiple organ failure and death. The opportunistic pathogen Pseudomonas aeruginosa, an organism inherently resistant to multiple antibiotics, is a common cause of sepsis in these patients.Aim. Development of a topical treatment unrelated to conventional antibiotics is essential for prevention of P. aeruginosa infection and sepsis, leading to a role for the direct application of probiotics or their by-products.Methodology. We examined the effectiveness of 20× concentrated supernatant from Lactobacillus gasseri strain 63 AM (LgCS) grown in de Man, Rogosa and Sharpe broth in inhibiting P. aeruginosa biofilms in vitro, as well as in reducing wound bioburden and P. aeruginosa sepsis in vivo.Results. LgCS inhibited the growth of P. aeruginosa strain PAO1, prevented its biofilm development and eliminated partially developed PAO1 biofilms. In the murine model of thermal injury, a single injection of LgCS following injury and PAO1 infection reduced mortality to 0 % and prevented systemic spread (sepsis). Furthermore, a second injection of LgCS 24 h after the first eliminated PAO1 from the wound. In the murine dorsal excision infection model, either LgCS or ceftazidime treatment of the PAO1-infected wound significantly reduced the mortality rate among infected mice, while combining LgCS with ceftazidime eliminated mortality.Conclusion. These results suggest the potential of LgCS in preventing sepsis from P. aeruginosa infection in severely burned and other immunocompromised patients.

Pseudomonas aeruginosa Alters Its Transcriptome Related to Carbon Metabolism and Virulence as a Possible Survival Strategy in Blood from Trauma Patients.

Trauma patients (TPs) are highly susceptible to infections, which often lead to sepsis. Among the numerous causative agents, Pseudomonas aeruginosa is especially important, as P. aeruginosa sepsis is often fatal. Understanding the mechanism of its pathogenesis in bloodstream infections is imperative; however, this mechanism has not been previously described. To examine the effect of trauma-induced changes in blood on the expression of P. aeruginosa genes, we grew strain UCBPP-PA14 (PA14) in blood samples from eight TPs and seven healthy volunteers (HVs). Compared with its growth in blood from HVs, the growth of PA14 in blood from TPs significantly altered the expression of 285 genes. Genes whose expression was significantly increased were related to carbon metabolism, especially malonate utilization and mannitol uptake, and efflux of heavy metals. Genes whose expression was significantly reduced included genes of the type VI secretion system, genes related to uptake and metabolism of amino acids, and genes related to biosynthesis and transport of the siderophores pyoverdine and pyochelin. These results suggest that during systemic infection in trauma patients, and to adapt to the trauma-induced changes in blood, P. aeruginosa adjusts positively and negatively the expression of numerous genes related to carbon metabolism and virulence, respectively. IMPORTANCE While a considerable body of knowledge regarding sepsis in trauma patients is available, the potential influence of trauma-induced changes in the blood of these patients on the pathogenesis of Pseudomonas aeruginosa is basically an unexplored area. Rather than using standard laboratory media, we grew P. aeruginosa in whole blood from either healthy volunteers or trauma patients. The specific changes in the P. aeruginosa transcriptome in response to growth in blood from trauma patients reflect the adaptation of this organism to the bloodstream environment. This knowledge is vital for understanding the strategies this pathogen uses to adapt and survive within the host during systemic infection. Such information will help researchers and clinicians to develop new approaches for treatment of sepsis caused by P. aeruginosa in trauma patients, especially in terms of recognizing the effects of specific therapies (e.g., iron, zinc, or mannitol) on the organism. Further, this information can most likely be extrapolated to all patients with P. aeruginosa septicemia.

Intrawound Vancomycin Powder Reduces Bacterial Load in Contaminated Open Fracture Model.

OBJECTIVES: To compare the effectiveness of both vancomycin powder and antibiotic bead placement to irrigation and debridement alone in prevention of infection in a contaminated open fracture model in rats. METHODS: In a previously described model of contaminated open fractures, 45 rats had simulated open fractures created, stabilized, and contaminated with Staphylococcus aureus. They were then treated 6 hours later with 3 interventions: irrigation and debridement alone (control group) or in combination with placement of polymethyl methacrylate beads containing vancomycin and tobramycin powders (antibiotic bead group) or placement of 10 mg of intrawound vancomycin powder (powder group). Rats were allowed to recover and then killed 14 days later for harvest of femurs and plates. Femurs and plates were both incubated overnight, and bacterial colonies were counted in each group for comparison. RESULTS: Quantitative counts of bacteria in bone showed significantly reduced growth in both bead and powder groups when compared with control group (P < 0.0001). Quantitative counts of bacteria in plates showed significantly reduced growth in both bead and powder groups when compared with control group (P < 0.0003; 0.029). No significant differences were seen in bacterial growth between bead and powder groups for either bones (P = 0.13) or plates (P = 0.065). CONCLUSIONS: When compared with irrigation and debridement alone, placement of intrawound vancomycin powder significantly decreased bacterial load in a contaminated open fracture model in rats similar to placing antibiotic beads. This may provide an additional adjuvant treatment that does not require a secondary surgery for bead removal.